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Advancing Healthcare
Through Personalized
Medicine
Priya Hays
Second Edition
123
Advancing Healthcare Through Personalized
Medicine
Priya Hays
Advancing Healthcare
Through Personalized
Medicine
Second Edition
Priya Hays
Personalis, Inc
San Mateo
USA
ISBN 978-3-030-80099-4 ISBN 978-3-030-80100-7 (eBook)

https://doi.org/10.1007/978-3-030-80100-7
© The Editor(s) (if applicable) and The Author(s), under exclusive license to Springer Nature
Switzerland AG 2021
This work is subject to copyright. All rights are solely and exclusively licensed by the Publisher, whether
the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of
illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and trans-
mission or information storage and retrieval, electronic adaptation, computer software, or by similar or
dissimilar methodology now known or hereafter developed.
The use of general descriptive names, registered names, trademarks, service marks, etc. in this publica-
tion does not imply, even in the absence of a specific statement, that such names are exempt from the
relevant protective laws and regulations and therefore free for general use.
The publisher, the authors and the editors are safe to assume that the advice and information in this book
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editors give a warranty, expressed or implied, with respect to the material contained herein or for any
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The registered company address is: Gewerbestrasse 11, 6330 Cham, Switzerland
Come to me whole: with
your flaws, your scars and
everything you consider
Imperfect.
Then let me show you what I
see. I see galaxies in your
eyes and fire in your hair.
I see journeys in your palms
and adventure waiting in your
smile. I see what you cannot:
you are absolutely, maddeningly,
Irrevocably perfect.---
Ariana Reines, Come To Me Whole
To Sherrill Hays, the dearest mother-in-law
that any daughter could wish for.
To the American Society of Clinical
Oncology, the American College of Medical
Genetics and Genomics, and the Society for
Immunotherapy of Cancer for granting their
elections to me.
Medical societies with character:
Great honors to be bestowed upon
high standards to live by
Preface to the Second Edition
The second edition of this book has benefitted appreciably from the recent advances
in precision medicine. Oncology is expanding to include cancer immunotherapies
and newer targeted agents with significant clinical outcomes; personalized medicine
technologies are having immediate impact in the development of wearable devices
that can monitor heart rate and other physiological parameters; and the implementa-
tion of artificial intelligence, digital pathology, and big data in personalized health-
care is ongoing. The storage of patient data is being revolutionized by electronic
health records, which, along with decision support tools, are allowing patients and
providers to have immediate access to genomic data; however, data privacy con-
cerns remain. Precision medicine clinical trials are presenting data that has signifi-
cant implications for the development of immune checkpoint inhibitors and adoptive
cell therapies. This book presents precision medicine trials as a snapshot in time and
leads the reader to “keep up” with subsequent data in their therapeutic area of inter-
est. Novel submissions to the FDA on precision medicine n-of-1 patients are also
changing the regulatory landscape considerably. Ethical considerations are still
paramount, especially concerning health disparities, data sharing, and the discovery
of variants of uncertain significance uncovered by sequencing. Financial toxicity is
a recurrent theme in the health outcomes and quality of life for precision medicines,
as they remain costly, but also demonstrate cost effectiveness.
This book is tailored for physicians and physician/scientists, and other members
of the medical community, who strive tirelessly in the interests of their patients’
health. Scientists who are invested in personalized medicine research also constitute
a primary audience. I consider this book as analogously performed in their efforts
towards advancing healthcare through personalized medicine.
San Mateo, CA, USA Priya Hays

March 2021
ix
Acknowledgments
This book would not have been possible without the outstanding resources offered
by the American Society of Clinical Oncology (ASCO), the American College of
Medical Genetics and Genomics (ACMG), and the Society for Immunotherapy of
Cancer (SITC). I am a member of all three medical societies and am a beneficiary
of their communication to their members, which provides cutting-edge, relevant,
and informative material on topics such as recent clinical trials evaluating precision
oncology regimens, translational genomic medicine, and new FDA breakthroughs
in cancer immunotherapies. This book is dedicated to these organizations, and I
remain indebted to them for their interactions with members. I also need thank
Cambridge HealthTech for providing access to their updated information on corpo-
rate involvement in personalized medicine, and Academia.edu, of which I am also a
member, for providing articles on the ethical, legal, and social implications of preci-
sion medicine.
I also thank my family and friends for their support while I was crafting the sec-
ond edition, and give a special thanks to Sherrill Hays, my mother-in-law, for her
guidance and encouragement. Finally, an acknowledgment goes out to the superb
editorial team at Springer Nature with whom I worked, and who provided steadfast
support to me throughout the course of this project: Sydney Keen, Eugenia Judson,
and Richard Lansing.
xi
Contents
1 “Introduction: Biomedical Innovation and Policy

in the Twenty-First Century”�� 1
Liquid Biopsy�� 23
Challenges and Incentives for Implementation�� 28
Components of the Book �� 30
References�� 33
2 “The Rise of Genomics and Personalized Medicine” �� 37
Human Genome Project, Sequencing, and Genomic Medicine�� 37
New Innovative Studies in NGS�� 41
Sanger Sequencing and Comparison with NGS�� 43
Single-Cell RNA Sequencing�� 43
Single-Cell Epigenetic Analysis�� 43
Application of Single-Cell Analysis in Precision Cancer Therapy�� 44
Genomic Medicine�� 44
Genomic Sequencing for Assessment of Disease�� 45
Potential Challenges of Genomic Medicine�� 47
ENCODE: Mapping the Functional Genome�� 48
Pharmacogenetics: Pharmacogenomics �� 48
Thiopurine-Methyltransferase (TPMT) �� 51
CYP2D6�� 54
CYP2C9 �� 55
UDP-Glucoronyltransferase�� 55
HIV and Abacavir�� 57
Targeted Panel Sequencing�� 59
Genomics-Enabled Medicine for Cancer Prognosis�� 62
The Emergence of Personalized Medicine�� 64
Biomarker Discovery �� 66
Tumor Mutational Burden (TMB) as a Biomarker�� 71
Germline Considerations �� 78
Conclusion �� 79
References�� 80
xiii
xiv Contents
3 “Patient Narratives: Personalized Medicine in the Field” �� 83

The Population of 1 Trial: The Story of Milasen�� 83
The Narratives of Personalized Medicine�� 86
Social Media’s Avenues for Personalized Medicine�� 91
Social Media for Patient Engagement�� 92
Role of Twitter in Identifying Barriers to Care Among
Patients with Metastatic Breast Cancer�� 94
The Narrative of Xin Zheng: A Lung Cancer Success Story �� 94
References�� 96
4 Alliances: Knowledge Infrastructures, and the Digitization
of Precision Health �� 99
Precision Medicine: Its Potential Development and Implementation�� 99
Implications of Precision Medicine for Patient Care and Disease
Treatment �� 103
Monitoring the Personal Genome�� 105
Macroeconomic Considerations of Precision Medicine�� 106
Global Alliance for Genomics and Health �� 109
Updates to Global Alliance for Genomics and Health:
GA4GH 2020 �� 112
Systems Thinking and -Omics Technologies�� 115
Data Portals and Data Analysis�� 121
Bridging the Gap for Translational Genomic Research�� 124
Database Efforts�� 125
Genomics and Artificial Intelligence �� 126
Deep Learning Methods in Precision Medicine�� 127
Digital Precision Health: The History of Machine Learning
and the Power of Big Data for Clinical Medicine�� 129
Digital Pathology �� 135
References�� 138
5 “Great Strides in Precision Medicine: Personalized Oncology,
Immunotherapies, and Molecular Diagnostics”�� 141
An Immunotherapy Case Report �� 141
A Target Therapy Case Report�� 142
The Basis for Targeted Therapies: Acquired Mutations
and Biomarkers in Cancers�� 145
Oncogenesis�� 149
Implications of Tumor Heterogeneity�� 150
Enrichment and Adaptive Strategies�� 151
Molecular Abnormalities in Solid Tumors�� 154
Germline Molecular Testing for Hereditary Cancer Risk�� 155
Cell Cycle Checkpoints�� 159
Understanding Clinical Trials in the Era of Precision
Oncology: Data, Biomarkers, and Diagnostic Testing�� 161
Contents xv
Analysis�� 162
Action�� 162
N of 1 Trial�� 165
Tumor Subtypes �� 165
Breast Cancer �� 165
Incidence and Mortality�� 165
Inherited Breast Cancer�� 166
Targeted Therapy Options in Breast Cancer Care�� 167
Tyrosine Kinase Inhibitor: Tucatinib �� 169
HER2+ /HER2-Positive Breast Cancer�� 169
Triple-Negative Breast Cancer (TNBC) and Metastatic
Breast Cancer (mBC)�� 183
Biomarker Predicting Response to PARP Inhibitors �� 183
“Study Supports Using MRD as a Stratification
Variable for Patients with TNBC” �� 186
“Unexpected Survival Benefit Reported With Trilaciclib
in Metastatic Triple-Negative Breast Cancer” �� 187
Olaparib�� 188
Talazoparib�� 189
Immunotherapy and Immunotherapy Combinations �� 190
Newer Generation Targeted Therapy Agents�� 195
Other Combination Agents �� 197
Recent Immunotherapies�� 198
Avelumab�� 200
Liquid Biopsy in Breast Cancer: A Systematic Review �� 200
Current Breast Cancer Diagnosis Standards�� 201
Oncotype DX �� 202
MammaPrint�� 202
PAM50 (Prosigna)�� 203
Genomic Grade Index (GGI) (Ipsogen) �� 204
Veridex Rotterdam Signature 76 Genes �� 204
Ventana HER2 Dual ISH DNA Probe Cocktail Assay
for Detection of HER2 �� 204
RSClin TOOL: 21-Gene Recurrence Score (RS)�� 205
HER2DX�� 205
Signatera MRD Test�� 206
Findings for Protein-Truncating and Missense Variants�� 207
“Study Suggests 86-SNV Be Incorporated in Breast Cancer
Risk Prediction Models”�� 208
Colorectal Cancer �� 210
Colorectal Cancer Classification Systems �� 211
Molecular Pathology and Diagnosis of Colorectal Cancer�� 211
“Molecular Testing in Metastatic Colorectal Cancer:
Understanding How, When, and What to Profile” �� 211
xvi Contents
Inherited CRC Syndrome�� 212

Hereditary Nonpolyposis Colorectal Carcinoma
(Lynch Syndrome) �� 213
Familial Adenomatous Polyposis�� 214
De Novo Colorectal Cancer �� 214
BRAF V600E �� 216
Additional CRC Biomarkers and Mutations�� 216
Targeted and Immunotherapies�� 217
Bevacizumab�� 217
Cetuximab�� 218
Panitumumab �� 219
Ramucirumab�� 220
Ziv-aflibercept�� 220
ASPECCT Trial �� 220
Immunotherapies for CRC: A Recent Development�� 221
Immunotherapy for the Nonimmunogenic Tumors �� 222
Liquid Biopsy �� 222
Melanoma �� 223
Targeted Therapies �� 224
BRAF-V600E Inhibitors�� 229
Immunotherapies for Melanoma�� 229
Multicenter Double-Blind Phase III IMMUNED Trial�� 231
The Phase III Double-Blind EORTC 1325/KEYNOTE-054
Trial (continued)�� 234
Immunotherapies in Combination with Targeted Agents �� 235
Adoptive Cell Therapy for Metastatic Melanoma�� 237
“Pretreatment ctDNA May Help Predict Outcomes
With First-Line—but Not Second-Line—Immunotherapy
for Patients With Melanoma”�� 238
Management of Brain Metastases from cuM�� 239
Lung Cancer�� 241
Targeted Therapies�� 243
“Pyrotinib for Pretreated Patients with HER2-Mutated
Advanced NSCLC”�� 244
Erlotinib �� 244
Gefitinib �� 245
Afatinib�� 246
Capmatinib�� 247
Dacomitinib�� 249
Summary of Erlotinib, Gefitinib, and Afatinib�� 249
Sotorasib�� 249
Contents xvii
Ceritinib �� 250

Alectinib�� 251
Brigatinib �� 251
Lorlatinib�� 251
Selpercatinib�� 253
Pralsetinib �� 254
Newer Targeted Therapy Agents�� 256
Immunotherapies�� 263
Addition of Nivolumab to Neoadjuvant Chemotherapy
in Resectable Stage IIIA NSCLC�� 265
KEYNOTE-024 �� 267
KEYNOTE-042 �� 267
KEYNOTE-189 �� 267
KEYNOTE-407 �� 267
KEYNOTE Trials Adverse Events�� 267
IMpower Trials�� 268
IMpower 110�� 268
Durvalumab�� 272
Comparative Analysis of Pembrolizumab or Nivolumab
in NSCLC�� 272
The ORIENT-11 Trial Showed Sintilimab Plus
Chemotherapy Improves PFS in Advanced NSCLC �� 273
The PD-L1 Status Dilemma�� 274
CheckMate Trials �� 275
CheckMate 227�� 275
CheckMate 9LA�� 276
Molecular Diagnosis�� 276
Molecular Genotyping in NSCLC �� 277
Role of Plasma-Based Molecular Genotyping in NSCLC�� 278
Case Studies of Liquid Biopsy in Lung Cancer�� 280
Prostate Cancer �� 281
Newer Generation Targeted Therapies�� 286
“Pan-BET Bromodomain Inhibitor Plus Enzalutamide
in Advanced Prostate Cancer”�� 289
Rucaparib �� 290
Non-metastatic Prostate Cancer�� 291
PROSPER�� 291
SPARTAN�� 294
ARAMIS�� 294
xviii Contents
Summary of PROSPER, SPARTAN, and ARAMIS�� 295

Immunotherapies�� 295
Molecular Diagnosis�� 296
Genetic Screening�� 297
Which Men Should Be Evaluated for Prostate Cancer?�� 298
Key Genomic Differences Identified in the Prostate
Cancer of African-American Men�� 299
“Series of Studies Identify Genetic Risk Factors
for Prostate Cancer”�� 299
“17q22 Loss Associated With Survival
in Enzalutamide-Resistant mCRPC” �� 301
Racial Differences in Genomic Profiles of Primary
and Metastatic Prostate Cancer�� 301
“Study Finds Differences in BRCA1/2 Male Pathogenic
Variant Carriers”�� 302
Diagnostic Tests �� 303
4 k Score �� 303
Prostate Health Index (PHI)�� 303
Prostate Cancer Antigen 3 Assay (PCA3)�� 304
SelectMDx �� 304
ExoDx Prostate Intelliscore (EPI)�� 304
ConfirmMDx�� 304
Prognostic Tests �� 305
Decipher (GC)�� 305
Post-operative Radiation Therapy Outcomes Score �� 305
Promark�� 305
Prolaris�� 305
OncotypeDx Genomic Prostate Score (GPS) �� 306
Precision Imaging �� 306
Biomarkers and Targeted Therapies�� 309
Tumor Immunology�� 310
Subset of Glioblastoma Tumors Identified as Immunotherapy
Candidate �� 312
Meningiomas�� 312
Hemangioblastomas�� 312
Hematological Malignancies�� 313
Hematologic Oncology Highlights 2019–2020
Almanac�� 314
Chronic Myeloid Leukemia and Acute Lymphocytic
Leukemia�� 316
Contents xix
Study Finds Asciminib Safer and More Effective

than TKI Bosutinib in Chronic Phase CML�� 317
“Effects of Stopping Tyrosine Kinase Inhibitor Treatment
for Chronic Myeloid Leukemia:�� 318
Focus on Molecular Response and Patient-Reported Outcomes” �� 318
“First-Line Dasatinib Plus Blinatumomab for Adult
Patients With Philadelphia Chromosome–Positive Acute
Lymphoblastic Leukemia” �� 320
Clinical Trials Investigating the Efficacy and Safety
of the Novel CART Therapies UCART 19 and UCART22
in R/R B-Cell ALL�� 321
“Chemotherapy Plus Blinatumomab for Philadelphia
Chromosome–Negative B-Cell ALL”�� 322
“FDA Grants Fast Track Designation to Allogeneic
CAR T-Cell Therapy for Advanced ALL” �� 324
Lymphomas�� 325
Newer Therapeutic Approaches Improving Outcomes
in Hodgkin Lymphoma�� 326
CAR T-Cell Therapy�� 328
“ASH 2020: ctDNA May Indicate Increased Risk
of Relapse After Stem Cell Transplant in Patients With DLBCL”�� 331
“Anti-CD30 CAR T-Cell Therapy for Patients With Relapsed
or Refractory Hodgkin Lymphoma”�� 335
Acute Myeloid Leukemia (AML) and Chronic Lymphocytic
Leukemia (CLL) �� 335
“Impact of NGS on Treatment Selection in AML”�� 338
“ADMIRAL Trial: Gilteritinib Improves Overall Survival
in Relapsed or Refractory FLT3-Mutated AML”�� 339
FDA Issues New Draft Guidance for Treatment of AML�� 339
“Acalabrutinib Plus Obinutuzumab Prolongs PFS Compared
to Other Targeted Agents in CLL” �� 342
“Genetic Prognostic Factors in CLL Treated With Novel
Compounds”�� 343
Ibrutinib for the Treatment of CLL�� 344
Prognostic Model Identifies Patients with CLL
at Risk for Ibrutinib Failure �� 345
BTK Inhibitors �� 346
Zanubrutinib�� 347
ELEVATE-TN�� 347
UNITY �� 350
Head and Neck Cancer �� 351
Earlier Treatments of HNSCC �� 353
xx Contents
Immunotherapy Clinical Trials�� 353

Frontline Therapy�� 354
Biomarker Testing�� 355
Biomarkers for HNSCC�� 356
“Tumor-Infiltrating Lymphocytes Plus Pembrolizumab
Active in Metastatic HNSCC” �� 358
Other Tumor Subtypes�� 359
Urothelial Cancer and Bladder Cancer �� 360
Immunotherapies in Metastatic Urothelial Carcinoma�� 362
“Enfortumab Vedotin Improves OS in Patients with Locally
Advanced, Metastatic Urothelial Carcinoma” �� 362
“Adjuvant Atezolizumab for Patients With Muscle-Invasive
Urothelial Cancer and Presence of Postsurgical ctDNA”�� 364
“A Noninvasive Assay Outperforms Standard Cytology
in Detection of Urothelial Carcinoma”�� 365
Multiple Myeloma�� 366
Melflufen: A New Alkylating Agent�� 366
BCMA-Targeted Immunotherapy: Focus on Belantamab
Mafodotin�� 366
Bispecific T-Cell Engagers�� 367
CAR T-Cells�� 367
“Analysis of Deep Whole-Genome Sequencing in Patients
With Multiple Myeloma Identifies Superior Outcome Subgroup”�� 368
“Daratumumab Regimen Benefits Black Patients
with Newly Diagnosed Multiple Myeloma”�� 370
Gain in Chromosome 1q Should Be Considered High
Risk in Myeloma�� 371
“Cilta-Cel Induces High Response Rate
in Relapsed/Refractory Multiple Myeloma”�� 372
Intermittent Dosing Schedules of RAF/MEK Inhibitor
Have Proven to Be Effective in RAS/RAF-Mutant
Multiple Myeloma �� 375
Cervical Cancer�� 378
Camrelizumab Plus Apatinib in Previously Treated Patients
With Advanced Cervical Cancer Showed Gains
in Objective Response Rate �� 379
Thyroid Cancer �� 380
Renal Cell Carcinoma�� 381
Gastric Cancer�� 384
Hepatocellular Carcinoma�� 388
First-Line Settings�� 393
Second-Line Settings �� 393
Ovarian Cancer �� 393
Contents xxi
6 Trends in Precision Oncology and Precision Medicine 2.0�� 419

Introduction�� 419
MicroRNAs�� 420
MicroRNAs as Diagnostic Markers in Cancer�� 420
MicroRNAs as Prognostic Indicators�� 421
Glioblastoma miRNAs�� 422
Colon Cancer miRNAs�� 422
Liquid Biopsy�� 423
Use of ctDNA to Direct Therapy in Advanced Breast Cancer�� 425
ctDNA May Be Reliable Predictor of Outcomes
with First-Line Immunotherapy for Melanoma �� 426
Exosomes �� 429
Immunotherapy�� 430
Immunological Mechanisms for Tumor Evasion�� 430
Escaping the Immune Response�� 431
The Development of Immune Checkpoint Inhibitors (ICIs)�� 432
Immune Checkpoint Blockade Inhibitors: The Landscape
of Treatments from Recent Clinical Trials: Solid Tumors
and Blood Cancers �� 433
Immune Checkpoint Inhibitors for Solid Tumors:
Advanced Melanoma, NSCLC�� 433
Meta-Analyses of Quality of Life in Patients Treated
with Checkpoint Inhibitors�� 435
Immune Checkpoint Inhibitors for Hematological
Malignancies (HMs)�� 441
Adoptive T-Cell Therapies �� 446
Adoptive T-Cell Therapy: History and Development�� 446
Clinical Trials for FDA-Approved CAR-T Cell Agents
in Hematologic Malignancies�� 448
Treatment for Populations with Urgent Need
with CAR-T Therapy �� 449
Anti-CD22 CAR T-Cell Therapy for CD22-Positive
B-Cell Malignancies�� 451
Immune-Related Adverse Effects Associated
with CAR T-Cell Therapy�� 457
Key Findings�� 457
Immune-Related Adverse Events for Immune
Checkpoint Inhibitors�� 459
The Intratumoral Microbiome�� 462
Systems Biology�� 463
Personalized Cancer Vaccines�� 464
Autologous and Allogeneic Tumor Cell Vaccines �� 467
Dendritic Cell Vaccines�� 468
Viral Vaccines�� 469
DNA Vaccines�� 469
xxii Contents
Personalized Vaccine Approaches�� 470

Epidermal Growth Factor Receptor Variant III
as a Vaccine Target for Glioblastoma�� 470
Recombinant Poliovirus�� 470
Personalized mRNA Cancer Vaccines �� 471
7 “Personalized Medicine’s Impact on Disease”�� 481
Type II Diabetes �� 481
GWAS and Type II Diabetes�� 483
Pharmacogenomics and T2D Treatments�� 484
DPP4 Inhibitors/GLP-1 Analogs�� 486
SGLT-2 Inhibitors�� 486
Thiazolidinediones�� 486
Gene-Environment Interactions �� 487
Neurological Disease: Alzheimer’s Disease
and Parkinson’s Disease�� 488
Alzheimer’s Disease�� 488
Genetic Testing for AD�� 488
GWAS and Genetic Meta-Analysis: The Discovery
of Rare Coding Variants �� 490
Fluid Biomarkers: Cerebrospinal Fluid (CSF) and Blood-Based
Biomarkers�� 490
Prevention and Sex-Gender Differences to Personalize
Treatment of AD�� 492
Parkinson’s Disease �� 493
The Personalized Parkinson Project (PPP) �� 495
Rheumatoid Arthritis�� 496
Multiple Sclerosis�� 497
Cardiovascular Disease�� 498
Clopidogrel Pharmacogenetics�� 499
Warfarin Pharmacogenetics �� 500
CYP2C9 �� 501
VKORC1�� 501
Genetic Testing of Monogenic and Polygenic CVDs�� 502
Novel Precision Medicine Options for Cardiovascular
Disease: Noncoding RNA as Biomarkers and Therapies�� 505
Infectious Diseases: HIV, the Novel Coronavirus and Hepatitis C�� 506
HIV�� 506
Novel Coronavirus (COVID-19)�� 507
Hepatitis C �� 511
Psychiatry�� 512
Major Depressive Disorder and Bipolar Disorder �� 513
The Role of GWAS in Psychiatric Disorders�� 515
Predictors of Treatment Response�� 515
Contents xxiii
Pharmacogenetics of Schizophrenia�� 516

Antipsychotic Response and Induced Side Effects�� 516
EMRs, Smart Wearable Devices, and Machine Learning�� 517
Big Data and Its Implementation in Psychiatry �� 519
8 The Genome in the Clinic and Boardroom: Biomarkers,
Diagnosis, Treatment, and Education�� 525
Companion Diagnostics: “Omics” and the Translation
of Biomarker Discoveries into Routine Diagnostics�� 526
Emerging Personalized Medicine in Disease: Diagnostic
Tests, Biomarkers, and Targeted Treatments �� 530
Cardiovascular Disease�� 531
Hypertension�� 531
Mental Health�� 532
Thyroid Cancer�� 533
Breast Cancer �� 533
Recent Trends in Precision Medicine’s Impact on Industry�� 534
Implementation of Personalized Diagnosis and Treatment:
Educating Patients and Healthcare Providers�� 553
Genomic Education�� 553
9 A New Set of Clinical Tools for Physicians�� 559
Precision Medicine in Community Healthcare Settings�� 560
Genomic Healthcare Team�� 560
Diagnostic Odyssey �� 566
Personalized Health Care�� 572
Finding Variants of Significance in Clinics and Sequencing Center�� 573
Clinical Utility and Diagnostic Yield of Exome Sequencing
in the Pediatric and Adult Settings�� 575
The Diagnostic Value of Immunohistochemistry�� 578
10 Legislation, Reimbursement, and the Regulatory Landscape �� 583
Recent Precision Medicine Legislation �� 583
Congress Expands Clinical Trial Access for Medicaid
Beneficiaries�� 586
FDA Role in Approval of Targeted Therapies�� 586
Laboratory-Developed Tests�� 589
Analyte-Specific Reagents �� 590
Clinical Laboratory Improvement Amendments�� 591
Challenges of Medical Device Framework for NGS �� 591
FDA Pharmacogenomics Drug Labeling�� 592
xxiv Contents
Evidence for Pharmacogenomic Testing: The Case

Example of Cancer�� 596
Regulation of Genetic Tests �� 598
Coverage and Reimbursement: Case Example of Cancer�� 599
Cost Sharing in Insurance Coverage�� 603
Pharmacogenetic Tests�� 604
Coverage of ctDNA Testing �� 604
Paying for Cell-Based and Gene Therapies �� 605
11 Translational Personalized Medicine: Molecular
Profiling, Druggable Targets, and Clinical Genomic Medicine �� 609
From Clinical Genetics to Personalized Medicine�� 609
Therapeutic Approach for Haploinsufficiency Disorders�� 621
Basic Genomic Research �� 628
Clinical Outcomes�� 636
Druggable Genome�� 637
Clinical Biomarker Discovery and Somatic Mutations �� 640
Alzheimer’s Disease�� 640
Atherosclerosis and Cardiovascular Risk�� 641
Colorectal Cancer�� 641
Translational Medicine and Technology�� 642
12 The Health Economics of Personalized Precision Medicine�� 655
The Case Study of Herceptin: Reasons for a Positive Outlook �� 658
Effect on Research Agenda and Availability of Drugs�� 659
Why Health Economics for Precision Medicine�� 660
Cost-Effectiveness: Precision Versus Conventional Medicines �� 661
Cash Flows and Value Degeneration�� 667
Establishing Value for Molecular Diagnostic Tests �� 667
Payer Perspective �� 669
The Implications of the Stratification of PM Drugs�� 670
Cost-Effectiveness of PM in the Literature�� 672
The Real-World Examples of Cancer Drugs: High Prices at Play�� 673
The Value Conundrum in Cancer Care: Creating Transparent,
Generalizable Value Models for Oncology�� 677
Embracing Necessary Complexity�� 678
Personalized Medicine and Pharmacogenomics: From
Micro to Macro�� 679
The Economic Value of Pharmacogenomic Information �� 680
Contents xxv
Economics of Precision Medicine Driven-Drug

Development Pipelines�� 681
Challenges for Economic Evaluations in Precision Medicine �� 682
Gene Therapy Payment Models �� 683
Equity and Equality�� 683
13 Moral, Societal, and Ethical Issues: Claims, Consequences,
and Caveats�� 689
Traditional Medical Ethics and the New Technologies
of Genomics �� 689
The Case of Genomic Privacy�� 693
Protection of Privacy and Confidentiality�� 694
The Human Genome in the Courtroom: Patents, Litigation,
and Physician Liability�� 698
WGS: Its Risks and Benefits�� 702
Incidental Findings and Return of Results: Debates and Discussion�� 703
“Return of Sequencing Results”�� 709
Patient Preference, Right “Not to Know” �� 711
Informed Consent�� 711
The Details of Data Sharing�� 712
GWAS: Older Technology, Newer Ethical Issues�� 715
Direct-to-Consumer Testing: Fraught with Irregularities�� 715
Ethics of Epigenetics�� 716
Health Disparities in Precision Medicine�� 718
Case Study 1�� 722
PGVs Can Have Relevance for Cancer Treatment�� 722
PGVs May Have Been Previously Missed�� 722
PGVs May Not Have Been Expected�� 723
14 Conclusion�� 727
Index�� 733
“Introduction: Biomedical Innovation
and Policy in the Twenty-First Century” 1
The development of Xalkori, also known as crizotinib, a small molecular inhibitor

encoded by the ALK gene targeted to treat non-small-cell lung cancer (NSCLC),
began several years ago. A fusion between the EML4 and ALK genes was evident
in a cDNA library of lung adenocarcinoma in a patient with transformed 3T3 fibro-
blasts. Five percent of patients with NSCLC harbored the protein resulting from this
fusion on biopsy analysis [1]. After this initial finding in 2007, crizotinib was dis-
covered to be an effective targeted therapy for patients whose NSCLC tumors har-
bored the ALK gene. According to Ranade et al., 90% of 82 patients carrying this
ALK fusion gene responded to crizotinib, causing tumors to shrink or stabilize in
30% of 57% of patients treated. Within 4 years of this discovery of this preliminary
clinical data, the FDA approved crizotinib on the basis of stratified phase II and
phase III clinical trials with selective enrollment of NSCLC patients harboring this
ALK fusion protein in late-stage metastasis. In turn, a companion diagnostic was
identified that detected the ALK fusion and was codeveloped with the medication,
as Ranade et al. noted [1, 2].
When the FDA approved the cancer drug known as Gleevec in 2001 for treat-
ment against chronic myeloid leukemia (CML), the agency did so after one of the
shortest drug review processes on record. Novartis, the maker of Gleevec, also
known as imatinib mesylate, had sponsored the clinical trials behind the drug. The
FDA approved the drug within 10 weeks of reviewing three separate studies on
1000 patients. On May 10, 2001, at the press conference announcing the FDA
approval of Gleevec, Health and Human Services (HHS) secretary Tommy
Thompson declared:
“Today I have the privilege of announcing a medical breakthrough. Like most scientific
breakthroughs, this one is not sudden, nor does it stand alone. Rather, like most scientific
advancement, it is a culmination of years of work and years of investment, by many people
in many different institutions, and even in different fields of medicine. We are here to
announce one dramatic product of all those efforts. But we believe many more products will
follow, based on years of scientific groundwork. So this is the right time to acknowledge
those efforts, to recognize that our investments in research are paying off, and to praise the
© The Author(s), under exclusive license to Springer Nature Switzerland AG 2021 1

P. Hays, Advancing Healthcare Through Personalized Medicine,
https://doi.org/10.1007/978-3-030-80100-7_1
2 1 “Introduction: Biomedical Innovation and Policy in the Twenty-First Century”
teamwork that has brought us here. It’s also the right time to talk about what this can mean
for our future—a future that promises a new level of precision and power in many of our
pharmaceutical products. Today the Food and Drug Administration has approved a new
drug, Gleevec, for treatment of chronic myeloid leukemia—or CML. Let me just say that it
appears to change the odds dramatically for patients. And it does so with relatively low
occurrence of serious side effects.”
With the details of the effectiveness of Gleevec and its implications, Thompson’s
announcement quickly gained the news media’s attention. CNN cycled the story
every half hour throughout the day of the press conference. The Associated Press
wrote and updated the story several times, and the news made the front page of
newspapers nationwide. In the weeks following the announcement, extraordinary
coverage was given to Gleevec and its effects on cancer, including a cover story in
TIME magazine (May 28, 2001) and reports in the New York Times, USA Today,
and Newsweek.
Gleevec was proven to be effective not just against CML, but also against another
cancer, gastrointestinal stromal tumor (GIST). Three days after Thompson’s press
conference, during the annual meeting of the American Society of Clinical Oncology
in San Francisco, Dr. Charles Blanke announced that the so-called “leukemia pill”
had stunning results against GIST [3] (https://liferaftgroup.org/wp-content/
uploads/2012/09/May2001newsletter.pdf).
According to a website for a GIST patient advocacy group a dozen years after its
approval, Gleevec is a relatively unknown pill. Why all the attention focused on one
orange pill against two relatively rare cancers (CML affects 4500 patients annually,
while GIST is even rarer)? Although primarily addressing CML rather than GIST,
Thompson broadly answered the question at the HHS press conference: Gleevec is
targeted therapy—it kills leukemia cells while sparing normal white blood cells.
Unlike other more strenuous chemotherapy regimens, Gleevec has relatively few
side effects. Gleevec targets the signal in the cell that causes cancer, acting as a
molecular switch. Gleevec is now the prototype of cancer drugs, and cancer research
laboratories around the world are trying to mimic the effects of Gleevec on other
types of cancers.
As reported by the National Cancer Institute, most of the 4500 Americans diag-
nosed with CML each year are middle-aged or older, although some are children.
CML does not present with symptoms in its early stages, as disease progresses
slowly. Bone marrow transplantation in the initial chronic phase of the disease was
the only known treatment for CML. However, suitable donors are difficult to find,
and many patients cannot tolerate transplantation due to health concerns, and even
among those patients, the side effects can be grave or even lethal. Prior to the devel-
opment of Gleevec, the drug interferon-alfa was prescribed and produced remis-
sions by restoring normal blood count in a majority of patients treated with the drug.
But still the prognosis remained bleak if the drug was found to be inefficacious.
With the advent of Gleevec, there have been higher remission rates in three short-
duration, early-phase clinical trials. In the results of one clinical trial, reported in
April 2001 in the New England Journal of Medicine, Gleevec restored normal blood
counts in 53 out of 54 interferon-resistant CML patients, a response rate rarely seen
1 “Introduction: Biomedical Innovation and Policy in the Twenty-First Century” 3
in cancer with a single agent. Fifty-one of these patients were still doing well after a
year on the medicine, and most reported few minor side effects. Imatinib mesylate
was invented in the late 1990s by scientists at Ciba-Geigy (which merged with
Sandoz in 1996 to become Novartis), in a team led by biochemist Nicholas Lydon,
and its use to treat CML was driven by oncologist Brian Druker of Oregon Health &
Science University. Druker led the clinical trials confirming its efficacy in CML [4].
The scientific story of Gleevec, which became known as targeted therapy, a med-
ical breakthrough that was a result of years of research, heralds back to the discov-
ery of the BCR-ABL fusion gene from translocation occurring between chromosomes
9 and 22, the Philadelphia chromosome or also known as the Philadelphia transloca-
tion (Ph). In 1959, the Philadelphia chromosome was first discovered and described
by David Hungerford from Fox Chase Cancer Center (then the Institute for Cancer
Research) and Peter Nowell from the University of Pennsylvania School of
Medicine, and was named after the city in which both facilities are located
(Correction from First Edition). Ph is a specific genetic abnormality present in chro-
mosome 22 of CML cells. An unusually short, defective chromosome results from
the reciprocal translocation between genetic material between chromosomes 9 and
22 and a fusion gene BCR-ABL1 results. The ABL1 gene of chromosome 9 is jux-
taposed onto the breakpoint cluster region BCR gene of chromosome 22, coding for
a hybrid protein: a tyrosine kinase signaling protein that is constitutively on, leading
to uncontrolled cell division and impairment of signaling pathways, a hallmark of
cancer. CML diagnosis is predicated on the observation of this translocation, and
the BCR-ABL gene is present in all cases of CML. The presence of this transloca-
tion is required for diagnosis of CML; in other words, all cases of CML are positive
for BCR-ABL [5].
Gleevec is targeted therapy, designed to attack cells with this BCR-ABL translo-
cation. “For the first time, cancer researchers now have the necessary tools to probe
the molecular anatomy of tumor cells in search of cancer-causing proteins,” said
Richard Klausner of the National Cancer Institute. “Gleevec offers proof that
molecular targeting works in treating cancer, provided that the target is correctly
chosen. The challenge now is to find these targets.”
Recent studies have continued to show successful outcomes for Gleevec. Three
years of adjuvant treatment with Gleevec can prevent almost 50% of deaths in high-
risk GIST patients who underwent surgery in the first decade, according to long-
term data from the Scandinavian Sarcoma Group XVIII/German trial. In a 10-year
follow-up data presented at ASCO20, 400 patients with resectable GIST and high
risk for recurrence were randomly assigned to imatinib dosing for 3 years [6].
Efficacy as determined by “recurrence-free survival (RFS) and overall survival (OS)
efficacy for three years of adjuvant imatinib is highly superior as compared to 1 year
of imatinib.” The intention-to-treat cohort had a 10-year OS of 79% for 3 years of
imatinib versus 65% in the 1-year group (HR 0.55, 95% CI [0.37, 0.83]; p = 0.004)
[6]. Additionally, at 119 months follow-up, RFS and OS remained significantly bet-
ter in patients assigned to 3 years of imatinib compared with 1 year of imatinib.
Comparably, the intention-to-treat cohort had a 10-year RFS of 53% for 3 years
versus 42% for 1 year of imatinib (HR 0.66, 95% CI [0.49, 0.87]; p = 0.003) [6].
There are hundreds of known mutations for cystic fibrosis (CF), an inherited
disease that affects the lung leading to complications such as pneumothorax. In
2012, the FDA approved a new therapy for CF called Kalydeco (known generically
as ivacaftor), which was approved for patients with a specific genetic mutation—
referred to as the G551D mutation—in a gene that is important for regulating the
transport of salt and water in the body. Approximately 4% cases are positive for the
G551D mutation. G551D is responsible for only 4% of cases in the United States
(approximately 1200 people). In these patients, the mechanism of action of Kalydeco
prevents the buildup of the sticky mucus on the surface of the lungs and can lead to
improper flow of salt and water, and is a precursor to life-threatening lung infections
such as pneumothorax. Kalydeco helps to restore the function of the protein of the
mutated gene that can lead to life-threatening lung infections and digestive problems.
However, the development and success of Kalydeco, even though targeted for a
relatively small group of patients, led to hope that a more “commonly occurring
mutation, a three-base deletion that results in a missing phenylalanine in codon
508” (Phe508del CFTR), would have effective treatments. The results of a pair of
phase III clinical trials recently published in an editorial in the New England Journal
of Medicine documented positive clinical outcomes from triple-drug therapy for
persons with cystic fibrosis and who had at least one copy of the Phe508del CFTR
mutation. These CF populations represent approximately 90% of persons affected
by this life-shortening autosomal recessive disease. Although only approximately
5% of patients with cystic fibrosis stood to benefited from ivacaftor, their “hearten-
ing clinical response served to stimulate vigorous pursuit of drugs for more com-
mon cystic fibrosis-causing mutations, especially the Phe508del CFTR mutation.
Two phase III, multicenter clinical trials reported now document the efficacy and
safety of elexacaftor–tezacaftor–ivacaftor triple-combination therapy (two correc-
tors, one potentiator) for patients with one or two copies of the Phe508del CFTR
mutation. [The editorial enthusiastically notes that] given the recent approval of this
therapy by the FDA, these findings indicate that it may be possible to offer safe and
effective molecularly targeted therapies to 90% of persons with cystic fibrosis” [7].
The profile of trastuzumab, more commonly known as Herceptin, also illustrates
many of the same principles of personalized medicine: targeted treatment for aggres-
sive breast cancer, making cancer cells retract while not damaging normal cells. The
history of Herceptin harks back to the discovery of the HER-2 gene by Robert
Weinberg in 1979, which he found to be involved in numerous cancer pathways. The
HER-2 gene makes a HER2 protein receptor that signals cells to grow by acting as a
growth factor. Dennis Slamon, a pivotal figure in the development of Herceptin,
announced at the May 1998 meeting of American Society of Clinical Oncology
(ASCO) that he had discovered a novel antibody therapy against the HER-2 protein,
Herceptin. The mechanism of action of Herceptin is that it attaches to HER2 recep-
tors on the surface of breast cancer cells and prevents the cells from receiving growth
signals. Much of what was discussed about Gleevec applies to Herceptin: it is molec-
ularly targeted therapy that influences genetic processes and turns off the actions of
mutated genes, with fewer side effects for patients. The FDA approved Herceptin for
the treatment of HER2-positive metastatic breast cancers in September 1998, and on

that same day granted approval to DAKO Corp for HercepTest, an in vitro assay to
detect HER2 protein overexpression in breast cancer cells [8].
Follow-up clinical trials of Herceptin have shown clinical efficacy of the mono-
clonal antibody. Cameron et al. reported in the journal Lancet a long-term study of
Herceptin in the adjuvant setting in the HERA (HERceptin Adjuvant) trial. In this
longitudinal study, HER2-positive early breast cancer patients were enrolled with a
median follow-up of 11 years [9]. Random assignment to 1 year of trastuzumab
significantly decreased the risk of a disease-free survival event (HR 0.76, 95% CI
0.68–0.86) and death (0.74, 0.64–0.86) compared with the observational cohort.
The clinical trial showed promising results in this longitudinal study as estimates of
10-year disease-free survival were 63% for observation, 69% for 1 year of trastu-
zumab, and 69% for 2 years of trastuzumab [9].
However, discussing the full impact of personalized medicine drugs cannot be
complete without mentioning Keytruda, or commonly known as pembrolizumab.
According to Biospace.com, Keytruda, a cancer immunotherapy, has been approved
for 21 indications, including some of the most prevalent tumor types, melanoma,
NSCLC, head and neck cancer, and classical Hodgkin Lymphoma. According to
Terry, “there are currently more than 1,000 clinical trials ongoing involving Keytruda
in a variety of cancers and treatment settings. The drug brought in $2.27 billion
alone in the first quarter of 2019 for Merck, hitting $2.63 billion for the second
quarter, exceeding Wall Street projections of about $2.5 billion” [10].
Keytruda is in a class of immune checkpoint inhibitor drugs that allows for the
immune system to attack tumor cells by preventing the evasion of tumor cells from
immune system attack. PD-L1 is a cell surface protein on T cells, the cellular immu-
nity of the immune system, which enables tumor cells to bypass the immune sys-
tem. Keytruda prevents the functioning of PD-L1 by binding to PD-1.
Keytruda received its FDA approvals through a series of KEYNOTE clinical tri-
als. Parker et al. conducted a meta-analysis of 12 KEYNOTE trials and compared
overall survival. The trials differentiated between six cancers, with a specific
Keytruda arm, chemotherapy arm, and Keytruda + chemotherapy arm [11]. Patients
were followed up at approximately 13 months. Mean overall survival was
12.5 months in the Keytruda arm versus 11.1 months for all control arms (HR 0.71,
95% CI: 0.59–0.85, p < 0.001, I2 = 0%). These results were observed across all set-
tings, in all the cancer types, advanced refractory or chemo-naïve cancer patients
[11]. In 2017, the US Food and Drug Administration (FDA) approved pembroli-
zumab for any tumor with deficiency in mismatch repair, regardless of histology.
Five stories, five drugs. Each of these highlights certain aspects of personalized
medicine and positive lessons learned: the discovery of driver mutations that drugs
could target, rapidly facilitated clinical trials that lessen FDA approval time for
breakthrough drugs, and co-development of drugs and companion diagnostics that
lead to effective predictive treatment for patients, and the secondary benefit of addi-
tional scientific and clinical research toward the discovery of molecularly targeted
treatments.
As the abovementioned examples indicate, by focusing on creating drugs for

patients that may not respond to conventional treatment, personalized medicine
aims to disrupt health care by treating these patients appropriately. The potential lies
ahead for lowering healthcare costs, fixing the ailing healthcare system, and provid-
ing options for preventing disease by early detection and risk prediction [12].
A notable success story of precision medicine is in the epidemiology and statis-
tics of NSCLC, the cancer originally mentioned early in this chapter, and where the
incidence and prevalence have diminished as a result of the development of targeted
therapies. In a recent article in the New England Journal of Medicine, a “study by
researchers at the National Cancer Institute investigating mortality trends in lung
cancer by subtype has found that population-level mortality from individuals with
non–small cell lung cancer (NSCLC) fell sharply from 2013 to 2016, and that sur-
vival after diagnosis improved substantially. The study findings suggest that a
reduction in incidence in conjunction with treatment advances likely explains the
decline in mortality” [13]. The study authors wrote “our analysis suggests that a
reduction in incidence along with treatment advances—particularly approvals for
and use of targeted therapies—is likely to explain the reduction in mortality observed
during this period” [13].
NSCLC Data from Surveillance, Epidemiology, and End Results database
(SEER) illustrates precision medicine in action as well by strongly suggesting that
advances in targeted therapy led to decreases in NSCLC mortality rates by a signifi-
cant percentage accompanied by increases in survival. The trends showed mortality
in men decreased 6.3% annually from 2013 to 2016, and the incidence decreased
3.1% annually from 2008 to 2016. Similar patterns were observed in women.
According to researchers, “[c]orresponding lung cancer–specific survival improved
from 26% among men with NSCLC that was diagnosed in 2001 to 35% among
those in whom the cancer was diagnosed in 2014. This improvement in survival was
found across all races and ethnic groups.” Precision oncology heralds a new future
in cancer outcomes for the better, with improved prognosis, and what has been dem-
onstrated in clinical trials is borne out by further analysis [13].
Some of the most remarkable changes have also occurred in the landscape of
clinical trials in the wake of personalized and precision medicine. By identifying
driver mutations in heterogenous tumors that could serve as targets for therapy, drug
companies could save time and money in drug development by “designing small but
highly effective trials targeted to those patients more likely to benefit from the ther-
apy” [1]. In a study led by researchers at Weill Cornell Medical College in 2015,
results “identified 684, or 8%, of eligible trials as precision cancer medical trials
that were significantly more likely to be phase II and multicenter; involved breast,
colorectal, and skin cancers; and required 38 unique genome alterations for enroll-
ment. The proportion of precision cancer medicine trials compared with the total
number of trials increased from 3% in 2006 to 16% in 2013,” and the number con-
tinues to increase until today [14].
As early as July 2015, clinical trials with 20 more arms were enrolling patients
with different cancer types that responded to a variety of molecular targets, which
reflected a classical clinical trial design to some degree [12]. Instead of focusing on
one cancer, as trials have for decades, the National Cancer Institute’s NCI-MATCH
(Molecular Analysis for Therapy Choice) trial included patients with any solid
tumor or lymphoma who have one of many genomic abnormalities known to drive
cancer, also known as a basket trial. Patients were matched with a targeted agent
that has shown promise against their abnormality, regardless of what cancer they
have. Known as a basket trial, the new design highlights the rapidly growing num-
ber of potential targets and agents in oncology and the urgency of finding more
efficient ways to evaluate them in trials [15].
According to a review published by the National Cancer Institute, “recent
advancements in cancer biomarkers and biomedical technology have begun to
transform the fundamentals of cancer therapeutics and clinical trials through inno-
vative adaptive trial designs. The goal of these studies is to learn not only if a drug
is safe and effective, but also how it is best delivered and who will derive the most
benefit” [16]. Heckman-Stoddard and Smith (2014) cite two trials in the early days
of precision medicine: I-SPY and BATTLE: The I-SPY-1 trial evaluated molecular
biomarkers of treatment and response and breast imaging to guide “adaptive” ther-
apy, that is, subsequent optimal treatments.
The I-SPY-1 trial assesses women with nonmetastatic advanced breast cancer
tested for molecular targets such as estrogen receptor, progesterone receptor, and
HER2. Mammaprint, a companion diagnostic, was utilized to predict recurrence
through analysis of a 70-gene signature [16]. The objective of I-SPY-1 was to dis-
cover molecular biomarkers associated with breast cancer and determine clinical
response. Chemotherapy was administered before surgery on a neoadjuvant basis,
and biomarkers were compared with tumor response based on magnetic resonance
imaging and 3-year disease-free survival. The results of the study were astounding
[16]. Pathologic complete response (pCR) (no invasive tumor present in breast or
adjacent lymph nodes) was distinguishable based on the subset of biomarkers tested.
HR-positive patients had the lowest pCR (9%), while HER2-positive patients had
the highest pCR (45%). The trial also determined that pCR predicted recurrence-
free survival in each molecular subset. According to the authors, the study estab-
lished the foundation for the integration of biomarkers with imaging and provided
real-time data for additional clinical trials [16].
The follow-up I-SPY 2 trial found that improved pathologic complete response
was observed in patients with high-risk HER2-negative stage II or stage III or triple
negative breast cancer on the combination regimens of the immunotherapy “dur-
valumab, the PARP inhibitor olaparib, and the taxane paclitaxel followed by doxo-
rubicin/cyclophosphamide as neoadjuvant therapy vs paclitaxel followed by
doxorubicin/cyclophosphamide alone. The study also found that immune-rich can-
cers showed higher pathologic complete response rates in all subtypes as well as in
the experimental and control arms” [17].
A 3-year follow-up study revealed that in the phase II I-SPY 2 trial, as reported
in JAMA Oncology, pathologic complete response or pCR had an association with
improved outcomes, including event-free survival and distant recurrence-free
survival regardless of neoadjuvant regimen and molecular characteristics, along

with 80% reduction in recurrence rate in women with high-risk stage II or III breast
cancer, encompassing nine novel therapeutic combinations. The 950 women cohort
included patients that had not undergone prior surgery and were not administered
prior systemic therapy with primary tumors ≥2.5 cm. The women were randomized
according to nine investigational agents that were used in combination and com-
pared to controls stratified by molecular subtype. Women with a low 70-gene assay
score that were HER2 negative and hormone-receptive positive were excluded from
the study. All patients were routinely administered neoadjuvant taxane treatment
with or without the investigational agent followed by doxorubicin and cyclophos-
phamide. A total of 34.7% or 330 women achieved pCR ranging “from 17% in
HR-positive/HER2-negative patients to 68% in HR-negative/HER2-positive
patients.” Compared to control arms, the investigational agent cohort had higher
rates. Three-year event-free survival trended toward 95% in pCR versus 78% with-
out pCR (HR = 0.19, 95% confidence interval [CI] = 0.12–0.31). “Range by molec-
ular subtypes was 93% to 97% versus 57% to 89%. By molecular subtypes, hazard
ratios were 0.14 (95% CI = 0.03–0.55) for HR-positive/HER2-negative disease,
0.15 (95% CI = 0.03–0.63) for HR-positive/HER2-positive disease, 0.18 (95% CI =
0.09–0.34) for HR-negative/HER2-negative disease, and 0.14 (95% CI = 0.05–0.41)
for HR-negative/HER2-positive disease.” For all patients, 3-year distant recurrence-
free survival was “95% in those with pathologic complete response versus 81% for
those without pathologic complete response (HR = 0.21, 95% CI = 0.13–0.34). By
molecular subtypes, hazard ratios were 0.16 (95% CI = 0.04¬–0.64) for HR-positive/
HER2-negative disease, 0.10 (95% CI = 0.01–0.77) for HR-positive/HER2-positive
disease, 0.20 (95% CI = 0.10–0.40) for HR-negative/HER2-negative disease, and
0.18 (95% CI = 0.06–0.53) for HR-negative/HER2-positive disease.”
These observations led investigators to conclude that the I-SPY 2 trial demon-
strated improved 3-year clinical outcomes for the nine novel therapeutic combina-
tions regardless of molecular subtype or treatment regimen, which led to pCR after
neoadjuvant therapy in association with a 80% reduction in recurrence rate. The
aims of the I-SPY 2 were to rapidly identify investigational agents that improve
pCR when validated in a phase III confirmatory trial. The investigators stated that,
“[w]hether pathologic complete response is a validated surrogate in the sense that a
therapy that improves pathologic complete response rate can be assumed to also
improve long-term outcome requires further study” [18].
The phase II Biomarker-integrated Approaches of Targeted Therapy for Lung
Cancer Elimination (BATTLE) program is a second example noted here of a clinical
trial to determine regimes for precision or personalized medicine with the utiliza-
tion of biomarkers to determine disease response [16].
“Biomarkers have emerged as an important factor in planning treatment for non-
small cell lung cancer (NSCLC) because of knowledge that specific epidermal
growth factor receptor (EGFR) mutations lead to improved outcomes with EGFR
tyrosine kinase inhibitors (TKI) [16]. The BATTLE program consists of an umbrella
trial plus four phase II protocols. The targets included EGFR (erlotinib), KRAS/
BRAF (sorafinib), retinoid-EGFR signaling (bexarotene and erlotinib), and
vascular endothelial growth factor receptor (VEGFR) (vantetanib) [16]. The pri-
mary endpoint of the study was the 8-week disease control rate (DCR) defined as
complete or partial response or stable disease via Response Evaluation Criteria in
Solid Tumors (RECIST)” [16].
“Patients enrolled in the umbrella trial underwent tumor biopsy and biomarker
analysis for 11 biomarkers: mutations in EGFR, KRAS, and BRAF; copy numbers
of EGFR and the Cyclin D1 gene (CCND1); and protein expression level of VEGF,
VEGF-2, RXRs α, β, and γ, and Cyclin D1. The results of the biomarker analysis
were used to classify patients into one of five groups: (1) EGFR mutation and/or
amplification; (2) KRAS or BRAF mutation; (3) VEGF and/or VEGF-2 over-
expression; (4) RXRs α, β, and γ, and/or Cyclin D1 overexpression and/or CCND1
amplification; or (5) negative for biomarker panel. During the first part of the study
patients were enrolled randomly to each of the four phase II studies except for
patients with prior erlotinib treatment, who were excluded from the erlotinib-
containing study arms. The results of this randomized portion of the study were
used to assess the association between a given marker group and disease con-
trol” [16].
Efficacy outcomes for these biomarker-driven trials have also been demonstrated
in one study published in the Journal of the National Cancer Institute led by inves-
tigators at the Moores Cancer Center at the University of California, San Diego:
In order to ascertain the impact of a biomarker-baesed (personalized) strategy, clinical out-

comes were compared between molecularly targeted treatments. The results included trials
with 58 drugs, of which 57 were randomized [32% personalized] and 55 were non-
randomized trials [47% personalized]. Analysis of experimental arms in all 112 personal-
ized medicine trials utilizing oral and single agents led to relatively higher response rates
versus non-personalized medicine trials. The study authors concluded that “a biomarker-
based approach was safe and associated with improved efficacy outcomes in FDA-approved
anticancer agents” [19].
Garralda et al. discuss new clinical trial designs in what has now come to be
known as the era of precision medicine. Traditional drug development studies with
safety and efficacy evaluations in phase I and phase II components, respectively,
and randomized phase III trials for determining treatment options are gradually fad-
ing away, being replaced by “basket and umbrella designs aimed at optimizing the
biomarker-co-development process.” Targeted agents are now matched with tumor
mutations, and these agents that are designed to interact with a specific target utiliz-
ing predictive biomarkers demonstrated high survival rates [20].
What are precision medicine-designed clinical trials? They are umbrella proto-
cols, wherein patient tumors are molecularly sub-stratified and matched to distin-
guishable anticancer agents, and basket trials, with patients that have differing
tumors but common molecular alterations and matched with the same treatment
(Fig. 1.1) [20].
I-SPY-1 has an adaptive trial design, while BATTLE is an umbrella trial. Other
umbrella trials include the ALCHEMIST (Adjuvant Lung Cancer Enrichment
Marker Identification and Sequencing Trial) trial that are planned to screen nearly
Fig. 1.1 (a) Umbrella a

trials design. (b) Basket B C A A C B
trials design. (Adapted
from Damodaran et al. Molecular Profiling
[21])
Biomarker A Biomarker B Biomarker C
A A B B C C
Treatment X Treatment Y Treatment Z

b Tumor A Tumor B Tumor C Tumor D
B A A B C C B C B C A
Molecular Profiling
Biomarker A Biomarker B Biomarker C
A A A B B B B C C C C
Treatment X Treatment Y Treatment Z
8000 patients with early stage non-small cell lung cancer to identify patients with
alterations in EGFR or ALK genes. Patients with EGFR mutations would be enrolled
on a randomized trial evaluating erlotinib, and patients with ALK fusions would be
enrolled on a randomized trial evaluating crizotinib. The trial involves genomic anal-
ysis of primary tumor and at the time of relapse along with circulating tumor DNA.
The Lung-MAP (Lung Cancer Master Protocol) will screen approximately 1000
patients with advanced squamous lung cancer who have progressed after one line of
chemotherapy to identify actionable genomic alterations using a commercial tar-
geted gene panel (Foundation Medicine, Cambridge, MA) [21]. Subsequently,
patients would be randomized to one of several phase II/III trials evaluating match-
ing targeted therapies with a control arm of standard therapy. Patients without any
genomic alterations of interest will be enrolled in the arm testing immunother-
apy [21].
The epitome of basket trials may be represented by the Basket of Baskets study.
The Basket of Baskets (BoB) study is the spearhead Program of the Cancer Core
Europe (CCE) [20]. “Antitumor activity of matched therapies is evaluated in small
CCE patient populations which are molecularly determined using a novel study
design in an international multi-center (basket) approach. The study consists of two
parts: (a) I-Profiler that provides the molecular characterization of tumors from
patients with metastatic or recurrent solid tumors using a new profiling tool that
selects the most suitable treatment for these patients; and (b) I-Basket determines
multiple cohorts based on genomically selected patients” [22].
Therapies are tested in multiple disease settings which utilize companion diag-
nostics developed in concert with biomarkers, and even circulating tumor DNA for
selecting patients for tested drugs, the objective being to increase efficacy.
Basket of Baskets study seeks to accomplish ambitious goals: to integrate the
discovery of treatments through consolidation of the networks of companies and
academic sites through the use of clinical trials, and implement this innovative
clinical research strategy to develop precision treatments for molecular targets
while utilizing diagnostic tools to determine the clinical benefit of the molecular
targets, all the while engaging in merged biomedical, translational science, and
clinical research and attempting to bridge existing gaps between “scientific discov-
ery in basic and translational research and its application in the clinical research
setting.”
These precision treatments and the outcomes of these clinical trials, or the devel-
opment of them over time, have led to awareness among sectors of the community
that personalized medicine would have a definitive impact on biomedicine. As early
as January 21, 2015, President Barack Obama unveiled the Precision Medicine
Initiative in his State of the Union to members of Congress. “I’m launching a new
nationwide Precision Medicine Initiative to bring us closer to curing diseases like
cancer and diabetes,” he stated. According to the mission statement of the Precision
Medicine Initiative, precision medicine:
“Enable[s] a new era of medicine through research, technology, and policies that empower
patients, researchers, and providers to work together toward development of individualized
treatments. The future of precision medicine will enable health care providers to tailor treat-
ment and prevention strategies to people’s unique characteristics, including their genome
sequence, microbiome composition, health history, lifestyle, and diet [23] PMI. To get
there, we need to incorporate many different types of data, from metabolomics (the chemi-
cals in the body at a certain point in time), the microbiome (the collection of microorgan-
isms in or on the body), and data about the patient collected by health care providers and the
patients themselves. Success will require that health data is portable, that it can be easily
shared between providers, researchers, and most importantly, patients and research partici-
pants” [24].
The fact sheet on the Precision Medicine Initiative reads like a primer on person-
alized medicine:
• A $215 million investment in the 2016 budget

• The development of patient-initiated research that will lead to novel biomedical
discoveries and new clinical tools and knowledge toward the development of
targeted treatment, obviating the need for “one-size-fits-all approaches”
• Treating patients according to their individual genes, lifestyle, and environments

using the innovative approaches of precision medicine that enables a better
understanding of patients’ complexities
• Using molecular approaches such as determining genetic makeup to profile a
tumor and selecting treatments that can improve survival and reduce adverse
effects [24]
Investments to launch this initiative include funding to the National Institutes of

Health (NIH), FDA, and National Cancer Institute to identify genomic drivers in
cancer. Key objectives include superior oncology treatments, the creation of a vol-
untary cohort of diverse participant data, including electronic health records,
genomic data, metabolite data and microbiomes, and environmental and lifestyle
data, and privacy protection. The NIH declares that the Precision Medicine Initiative
is one small part of precision medicine. The PMI will consist of building a cohort of
one million volunteers to facilitate building effective preventive strategies, diagnos-
tics, and treatments. Stakeholders and drivers in the PMI would like this program to
treat participants as partners rather than subjects, and look beyond the genome to
integrate other sources of knowledge about health, including diet and environment,
to develop quantitative estimates of risk for a range of diseases by integrating envi-
ronmental exposures and genetic factors. Concerning this cohort study, in a survey
of public opinion conducted in 2017, 79% agree that a cohort should be done and
54% would participate, with 83% agreeing to receive the results of the survey. One
of the concerns of this cohort study is the return of data and who should have access
to it. Participants agreed that data can be accessed broadly for research purposes.
Francis Collins, Director of the National Institutes of Health, has set up a working
group for implementing this cohort study with a steering committee.
The “omics” era has entered.
Figures 1.2 and 1.3 illustrate personalized medicine in action. Figure 1.2 shows
precision medicine workflow: omics data are generated into clinically useful infor-
mation, these data are processed, and patients are stratified and personalized medi-
cines are developed individualized for each patient genetic/proteomic/epigenetic/
transcriptomic/metabolomic status. Figure 1.3 is a variation on this same theme,
with a diagram comparing to reactive, traditional medicine. Patients are stratified
according to clinical and sociodemographic characteristics for therapies; however,
in true individualized precision medicine, a diagnostic is utilized with the treatment,
also known as a companion diagnostic, which is based on the biomarkers present in
the patient.
Terms such as “genomics,” “epigenomics,” “glycomics,” “lipidomics,” “tran-
scriptomics,” “proteomics,” “metabolomics,” and “pharmacogenomics” have
emerged and are transforming medicine, allowing for prediction of patient response
to therapies through stratification. Omics profiling of populations on a wholescale
level can enable the detection of individual variations. “These high-throughput anal-
yses performed on different biomolecules can detect the variation in an individual’s
genetic and metabolic profiles related to certain disorders, thus affecting their
response to a specific therapy. These -omics facilities have wide applications
Genomics/epigenomics/pharmacogenomics
Patient’s sampling
(DNA)
Transcriptomics
(RNA)
Omics
technologies Proteomics
(proteins)
Personalized medicine
Disease susceptibility, Prevention, diagnosis, Metabolomics
treatment (glycomics, lipidomics, amino acids,
nucleotides)
Clinical data
generation
Detailed information about
Stratification of subjects patient’s health status
Biomarker identification
Variations in drug responses
Common genetic markers
Data analysis, integration and

interpretation
Computational and
bioinformatic tools
Fig. 1.2 The complete workflow of precision medicine Omics information is integrated into clini-
cal and data processes to stratify patients leading to personalized treatment, based on patient char-
acteristics. (Adapted from Qazi et al. [25])
Traditional Medicine Stratified Medicine Precision Medicine

Patients are grouped by: Individual patient level:
• Disease Sub-types • Genomics and Omics
• Risk Profiles • Lifestyle
• Demographics • Preferences
• Socio-economic Factors • Health History
• Clinical Features • Medical Records
• Biomarkers • Compliance
• Molecular Sub-populations • Exogenous Factors
Companion Diagnostic
(CDx) Biomarker
Therapy (mainly Rx) Therapy (mainly Rx)
Therapy (Rx + Dx = CDx)
Adverse No Benefit Patient benefit from more Each benefit from Individualized
Event Benefit groups treatment targeted patient treatment
Precision medicine research enables development and delivery of the right

patient intervention
Fig. 1.3 Precision medicine, the right treatment for the right patient at the right time, is contrasted
with reactive, trial-by-error traditional medicine, where the benefits of a medication are varied in a
patient population. In stratified medicine, therapies are targeted according to interindividual varia-
tion of patients. In personalized medicine, companion diagnostics, based on biomarker detection
in patients, are utilized with the targeted therapy to further individualize treatment. (Adapted from
Linguamatics.com)
throughout the life span in the prediction of disease risk factor, diagnosis, and pro-
gression as well as the targeted therapeutic approaches. This also provides a better
understanding of molecular and cellular mechanisms involved in human health and
disease. The integrated -omics approach in clinical practice as well as the availabil-
ity of different biomarkers have greatly facilitated the stratification of patient based
on their responses to specific treatment, in order to decide the best-suitable thera-
peutic regimens” [25].
The advent of precision medicine heralds back to the days when the first targeted
treatment approaches were just appearing on the horizon. Xalkori, Gleevec,
Kalydeco, Herceptin, and Keytruda are part of a new wave of targeted therapies and
immunotherapies. Gleevec serves as a significant example of personalized medicine
because it was one of the first therapies designed to attack cancer by confronting the
molecular causes of cancer.
These targeted treatment approaches meant a new wave of medicine called per-
sonalized medicine. Personalized medicine means many things to many people,
such as patients, clinicians, and other healthcare workers. As described earlier, per-
sonalized medicine concerns utilizing the genetic variation in individuals to tailor
therapies according to the constitution of each individual’s DNA, for which one can
identify the molecular causes of disease and genetically diagnose patients by recog-
nizing biomarkers that reveal (predisposition to) disease.
While there are many definitions of personalized/precision medicine, “its most
marked qualities are connecting the genetic and lifestyle factors of an individual to
determine their disease predispositions and prevention, how they respond to treat-
ment and cures, and having the capability of predicting these aspects of patient care.
The application of pharmacogenomics prevents adverse drug reactions, or drug tox-
icity, since patients can now be categorized as extensive, poor, and intermediate, and
fast drug metabolizers through genetic testing of alleles. Patient response to therapy
can now be predicted through genetic profiling. PM can be encapsulated as “the
right treatment for the right person at the right time.” The upshot: stratification of
populations according to their genome leads to individualized therapies [24].
Another way healthcare can be viewed through the lens of personalized medicine is
as “a profound shift in thinking from genetics as a specialist interest addressing rare
disorders to the use of genetic information in all aspects of health care” [26].
Personalized medicine is characterized as both innovation and atavistic, as both
elegant and convoluted, and as both revolutionary and static. Controversial nonethe-
less, personalized medicine is both encouraged and supported by some, and recon-
sidered and questioned by others. Consider a Journal of the American Medical
Association article arguing for reason to ask cogent questions about the implemen-
tation of personalized medicine [27]. Donna Dickenson, a noted bioethicist, has
written a monograph arguing that investment in personalized medicine may come
about at the expense of public health [28]. The argument that personalized medi-
cine, or personalized medicine’s potential, can lead to safe and efficacious cures
that, in the long term, may lead to less cost for the healthcare system, as evidenced
by the example of Gleevec and other drugs, is under debate. However, less scrutiny
is given to the socioeconomic impact of personalized medicine, that is: What are the
effects of personalized medicine on patient clinical outcomes and overall quality of

life? How will personalized medicine influence healthcare, through a confluence of
networks among the biomedical and pharmaceutical industry, government regula-
tors, and clinicians and researchers in academia and major medical centers?
The thesis behind this book, similar to that explored in the first edition of
Advancing Healthcare through Personalized Medicine, is that a paradigm shift is
taking place in biomedicine, driven by personalized medicine, pharmacogenetics
and pharmacogenomics, and genetic testing. The clinic, the pharmaceutical and bio-
tech industry, translational medicine, and basic research will undergo major trans-
formations, and a new set of moral and ethical issues will arise with these
socioeconomic changes. How will knowing that a genetic predisposition prevents
Alzheimer’s, heart disease, and cancer through medication and lifestyle changes
improve healthcare? Sparking a debate in society is necessary to confront the seri-
ous changes and deep consequences that will ensue from pursuing personalized
medicine.
Personalized medicine is partly about addressing inter-genetic variation in the
population. The current strategy in drug development and treatment is the “one-
drug-fits-all” kind, which has its limits in safety and efficacy, and possibly contrib-
utes to high costs for the healthcare system. Personalized medicine has the potential
to produce safe and effective medicines and to lower costs and risks, including
adverse drug reactions, which run rampant in the elderly population.
Pharmacogenetics and pharmacogenomics examine how genetic composition
affects both disease predisposition and response to therapy and bring the promise of
a new era of “personalized medicine”: delivery of the right drug to the right patient
at the right dose [29].
Thus, personalized medicine and pharmacogenomics represent a new era of
healthcare. Ideally, a medical provider can prescribe a unique therapy for a patient,
basing the therapy on the provider’s diagnosis. Patients can now receive unique
therapies for their diseases or conditions, and genetic tests can reveal information
about genomic makeup that allows for personalized care. Additionally, translational
medicine centers (translational medicine, also known as bench-to-bedside medi-
cine, “translates” discoveries from basic science laboratories into medicine for
patients) have formed or are forming in major medical centers across the United
States, with the intent of delivering clinical care though basic research (Fig. 1.4).
During the period 2011–2013, there was the possibility that federal funding in
the United States for scientific research would drop dramatically. Scientific grant
acceptance rates of research proposals plummeted to single digits. At the time of
writing of the first edition in 2012, biomedical science was at a crossroads. While
the federal government cut funding drastically for biomedical research and pharma-
ceutical companies were being criticized for pursuing profits over patients, some
scientists and clinicians were heralding that personalized medicine and translational
medicine were the future of research and treatment.
For some years now, federal funding for basic science research has been some-
what constricted. According to a study by [30] Dorsey et al. (2010), “federal fund-
ing for research has not kept up with inflation. Their conclusion: After a decade
But we live in a very exciting time
Cheap “omics” and other

100s of exciting new oncology drugs
diagnostic technologies
and immunotherapies with
enabling precision medicine
strong scientific rationales
1000s of non-cancer drugs that Remarkable

could be repurposed Convergence AI, Machine Learning, and data
of Events analytics coming of age
Clinical trial innovations: Favorable regulatory

e.g., master protocols, RWE evironment
Fig. 1.4 The diverse outcomes of personalized medicine. (Adapted from Newton, XCures, 2019)
(1994–2003) of doubling, the rate of increase in biomedical research funding slowed

from 2003 to 2007, and compared to the 2007 figure with an adjustment for infla-
tion, the absolute level of funding from the National Institutes of Health and indus-
try appears to have decreased by 2% in 2008”* [30].
A 2011 editorial in the San Jose Mercury News points to the concern over this
state of affairs in funding. The opinion writers, Roger Kornberg and Scott Bruder,
cite the discovery of flow cytometry as an example of a pivotal event that resulted in
part from federal funding of biomedical research, correlating the economic prosper-
ity of California and the United States as a whole with advances in research.
Kornberg and Bruder argue that investment in NIH promotes long-term growth and
prosperity in the national economy, particularly in the biotech sector, which inno-
vates at a record pace. They conclude that “our nation and region are at a critical
juncture. As we struggle to cope with an aging population, debilitating chronic con-
ditions, and deadly infectious diseases, sustained and strong NIH support will be
crucial to the health of California and the nation” [31].
This editorial echoes the sentiments of many biomedical researchers in aca-
demia, where much NIH-funded research occurs. It may in fact be the mainstream
view of the biomedical community and industry as well that support for academic
research will lead to economic prosperity and patient cures. This support would
probably translate to more funding and discoveries in commercial research, leading
to more jobs and global health improvements. In short, this editorial is exemplary of
efforts to increase federal funding of biomedical research on the basis of improved
health and job opportunities as an antidote to the economic recession (note that
unemployment was near historical highs when the editorial was published).
*
However, through the 21st Century Cures Act passed by Congress in December 2016 and signed
by President Obama, the NIH is authorized to receive $4.8 billion over 10 years, and the FDA
receives $500 million. The $6.3 billion act was sponsored by Republican Representative Fred
Upton and was voted overwhelmingly by the Senate and House of Representatives. The bill also
designates $1.8 billion in funding for then Vice President Joseph Biden’s Cancer Moonshot initia-
tive for cancer research.
There is a certain level of dissonance present. With basic science threatened by

imminent down-sizing due to cuts, and Big Pharma and biotech in marked transition
in terms of their business models, clinicians, medical researchers, and scientists are
touting a new wave of medicine that offers no less than a revolution in patient diag-
nosis and therapy. In other words, upstream and downstream of therapy are bottle-
necks, and in media res are growing and thriving branches of medicine. This
disconnect could well impede the process of innovation in biomedicine.
However, through the 21st Century Cures Act passed by Congress in December
2016 and signed by President Obama, the NIH is authorized to receive $4.8 billion
over 10 years, and the FDA receives $500 million. The $6.3 billion act was sponsored
by Republican Representative Fred Upton and was voted overwhelmingly by the
Senate and House of Representatives. The bill also designates $1.8 billion in funding
for then Vice President Joseph Biden’s Cancer Moonshot initiative for cancer research.
The latest developments in precision medicine funding trends may upend this
disconnect. The All of Us Research Program announced a series of precision medi-
cine research initiatives, which have led to new funding opportunities. A few of
them, although not an exhaustive list, are: The All of Us Research Program Genome
Centers aims to establish genome centers around the country that will generate
genomic data from bio-samples, and contribute to the program’s participants while
awarding grants to the Baylor College of Medicine, The Broad Institute, and the
Northwest Genomics Center at the University of Washington in 2018. The All of Us
Data and Research Center grant provided Vanderbilt University Medical Center,
Verily Life Sciences, and the Eli and Edyth L. Broad Institute funding to acquire,
organize, and provide secure access to datasets and provide research support for the
scientific data and analysis tools for the All of Us.
And, the All of Us Health Care Provider Organizations Regional Medical Centers
funding opportunity awarded the Marshfield Clinic Research Institute in Wisconsin,
the University of Miami Miller School of Medicine, and the University of Alabama
for engaging patients in the Research Program and collecting essential health data
and biological specimens.
One of the arguments of this book is that increased investment in personalized
medicine would counter the healthcare crisis, partly by promoting preventive medi-
cine in patient care. Ideally, if personalized medicine is implemented in health care,
this would most possibly lead to disease prevention and, over time, a decrease in
healthcare costs. This is mainly because increased investment in personalized medi-
cine would reconstruct the ailing business model of the biotechnology and pharma-
ceutical industry, decrease the costs of clinical trials and thus lower the prices of
drugs, take advantage of a growing IT infrastructure, and promote economic prosper-
ity through increased growth in related manufacturing and industrial sectors and
through the growth of medical and scientific centers devoted to the practice of per-
sonalized medicine. Furthermore, personalized medicine would lead to a more effi-
cient healthcare system, thus reducing the rate of cost inflation in health care and
leading to increased economic prosperity and more jobs. Personalized medicine will
accomplish this not only through the development of targeted therapies such as
Gleevec and personalized genetic profiles, but also through the “role of companion
diagnostics which can simplify the drug discovery process, make clinical trials more
efficient and informative and be used to individualize the therapy of patients” [32].
According to Precision Medicine Industry 2019 Global Market research report
studies, “latest precision medicine economic impact and states that the global preci-
sion medicine market was valued at USD 48,554.1 million in 2018, and is estimated
to be valued at USD 84,590.21 million in 2024, witnessing a CAGR of 9.7%.”
According to the report, “the key factors propelling the growth of this market are
increasing online collaborative forums, increasing efforts to characterize genes, and
advancements in cancer biology. “The report states that” the functional character-
ization of identified genomic mutations, coupled with comprehensive clinical data
available in electronic health records, has the potential to provide compelling evi-
dence to implicate novel disease and/or drug-associated mutations in phenotypi-
cally well-characterized patients. There have been increasing efforts made by both
the government and private players to establish databases that contain information
of characterized genes of a large population. Some countries are establishing a
national repository of genetic information to accelerate the research in precision
medicine. Methods, such as meta-analysis, transcriptome data analysis, and RNA-
seq data analysis, are employed to characterize genes, generally” [33].
At the 2013 ASCO annual meeting, Janet Woodcock of the FDA described a num-
ber of challenges of precision medicine, including scientific, policy, logistical, and
value-related options for health care. Diagnosis is the basis of therapy and personal-
ized medicine, and reliable diagnostics are needed to relay understandable informa-
tion to clinicians. Next-generation sequencing, as will be described in Chap. 2, will
generate huge amounts of data that will need to be analyzed and stored. As the number
of individual biomarkers within a companion diagnostics assay increase, so do the
complexity, cost, time, and risk of development. How do we find a balance between
finding the “best” set of biomarkers and the practical requirements of companion
diagnostic development and commercialization? One option would be to choose the
best set of biomarkers that are compatible with the development timelines, or finding
the right patients to aim for the best coverage, even if this means slowing drug devel-
opment and missing a few patients. Gene panels are now being used by diagnostic
companies, but they may miss mutations that next-generation sequencing would find.
New technologies are enabling new paradigms in the way we think about disease
management and patient care. We seek to expedite ways to safely translate these
technologies for diagnostic and therapeutic use indiscriminately. The demand from
pharma for regulated companion diagnostic tests and the complexity of the develop-
ment process have resulted in the formation of many physician–pharma partner-
ships and partnership models. The central element of these collaborations is nearly
always the same—“develop and commercialize an FDA approved companion diag-
nostic alongside the drug … (without delay!).” Although the basic framework is
very clear, there are still many related questions which are as yet unresolved.
The history of personalized medicine goes back to December 1984, when geneticists
set out to sequence the entire human genome. In April 2003, this moonshot was achieved.
A number of trends have emerged since then, which are detailed in this book.
Personalized medicine by the numbers report FDA report 155 drug labels of
genomic-guided therapy. According to Geoffrey Ginsburg, MD, PhD, of the Duke

Center for Genomic and Personalized Medicine, personalized medicine will lead to pre-
natal sequencing and newborn screening and the use of circulating fetal DNA to detect
fetal abnormalities. Ginsburg cites the proliferation of targeted therapies, including:
• Vermurafenib (ZEBORAF®) approved for BRAF V600E–positive melanoma

• Trametinib (MEKINIST®) approved for BRAF V600E- and V600K-positive
melanoma
• Ado-trastuzumab emtansine (KADCYLA®) approved for HER2-positive meta-
static breast cancer
• Currently, an FDA-approved CTC test by CELLSEARCH R system (Menarini
Silicon Biosystems) is being used to detect circulating tumor cells (CTCs) in
peripheral blood, which can longitudinally monitor oncology patients by detect-
ing CTCs. This test relies on a single marker (EpCAM+) for CTC isolation of
epithelial origin in blood and can be used in conjunction with other monitoring
agents, such as physical exam, imaging, and laboratory tests.
• The Maintrac R CTC count test measures cancer stem cells in blood and assesses
cancer aggressiveness in terms of risk of spread and relapse. The test, used every
3 months, can monitor treatment efficacy as well. Studies have demonstrated the
clinical utility of the Maintrac diagnostic in solid tumors.
There have also been gains in cancer diagnosis and prognosis, as well as for other
diseases such as cardiovascular disease:
• Oncotype DX®: 21-gene RNA signature from breast tumor, 12-gene RNA sig-
nature from colon tumor
• MammaPrint®: 70-gene RNA signature from breast tumor (FDA approved)
• BluePRint™: 80-gene RNA signature that distinguishes basal, luminal ERBB2
subgroups of breast cancer
• Pathwork® Tissue of Origin test: 2000 RNAs to classify cancer of unknown
primary (FDA approved)
• AlloMap®: 11-gene RNA signature for rejection following cardiac transplant
(FDA approved)
• Corus™ CAD: 23-gene blood RNA signature for coronary artery disease (CAD)
obviates the need for coronary angiography
• Triage® Cardiac Panel: 5-blood-protein signature for assessment of chest pain
and shortness of breath
• NanoString nCounter system enables digital quantification of multiplexed RNA
from small amounts of blood and formalin-fixed tumors
FoundationOne, a molecular diagnostics company, has a companion diagnostic

test, CDx, for patients with any type of advanced cancer. It uses comprehensive
genomic profiling to search for 324 genes for cancer-relevant mutations in tumor
DNA. According to their patient literature, FoundationOne®CDx “is a next gen-
eration sequencing vitro diagnostic device for detection of substitutions, insertion
and deletion alterations (indels), and copy number alterations (CNAs) in 324 genes
and select gene rearrangements, as well as genomic signatures including microsat-
ellite instability (MSI) and tumor mutational burden (TMB) using DNA isolated
from formalin-fixed paraffin embedded (FFPE) tumor tissue specimens. The test is
intended as a companion diagnostic to identify patients who may benefit from
treatment with the targeted therapies.” Additionally, F1CDx is intended to provide
tumor mutation profiling to be used by clinicians in accordance with professional
guidelines in oncology for patients with solid malignant neoplasms. The test
requires biopsy and identifies biomarkers, also known as genomic profiling (as
opposed to genetic testing that looks at inherited traits in a person’s genes and
certain cancer-relevant mutations that a person could get from their parents).
Recommendations for targeted therapy, immunotherapy, or a clinical trial are
potential options [34].
Additionally, FoundationOne’s Liquid CDx received expanded approval for
“PIK3CA, BRCA, and ALK alterations, for determining if a patient is a potential
candidate for treatment with alpelisib, a kinase inhibitor, rucaparib, a PARP inhibi-
tor, for the treatment of adult patients with a deleterious BRCA mutation-(germline
and/or somatic)-associated epithelial ovarian, fallopian tube, or primary peritoneal
cancer who have been treated with two or more chemotherapies or alectinib, a tyro-
sine kinase inhibitor.” Liquid CDx utilizes high-throughput sequencing for a panel
of 300 genes. Prior approvals for Liquid CDx included “detecting EGFR mutations
in lung cancer to select for EGFR-targeting tyrosine kinase inhibitors, and BRCA1/2
mutations in prostate cancer for selection of rucaparib.”
“Expanded approvals were extended for the detection of PI3KCA mutations in
HR-positive (PIK3CA is the most commonly mutated gene in HR-positive/HER2-
negative breast cancer; approximately 40% of patients with this subtype of breast
cancer have the mutation), HER-negative advanced breast cancer among men or
postmenopausal women for selection of second-line alpelisib in combination with
fulvestrant; detection of BRCA1/2 mutations in ovarian, fallopian tube, or primary
peritoneal cancer for selection of third-line rucaparib; and detection of ALK rear-
rangements in metastatic non-small cell lung cancer for selection of first-line alec-
tinib, indicated for the treatment of patients with ALK-positive mutated NSCLC as
detected by an FDA-approved test. ALK rearrangements are identified in approxi-
mately 5% of patients with NSCLC” [35].
The FoundationOne CDx test was also approved as a companion diagnostic for
identifying NTRK fusions in patients with “solid tumors eligible for treatment with
larotrectinib” approved by the FDA. Approval for larotrectinib was based on multi-
center clinical trials including LOXO-TRK-14001, SCOUT, and NAVIGATE and
was approved for adult and pediatric patients that have solid tumors that harbor the
NTRK gene fusion. Patients have metastatic cancer and poor outcomes predicted on
surgical resection, whose tumors regress and have no real treatment options. The
approval was based on retrospective studies utilizing the CDx assay on tissue sam-
ples available from the three clinical trials. According to the data “efficacy for
larotrectinib was shown to be maintained in patients with confirmed NTRK fusion-
positive results by FoundationOne CDx” [36].
Another real advance in precision medicine came with the discovery and devel-
opment of adagrasib that targets the KRAS mutant, also known as the most undrug-
gable oncogene. In the phase I/II KRYSTAL-1 trial, the efficacy of adagrasib was
tested in patients with NSCLC, colorectal cancer, and other solid tumors such as
pancreatic, endometrial, and ovarian cancers. According to the report, “all patients
in the trial had advanced cancer and had previously received standard treatment for
their disease, including chemotherapy and immunotherapy.”
Adagrasib’s mechanism of action is to irreversibly and selectively bind to the
KRAS G12C mutation in its inactive state, thereby blocking cell-signaling path-
ways and leading to apoptosis of cancer cells. It has an incidence of 14% in lung
cancers, 3–4% in colorectal cancers, and 2% of pancreatic cancer. The discovery of
the drug is all the more encouraging because the KRAS G12C mutation has a poor
prognosis with lack of response to most treatments.
While there were serious adverse events, such as nausea, fatigue and diarrhea,
and vomiting, observed in the trial, the clinical outcomes demonstrated potential
hope for this population with few options [37].
As an example, classification tools for lymphoma genetic subtypes have also
emerged for precision diagnosis.
“[A] probabilistic classification tool for genetic subtypes of diffuse large B-cell
lymphoma (DLBCL)—may offer valuable insights into individual responses to tar-
geted therapy, according to a [recent] study published in Cancer Cell.”
“Development of precision medicine approaches for DLBCL has proven difficult
due to its genetic, phenotypic, and clinical heterogeneity, a recurring theme in preci-
sion medicine, particularly in oncology. Recent genomic research has revealed the
existence of multiple genetic subtypes of DLBCL.” Probability algorithms were
developed to determine genetic aberrations in DLBCL and classify them into seven
subtypes, for example, mutations, copy-number alternations, or fusions that predict
the tumor. The algorithm also called LymphGen identified subtypes already in exis-
tence (MCD, BN2, N1) and identified four new ones (A53, ST2, EZB-MYC-
positive, and EZB-MYC-negative) in an NCI patient cohort (n = 574). The
percentage of molecular classification into one of these subtypes was significantly
higher than previous tools, leading to the potential for targeted trials and treat-
ments [38].
There are other diagnostic tests in the pipeline for lupus, breast cancer, viral
infection, pulmonary fibrosis, and Alzheimer’s disease; however, there remain chal-
lenges in developing companion diagnostics for certain biomarkers. In 2005, the
genetic etiology of age-related macular degeneration (AMD) was revealed through
genome-wide association studies: a variant complement factor H responsible for
AMD. Fast forward to today, and thousands of variants have been found for chronic
disease and complex traits, a stunning accomplishment and underpinning of dis-
ease, even if these results have been accompanied by a disappointing odds ratio of
less than 2.
Today, cancers such as lung adenocarcinoma are being diagnosed based on their
genetic alterations rather than histologic subtypes and treated with targeted thera-
pies based on their underlying somatic mutations, as evidenced by Xalkori.
Mobile health and social networks are also undergoing a transformation, with
technology playing a large role. Cell phones now have the capacity to delivery
important health information. There now exist a shear proliferation of products that
can monitor phenotypes and different patterns of disease, such as Google’s glucose
sensors and contact lenses sensing interocular pressure, and KardiaMobile, a por-
table EKG monitor. Social media will indicate health trends, such as Google Flu
Trends and Twitter Data Models. Ginsburg predicts that this summation of data
streams will constitute a better and more precise representation of health and dis-
ease states through social network data, sensor data, and predictive modeling. He
added that community hospitals need to keep up with the technology of personal-
ized medicine and incorporate its advances.
The principles behind this “omics testing” has demonstrated the potential to
detect disease through a simple blood draw. PanSeer, a blood test developed by
scientists, can detect five prevalent cancers within a 4 year time span, “stomach,
esophageal, colorectal, lung and liver—in 88 percent of patients who were already
diagnosed, with 96 percent accuracy”—through determining the methylation status
of the patients’ genomes. PanSeer was deemed accurate in detecting cancer in
asymptomatic people later diagnosed with the disease. These results, however, need
to be validated, according to the study authors in Nature Communications.
Participants had their blood drawn, and the samples were stored for a period of
7 years. Samples from people with and without cancer who went on to develop the
disease underwent testing for methylation status, that is, CpG islands. The goal of
PanSeer, according to the study authors, is the detection of cancer in people without
symptoms but may be at higher risk for cancer due to family history or age, other
factors, allowing for early detection, which could ideally take place in annual check-
ups [39].
Another experimental multicenter blood test was shown to “detect cancer in
women who had no symptoms of the disease, and detected several different kinds of
cancer,” researchers at Johns Hopkins University reported, who developed the test.
The new tool, a multi-analyte blood test called DETECT-A from Thrive Earlier
Detection Corp., relies on DNA and protein biomarkers. Radiological scans were
performed on women who had positive results from the blood test to look for
tumors. Out of the 10,000 women, ranging in ages from 65 to 75 with no history of
cancer and high adherence to standard-of-care screening, “the test turned up 26
cancers that were later confirmed with PET/CT” as reported in Science. All partici-
pants were enrolled through the Geisinger Health System. Researchers paid particu-
lar attention to ovarian cancers, which lack methodology for early detection and
have a favorable prognosis. Central nervous system cancers were excluded as they
are less likely to be detected by a blood-based screening test. This test has remark-
able implications for cancers that are usually caught after metastasis due to their
subtle symptomology, and could serve as the standard screening for cancer in com-
parison to standard of care. The study authors add that this screening test should
supplement and not replace standard of care diagnostic approaches [40].
A 31-gene expression profile test was found to be of robust prognostic value in
detecting melanoma at risk for recurrence or distant metastasis in a systematic
Liquid Biopsy 23
review in comparison with the American Joint Committee on Cancer staging. 211
patients from a novel cohort were joined with patients from previously conducted
studies for a total of 1479 patients. 5-year prospective results determined a
recurrence-free survival of close to 91%. “Regarding the 31-gene expression pro-
file, the researchers reported that this test identified American Joint Committee on
Cancer stage I to III patient subsets who were likely to recur or experience distant
metastasis. Concurrent consideration of both the 31-gene expression profile and
sentinel lymph node biopsy results yielded improvements in both the sensitivity and
negative predictive value for distant metastasis-free survival.”
“The 31-gene expression profile test demonstrated consistent sensitivity in iden-
tifying patients with melanoma at risk for recurrence or distant metastasis, accord-
ing to a study.” The study authors concluded that “[the 31-gene expression profile
test] augments current risk stratification by reclassifying patients for heightened
surveillance who were previously designated as being at low risk” [41].
Liquid Biopsy
Another highly significant development in precision medicine is the use of liquid

biopsy as alternatives to standard tissue biopsy. Liquid biopsy presents opportuni-
ties and challenges particularly in the fight against cancer. Liquid biopsy is a test
performed on a sample of blood or biofluid to capture and characterize circulating
tumor cells (CTCs), tumor-associated circulating cells, nucleic acids, proteins, and
extracellular vesicles. It is driven by the benefit of being a noninvasive method of
cancer detection, allowing for longitudinal samples, which are generally less risky
for patients [42]. “Liquid biopsies offer an alternative means of collecting represen-
tative and highly relevant information in a safe, easily repeatable, low cost manner
in the form of a simple peripheral blood draw, enabling examination of various
analytes including circulating tumor cells (CTCs), cell free DNA (cfDNA), cell free
RNA (cfRNA), and extracellular vesicles with a widening array of potential clinical
applications. Due to their easy accessibility, liquid biopsies enable monitoring of
disease course in real time to gauge response to treatments, elucidate temporal evo-
lution of genetic or cellular adaptations in response to therapies, and guide subse-
quent treatments. Hence, this approach may ultimately serve a pivotal role in
translating precision medicine principles to clinical practice” [42].
The standard of care for diagnosis of cancer is tissue biopsy. However, tissue
biopsy has shortcomings; it is invasive and expensive. Liquid biopsy, the detection
of blood biomarkers using sequencing, is coming into foreground for the diagnosis,
especially in cancer patients. Liquid biopsy enables signatures of patients’ tumors
that can be characterized and may aid in the understanding of tumor biology. Recent
advancements in liquid biopsy have recently resulted in the FDA approval test for
EGFR mutations blood for lung cancer.
First discovered in 1948 by Mandel and Metais, liquid biopsy relies on the detec-
tion of fragmented DNA or cfDNA and is correlated with cellular apoptosis and
damage. Determining the levels of cfDNA can provide information on survival of
cancer cells because as tumors grow, macrophages infiltrate the tumor site to clear
products of cellular injury. However, the growth of the tumor outpaces the capacity
of macrophagic clearing, and ctDNA is passively released into the bloodstream.
Active release can also occur, which has been observed in vitro, and can account for
between 0.01% and 90% of all DNA present in plasma.
In addition to CTCs, therapeutic response for cancer can be monitored through
circulating tumor DNA (ctDNA) and allows for early diagnosis and intervention
through cancer screening. Localized, metastatic, and refractory forms can be identi-
fied early through liquid biopsy and used to guide treatment selection.
One of the most remarkable advances in the application of liquid biopsy is in the
diagnosis and treatment of NSCLC. NSCLCs have responded to recently developed
targeted therapies for actionable mutations and immunotherapies and have demon-
strated robust clinical outcomes, which make use of liquid biopsy platforms even
more relevant since they can identify minimal residual disease and longitudinally
monitor disease. According to Levy, recent prospective clinical trials demonstrated
the clinical utility of liquid biopsy in NSCLC and “suggested that simultaneously
adding a plasma ctDNA analysis to tissue biopsy in treatment-naive patients may
enhance the chances of detecting a relevant actionable mutation.
The initial clinical utility of cfDNA in advanced NSCLC involved the identifica-
tion of resistant T790M mutations in patients harboring EGFR mutations who expe-
rienced disease progression while being treated with a first- or second-generation
tyrosine-kinase inhibitor (TKI),” who were treated with osimertinib and compared
with a chemotherapy-treated cohort.
Advanced cancer patients with ALK rearrangements, BRAFV600E mutations,
and MET exon ctDNA platforms participated in clinical trials comparing (NGS)-
based ctDNA, and tissue biopsy demonstrated that plasma-based testing could serve
as a source for accurate diagnosis. In the FLAURA trial, liquid biopsy showed the
potential for predicting cancer recurrence, identifying minimal residual disease, and
monitoring patients longitudinally. Patients were treated with tyrosine kinase inhib-
itors or immune checkpoint inhibitors. “The trial demonstrated the clinical superior-
ity of osimertinib versus first-generation TKIs in treatment-naïve patients harboring
EGFR mutations” [26]. ctDNA is also serving as an evaluative strategy of whether
to switch between TKIs and ICIs or adding ICIs to TKIs based on connections
between clearance of ctDNA and progression-free survival and improved outcomes
[36]. Based on these findings, clinical consensus may be that liquid biopsy testing
“should be considered in all patients with metastatic lung cancer for whom tissue
sampling may be infeasible or insufficient, or cost ineffective” [43].
However, “[s]ignificant technical, biological and clinical challenges remain: In
the technical realm, optimal methods for enrichment, recovery, amplification, and
analysis of rare cells or cell-free material are still being refined and analytically vali-
dated. Biologically, the significance of detected alterations is still being elucidated:
How do they compare with findings in tumor biopsies? How do they change over
time as tumors evolve in response to treatment? What bioinformatics approaches are
most suitable for addressing cellular heterogeneity and extrapolating from highly
Another random document with
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käskystä oli hän saapunut näihin häihin; sitä luonteen lujuutta, jonka
kuningas luuli hänellä olevan, tahtoi hän näyttää omaavansakin ja
hän lähestyi noita äsken vihityltä toivottaakseen vuorostansa heille
onnea. Tässä silmänräpäyksessä loi Armfelt häneen silmäyksensä,
ja Magdalena nosti hymyillen päätänsä, samalla kuin laattia
ikäänkuin kiikkui hänen allansa, ja hän olisi kaatunut kumoon, jos ei
pari pehmeätä käsivartta olisi kietounut hänen ympärillensä, ja
muuan ääni samassa kuuluvasti lausunut:
— Ruhtinatar kutsuu teitä, Magdalena!
Ja nyt tunsi hän itsensä samojen suojelevien käsivarsien

poisvetämäksi, ja sulhanen sekä morsian ja tuo iloinen hääjoukko
sekä kirkko olivat kaukana hänen takanansa.
— Se ei olekaan totta, Magdalena, teitä ei olo kukaan kutsunut;

mutta minä ajattelin… niin, minä huomasin teidän voivanne pahoin
ja…
Magdalena, joka yhä antoi taluttaa itseänsä kauemmaksi, näki

hämmästyneenä vapaaherratar Rålamb'in, joka säälien häntä
katseli, ja nojaten hänen olkapäähänsä antoi neiti Rudenschöld
muutaman sekunnin kyyneleensä vapaasti vuotaa.
Mennyttä, ijäti mennyttä oli Armfelt hänelle, tahi tuli tämän

kumminkin hänelle olla! Ah, mitä oli hänellä enään jäljellä maan
päällä, sittekun tämä oli toisen oma.
— Päätäni vaan vähän huimasi, pyörrytti,— kuiskasi Magdalena,

nähdessään ne levottomat silmäykset, joita tuo suojeleva
ystävättärensä häneen kiinnitti, — kirkossa oli kentiesi liian lämmin,
tahi kentiesi kukkakimppujen tuoksu tahi…
— Älkäämme ajatelko teidän pahoinvointinne syytä, pääasia on,
että jälleen tulette terveeksi…
— Luuletteko te kenenkään huomanneen,., oi, minä en soisi…
— Vihkimisen aikana olin minä teitä lähinnä, ja muiden huomio oli

niin kiinnitetty morsiuspariin… juokaa hiukka vettä, Magdalena, ja
sallikaa minun hautoa ohimoisiannne…
— Minä voin nyt paremmin…
— Kuten sanoin, minä luulen sangen harvan teitä katselleen ja jos

te pian voitte mennä sisään, tulee teidän poissaolonne tuskin
huomatuksikaan. Niin, mikä vaihetteleva päivä, hoijakoita ja
näytelmiä; mutta näistä puhuen teillä oli miellyttävä osa tuossa
"Lumotun metsän valloittaminen" -nimisessä näytelmässä!
Katkera tuskan vivahdus ilmestyi Magdalenan huulille. Eikö

elämäkin ollut näytelmää ja eikö seuraelämä pakoittanut kutakin
näyttelemään eri osaansa? Eikö hänenkin täytynyt hymyillä ja laskea
leikkiä, samalla kuin tuska ahdisteli hänen sydäntänsä! Mutta tulisiko
hänellä olemaan nyt voimaa näyttämään Armfelt'ille
välinpitämättömyyttä ja kylmyyttä? Hänen täytyi, — tuo suuri uhri oli
täytettävä!
Hän nousi äkkiä tuolta pitkältä, kapealta sohvalta, jolle hän

tuskansa valtaamana oli vaipunut, ja mennen kuvastimen eteen,
katseli hän itseänsä tarkoin. Niin, tuolla huulillansa olevalla hymyllä
voi hän esiytyä hänen silmäinsä edessä ja kohdata häntä.
Magdalenan kasvot olivat kalpeat ja hiukan laihahkot, hänen
silmänsä näyttivät suuremmilta ja loistavammilta — hänen
ajatustensa tuskallinen ristiriitaisuus oli jättänyt nämät jäljet
itsestänsä. Tulisiko Armfelt tätä huomaamaan ja mitä tulisi hän siitä
ajattelemaan? Voisiko hän tänä päivänä puhua Magdalenalle, ja mitä
olisi hänellä tälle sanomista? — Täten kiusasi hän itseänsä
tuskissaan alituisesti uusilla, ikävillä kysymyksillä ja antoi vaan
lyhyviä, katkonaisia vastauksia niihin kysymyksiin, joita
vapaaherratar Rålamb hänelle teki ja jotka koskivat joko sitä
hoijakkaa, jota he edellä puolenpäivää olivat olleet katsomassa tahi
sitä näytelmää, jossa he vähää ennen vihkimistä olivat olleet sekä
johon kuningas itse oli tehnyt suunnitelman ja jonka aineena oli ollut
Tasson "vapautettu Jerusalem"; siinä oli näyttelijänä ottanut osaa
kuningaskin ynnä hänen molemmat veljensä, ruhtinatar Sofia
Albertina sekä kaikki, joita, ollen heidän läheisessä seuruessaan,
voitiin pitää nuorina ja kauneina.
— Nyt olen minä jokseenkin voimissani… oi, kiitoksia kaikesta

osoittamastanne ystävyydestä! — lausui vihdoin Magdalena, joka
iloisen näköisenä kääntyi vapaaherratar Rälamb'in puoleen.
— Morsiusväki on nyt kokoontunut saliin, tulkaa, rientäkäämme

sinne!
Pian oli Magdalena ruhtinatar Sofia Albertinan lähellä, joka

ystävällisesti viittasi nuorta hovineitiään luoksensa ja nyt alkoi jutella
puoliääneen hänen kanssansa.
Mutta Magdalenan selän takana viuhdottiin ahkeraan viuhkoilla ja

kuiskaeltiin, että neiti Rudenschöld'illä oli äsken kirkossa ollut joku
taudinkohtaus.
— Se oli veikistelemistä ja huomioa herättämistä, — lausui

naurahtaen kreivitär Vrede.
— Anteeksi armollinen kreivitär, — keskeytti Axel Fersen, joka
seisoi niin lähellä, että hän oli kuullut nuot muistutukset, —
haluaisinpa nyt lausua erään vanhan sananparren.
— Mikähän se olisi? — kreivitär Vrede kumartui veikistellen

eteenpäin, jolloin kreivi Fersen hiljaa kuiskasi:
— Muita luullaan itsensä kaltaiseksi.
Tämän lausuttuansa riensi hän pois ja silmänräpäys sen jälkeen

seisoi hän Magdalenan luona.
— Neiti-hyvä — lausui tuo nuori jalosukuinen nähtävällä

ihastuksella, — sallitteko minun johtaa teitä valtiosaliin, jossa tanssi
soihtujen valossa kohta alkaa?
Magdalena katsoi häneen lempeällä hymyllä, hyvin huomaten

Armfelt'in silmäykset, jotka toistamiseen vihkimisen jälkeen olivat
häneen luodut, mutta hän ei menettänyt tuota mielensä malttia,
jonka hän oli päättänyt pitää, — hetki oli tullut, jolloin hän tahtoi
näyttää tuolle petolliselle, kuinka vähän hän tästä piti lukua ja kuinka
täydellisesti hän oli Armfelt'in unohtanut; hän valmistiihe tuohon
tuskalliseen riemuun, että hän tämän silmäin edessä sai näyttää sitä
viehätystä, jonka hän sillä hetkellä voi, ja hänen paisuvilta
rusohuuliltansa luiskahti hiljaa sanat:
— Enpä tiedä ketään, jota teitä mieluisammin ottasin johtajakseni.
— Jos rohkenisin teitä uskoa olisin onnellisin ihmisistä.
— Naisen sanojen epäileminen ei sovellu hyvin teidän tunnettuun

ritarillisuuteenne.
Neiti Rudenschöld'in ja kreivi Fersen'in sanat luikuivat helposti
heidän huuliltansa, ja tämän rinnalla liihoitteli Magdalena eteenpäin
sekä saapui valtiosaliin, jossa Armfelt ikäänkuin sattumasta meni
aivan hänen ohitsensa.
Magdalenan silmät loistivat, hänen kosteat huulensa olivat

puoleksi auki ja hänen huikaisevan valkoisella hipiällänsä oli vieno
ruso. Mitä ajatteli Kustaa Mauritz Armfelt hänestä tässä
silmänräpäyksessä? Eikö hänen silmänsä viipynyt Magdalenassa
mustasukkaisen mielikarvauden vivahduksella? Oi, jospa tämä vaan
voisi oikein hymyillä Axel Fersen'ille! Ja sitten?… oli koko maailma
vaan synkkä pimeys ja lemmen "tulkoon valkeus" ei koskaan enään
tulisi soimaan hänelle.
— Ettekö tekin mene soihtukulkuun?… Ettekö ai'o olla mukana

tanssimassa ruusua sulhasen rinnasta? — Magdalena kääntyi näillä
sanoilla kreivi Fersen'iin, joka koko illallisaterioimisen ajalla ei ollut
poistunut hänen luotansa.
— Jos sallitte, viivyn mieluummin teidän luonanne.
Vielä yksi niitä hymyilyjä, joita Magdalena tänä viimeisenä

tuntikautena niin runsaasti oli tuhlannut, valaisi taaskin hänen
kasvojansa, samalla kun hän tunsi todellista helpoitusta ovien
lentäessä auki ja kun uudet menot vetivät puoleensa kaikkien
huomion sekä saivat tuon häntä kohteliaasti palvelevan kavalieerin
silmät muutamaksi minuutiksi käännetyiksi pois Magdalenasta.
Käyttäen hyväksensä tätä tilaisuutta, vetäytyi hän kenenkään
huomaamatta erääsen akkunanloukkoon.
Eräs viehättävä keijukainen, jolla oli kreivitär Höpken'in ihanat

kasvot, oli ovien auetessa näyttäynyt niiden aukossa; astuen salin
keskelle seurasi häntä hovipojat puetut — tahi kentiesi oikeammin:
pukemattomat — villeiksi, jotka, soihdut käsissänsä, alkoivat tanssin,
minkä loputtua keijukainen alkoi laulun, jossa hän kehoitti sotaan
noita kristityitä ritareja, jotka villein tanssiessa olivat tulleet sisään
salin vastakkaiselta puolelta. Kovalla torven toitahduksella vastasivat
ritarit kristittyjen puolesta tuohon kehotukseen. Tässä
silmänräpäyksessä säpsähti Magdalena, muuan ääni tunki hänen
korviinsa, eräs tuttu, rakas ääni kuiskasi:
— Hyvästi Magdalena! Lempeni jää sinulle ikuisesti!
Magdalenan suoniin lensi ikäänkuin tuli, hänen hengähdyksensä

pysähtyi muutamaksi sekunniksi, hänen kosteat huulensa muuttuivat
polttaviksi ja hänen vajonneiden silmäluomiensa alta loisti hehkuvia
säteitä.
Kun Magdalena taaskin loi silmänsä auki olivat ritarit, villit sekä
keijukainen kadonneet ja hänen edessänsä seisoi kreivitär Piper.
— Tulkaa, tulkaa, neiti Rudenschöld, siellä tanssitaan morsiamen

päästä seppelettä!
Magdalena meni hänen mukanansa ikäänkuin unessa, ja yhä soi

hänen korvissansa: — lempeni jää sinulle ikuisesti!
KAHDESKYMMENESSEITSEMÄS
LUKU.
— Teidän majesteettinne puhuu lähestymisestä Tanskaan — lausui

Armfelt, joka päivä päivältä näytti nousevan kuninkaan suosiossa ja
nyt oli kuunnellut eräitä tämän salaisempia mietteitä; — mutta
sellainen lähestyminen, —jatkoi suosikki — on sen mukaan, mitä
olen kuullut paroni d'Albedyhl'iltä, peräti hyödytöin. Tanskan
perintöruhtinas on, lieveimmin sanottu, vähäpätöinen henkilö, siinä
maassa on viisaiden ja älykkäiden miesten puute, siellä ei ole
voimaa eikä rahaa, ja ministeristö on Venäjän orjana.
— Siitä huolimatta välittää nyt paroni d'Albedyhl minun puolestani

avioliittoa poikani ja perintöruhtinatar Lovisa Augustan välillä.
— Mutta onhan ruhtinatar vähintänsäkin kahdeksan vuotta meidän

perintöruhtinastamme vanhempi ja sitäpaitsi kihloissa erään
Augustenburgin ruhtinaan kanssa…
Kuningas hymyili. — Noh, mitäs kaikki tämä asiaan vaikuttaa?
— Minusta näyttää se mahdottomalta… onhan hänen

kuninkaallinen korkeutensa, Kustaa Adolf, vielä lapsi.
Nyt nauroi kuningas ääneensä, ja muutaman silmänräpäyksen
kuluttua yhtyi Armfelt mitä iloisimmasti tuohon nauruun sekä lausui:
— Luulenpa nyt rohkenevani käsittää…
— Gösta-hyväni, eihän minulla silloin, kun mitä innokkaammin

koen saada avioliittoa toimeen, voitolla vihollisia hankkeita. Tanska
on minua epäillyt, mutta minä toivon meidän hyvän välimme jälleen
palajavan. Niin, sen täytyy minussa nähdä ystävänsä, jos… —
kuningas keskeytti puheensa, ja hänen loistavat silmänsä kohtasivat
Armfelt'in ihmetteleviä silmäyksiä.
— Minä olen ikäänkuin pimeydessä, jota teidän majesteettinne

suurten, nerokkaiden hankkeiden salama väliin valaisee.
— Kuulehan, Gösta, muuan salainen asiamies on ollut Norjassa

tutkimassa kansan mieltä olisiko se halukas liittymään Ruotsiin, ja
tämä asiamies on tuonut minulle sangen mieluisen viestin.
— Tunnettu asia on koko Europan katsovan teihin ihmetellen, ja

norjalaisten salaisen suosion johdosta näen jo hengessä suuren
kuninkaani sotajoukon eturivissä voittajana tunkeutuvan tuohon
maahan.
— Ei aivan niinkään. Ilman syyttä on tahdo hyökätä heidän

kimppuunsa, minä en tahdo alkaa sotaa, minua miellyttää enemmin
itseni puolustajana oleminen. Sinä panet silmäsi suureksi, Gösta…
leskikuningatar Juhana Maria on minun välikappaleeni; hän pelkää
valtansa menettämistä; jos hän rupeaa luulemaan sen olevan
vaarassa samoin kuin henkensäkin… niin, että kaikenlaatuiset
vaarat häntä ympäröivät — ja hän tulee uskomaan sen, siitä minä
kyllä pidän huolen, — niin tulee hän peljästyksissänsä olemaan
halukas käyttämään tehokkaita keinoja, joihin valtansa suojelemisen
halu häntä tulee pakoittamaan. Jos ei hän onnistuisi, tulee hän
kiitollisuudella vastaanottamaan sen turvan sekä avun, jota Ruotsi
hänelle tarjoaa. Missä tapauksessa hyvänsä saamme tilaisuutta
näyttää laivastomme, Köpenhamina hämmästyy ja sitte…
— Sitte, teidän majesteettinne?
— Tulen kentiesi säätämään lakeja — Kristianborg'issa.
— Teidän majesteettina ajatuksien juoksu on kuin kotkan lento,

kun minä tahdon sitä seurata. Oi, jalo kuninkaani, minulta puuttuu
sanoja osoittaakseni teille kunnioitustani ja kiitollisuuttani.
— Kuninkaan silmät loistivat. Hänen kunnianhimosta kuohuva

verensä oli saattanut hänen aikeensa puhkeamaan sanoihin, yksi
noita rohkeita, vaihettelevia yrityksiä, jotka silmänräpäyksessä tulivat
hänen mieleensä, kentiesi yhtä helposti sieltä poistuaksensa, ja
hänen vilkkaat, avonaiset, nuoruutta osoittavat kasvonsa kääntyivät
suosikkiin, samalla kun hän mielihyvällä kuulteli tämän sanoja.
— Niin, minun täytyy saada sotaa, — kertoi Kustaa-kuningas, —

sotaa jonkun naapurini kanssa, olkoonpa se sitte Venäjän tahi
Tanskan kanssa; sisälliset olosuhteet tässä maassa ovat todellakin
sellaiset, että ne pakoiltavat minua sellaisiin hankkeisin.
Tyytymättömyys enenee enenemistänsä, ja nuot rahalliset pulat sitä
lisäävät.
— Jos meidän maassamme onkin tyytymätöin puolue, — riensi

Armfelt keskeyttämään,— on tämä vähälukuinen, sillä teidän
majesteettinne alamaiset tuntevat rakkautta jaloa sekä nerokasta
kuningastansa kohtaan.
— Tähän rakkauteen luottaminen on mieluista minun sydämelleni,
— lausui Kustaa-kuningas lempeästi ja jatkoi tuolla terävällä ja
asiaansattuvalla aistilla, joka loi hänelle ikään kuin vaistonaisen
aavistuksen tulevaisista seikoista sekä tapahtumista, — mutta ne,
joille enin olen loivannut kunnianimiä ja virkoja, ovat kuitenkin
salaisia vihamiehiäni, ja tekevät työtä minun kukistamisekseni… Ei,
ei, Gösta, älä koetakaan saada minua muuhun luuloon! Minä tiedän
ja tunnen näiden jalosukuisten ylimysten ainoastansa vartoavan
tilaisuutta, saadakseen muuttaa hallitusmuotoa, joten he jälleen
voisivat määrätä lakeja kuningasvallalle. Mutta se ei saa tapahtua,
se ei tule tapahtumaan, minä olen mies pitämään heitä aisoissa, ja
niinkauan kuin pääni on pystyssä, tulen kyllä pysymään ohjaksissa.
Kuningas vaikeni muutamaksi silmänräpäykseksi, jonka jälkeen hän
jatkoi: — Mutta ah, soisinpa asiat olevan taasen entisellään ja
ystävällisellä kannallansa! Niin, onnellinen sota tulee taas kaikki
selvittämään, tulee antamaan minulle takasin, mitä olen menettänyt,
ja minun lippuni alle tulevat taas kaikki puolueet kokoontumaan!
Mutta ensiksikin pitää minun pitämäni valtiopäiviä… säädyt ovat
kutsuttavat ko'olle, sen olen nyt päättänyt;… rahaa täytyy minun
saada, paljo rahaa!
Luottamus pettämättömään onneen kaikissa hankkeissansa oli

kuninkaan sanoissa, ja kentiesi vielä enemmän äänessä, jolla hän
ne lausui. Tämä sisällinen vakuutus levitti ikäänkuin loiston hänen
hienojuonteisille kasvoillensa, samalla kuin hänen tummat, vilkkaat
silmänsä säkenöi nerosta, tuosta miltei kiihkoisesta haaveilusta, joka
niin helposti näytti saavuttavan hänessä vallan, kun hän selitteli
jotakin asiaa, joka oli voittanut hänen täydellisen mieltymyksensä, ja
hän alkoi antaa viittauksia sukkelasti sommitelluista juonista, joiden
menoa hän niin hyvin osasi johtaa, kunnes hän vihdoinkin aseet
kädessänsä tulisi rientämään voitosta voittoon, tulisi saavuttamaan
tarkoituksensa, ja ulkonaisten sekä omassa maassakin olevien
vihollisten täytyisi taipua, heittäen pois aseensa, hänen tahtoonsa.
Harvoin oli Kustaa-kuningas tällä tavoin puhunut suosikillensa,

jonka seikkailuja rakastava mieli helposti käsitti ja seurasi häntä
hänen ajatustensa haaveilevassa lennossa, jossa hänelle ei näkynyt
mitään voittamatointa eikä mikään tarkoitus liian vaikealta saavuttaa.
Nämät tunteet elähyttivät vielä kuninkaan kasvoja, hänen vielä

kepeästi kiekahtaen kengänkorollansa kääntyessään Armfelt'iin ja
tarttuessaan hänen käsivarteensa sekä mennessänsä suorastaan
kuningattaren luokse, joka tänä iltana oli kutsunut luoksensa
suuremman hoviseuran.
Tuolla vastustamattomalla viehätysvoimalla, joka oli Kustaa-

kuninkaan käytöksessä, jutteli hän hetken kuningattaren ja Sofia
Albertinan kanssa. Sitte kääntyi hän Magdalena Rudenchöld'iin, jota
hän aina siitä asti, kuin hän oli keskustellut tämän kanssa Armfelt'in
avioliitosta, oli erinomattain ystävällisesti kohdellut. Ollen yhä yhtä
ystävällinen, lausui hän Magdalenalle kohteliaita sanoja tavalla
semmoisella, joka veti hänen puoleensa monta kateellista silmäystä,
ja siellä täällä alettiin kuiskeella keskenänsä, että neiti
Rudenschöld'in virnasteleminen jo meni kaikkein rajain yli.
Erittäinkin oli näiden ylhäisten naisten joukossa yksi, joka seurasi

Magdalenan kaikkia liikkeitä, kuitenkin enemmän tutkivilla ja
utelevilla kuin moittivilla silmäyksillä. Tämä nainen oli kreivitär Piper,
tuo miellyttävä Sohvi Fersen, ja kuninkaan istahtaessa pitkän,
vihriällä veralla peitetyn pöydän ääreen pelataksensa hiukan trente
et quarante-peliä (kolmea- ja neljääkymmentä, erästä uhkapeliä),
lähestyi kreivitär Magdalenaa ja vei hänet erääsen syrjähuoneesen,
jossa hän hymyillen veti tämän erääsen matalaan sohvaan
istumaan.
— Minulla on tuhansia terveisiä teille Magdalena, — lausui

kreivitär
Piper hymyillen, — keneltä, sen saatte itse arvata.
— Arvaamistaidossa en ole juuri etevä, — vastasi Magdalena

väistellen.
— Mutta koska olen ottanut tuodakseni teille terveiset, niin… —

Kreivitär leikitteli veitikkamaisesti viuhkallansa, silmäillen
Magdalenan viehättäviä kasvoja, jotka, puolittain poispäin
käännettyinä, ilmaisivat hämmästystä, joka pusersi punan hänen
poskillensa. — Noh, tuntuuko se nimi niin vaikealta saada
huuliltanne? — Kreivitär nojausi, lempeällä äänellä kysäisten,
Magdalenaan, joka, silmät luotuna laattiaan, vastasi:
— Terveisenne ovat Axel Fersen'iltä.
— Noh, vihdoinkin! Niin, Magdalena… ja kuulkaahan nyt: veljeni

on pyytänyt minua puhumaan teille eräästä asiasta, jonka
keskustelemisen hän sanoo teidän aina keskeyttävänne.
— Ja jos minä sen keskeytän, niin…
— Niin se tapahtuu kahdesta syystä, — lausui vilkkaasti Sohvi

Piper, — joko te ette häntä lemmi, tahi olette te liian ujo, ystäväiseni.
Heleä nauru avasi Magdalenan huulet.
— Eipä minua juuri moitita siitä, — vastasi hän tuolla vilkkaalla,

vapaalla tavalla, joka hänellä oli ja jota väliin nimitettiin
"viehättäväksi," väliin, "sopimattomaksi" tuota vilkasta hilpeyttä, joka
hänellä oli synnyinnältänsä ja jonka edessä tuo hämmästys, mikä
tämän keskustelun alussa hänet oli vallannut, nyt oli poistunut, ja
hän katseli kreivitär Piperiin kirkkailla, vilkkailla silmillänsä, lisäten: —
Teeskennellä en osaa ja, totta puhuen, sitä halveksinkin; tosin olen,
— jatkoi hän vitkallisesti, — koettanut näyttää iloiselta murheellisena
ollessani ja välinpitämättömältä, kun olin levottomana; onhan
elämässä sellaisiakin kohtia, joissa se on velvollisuuskin, mutta
halusta tahi taipumuksesta… ei, ei!
— Siis te ette lemmi Axelia?
— Minä kunnioitan hänen jaloa luonnottansa sekä pidän arvossa

hänen ritarillista käytöstänsä…
— Noh, siispä tahdon tehdä hänet onnelliseksi tämän

tunnustuksenne kertomisella… te tulette kerta oppimaan häntä
rakastamaan, Magdalena; hän on todellinen jalosukuinen, jota te
ylpeydellä voisitte nimittää puolisoksenne.
— Oi, en, en, sitä en voi, sitä en koskaan tule oppimaan!

Todellisesta ja vilpittömästä rakkaudesta voi sydän sykkiä
ainoastansa yhden kerran elämässä… olenhan kerta tuntenut tätä
autuutta… sanokaa se hänelle… ja minun lempeni on yhtä
uskollinen kuin toivotoinkin.
Kirkkaat kyyneleet kimaltelivat Magdalenan silmissä ja hellästi

puristaen piti hän kreivitär Piperin kättä omassansa.
— Te puhutte ikäänkuin mieletöin lapsi,— vastasi Sohvi Piper,

puolittain sääliväisesti puolittain loukatusta ylpeydestä, että hänen
veljensä, hänen, jolle eräs ruhtinas turhaan oli kosijana kumarrellut,
ja erään Ruotsin mainiomman jalosukuisen poika, eräs tämän maan
etevimmistä ylhäisistä tuli hyljätyksi eräältä köyhältä ja toisen
vallassa olevalta hovineidiltä, jonka ainoa etevyys ainoastansa oli
harvoin nähty kauneutensa. — Niin, Magdalena, ikäänkuin mieletöin
lapsi,— kertoi kreivitär — ja uskokaa minua, te työnnätte luotanne
tällä hetkellä todellisen onnen ja varman menestyksen; te tulette
joskus selvästi huomaamaan tämän, mutta silloin se jo on liian
myöhäistä!
Liikutettuna nojausi Magdalena sohvan patjoihin; jonkinlainen

ahdistuksen kaltainen epäröiminen oli hänet vallannut. — Entä jos
hän ehkä… entä jos hän lupaisi koettaa lempiä Axel Fersen'iä?
Tässä silmänräpäyksessä kävi Armfelt tuon pienen syrjähuoneen

läpi, jossa kreivitär Piper ja Magdalena tähän asti häiritsemättä olivat
vaihettaneet ajatuksiansa; kylmästi ja vieraasti tervehtivät hän ja
Magdalena Rudenschöld toisiansa.
— Ei, ei, en voi! — kuiskasi Magdalena puristaen vielä kerran

helleydellä kreivitär Pipetin kättä.
KAHDESKYMMENESKAHDEKSAS
LUKU.
Haga'n tuuheassa puistossa astuskeli kuningas Kaarlo-herttuan

rinnalla. Päättäen herttuan synkeästä otsasta ja tuosta vähäisestä
värisemisestä, joka väliin pudistutteli hänen käsiänsä, ei keskustelun
aihe ollut hänestä mieluisinta laatua.
— Tunnenpa jokseenkin hyvin tuon Ulfvenklou'n sekä kaikki hänen

juonensa, — lausui kuningas, joka ivallisella silmäyksellä katseli sitä
hämmästystä, jonka hänen sanansa vaikuttivat Södermanlannin
herttuassa. — Sinulla voi kentiesi hänen ennustuksissansa ja
haaveiluissansa olla jotakin mieluista; mutta foi de gentilhomme,
jopa sanotaan sinun siinä kohdassa toimittavan sangen naurettavaa
osaa, — osaa, joka loukkaa minua kuninkaana ja huolettaa minua
veljenä.
Kaarlo-herttua heitti nopean silmäyksen kuninkaasen, silmäyksen,

joka koetti tutkia, kuinka paljo kuninkaalla voisi olla tiedossa näitä
Ulfvenklou'n ennustuksia, mutta hänen silmäyksenpä luikui arasti
ohitse, kohdatessansa ainoastaan Kustaa-kuninkaan läpitunkevat ja
kirkkaat silmät ja tuon milt'ei ylenkatsetta osoittavan virnistyksen
hänen huulillansa.
— Sinun täytyy karkoittaa tuo seikkailija luotasi, — jatkoi Kustaa-
kuningas lempeällä innolla, joka ei sallinut yhtään vastustelemista,
— ja että hän sitte tulee pois maasta, siitä minä kyllä tulen huolta
pitämään. Niin kauan kuin häntä pidetään sinun suosikkinasi en
tahdo ryhtyä asiaan; sellaista menettelyä voisivat käsittää väärin…
minua koetetaan kyllä ilman sitäkin lahmata. Vetoaan siis sinuun
avosydämisesti siinä toivossa, että sinä huomaisit, mitä laatua tuo
Ulfvenklou oikeastansa on.
— Tähän asti olen huomannut hänen olevan sangen merkillisen

miehen, — vastasi herttua, ottaen vakavan ryhdin, — ja minä olon
nähnyt hänen tekevän sangen kummallisia temppuja ja seikkoja,
jotka, sen tunnustan julkisesti, ovat vaikuttaneet minussa todellista
ihmettelyä ja kummastusta, voimattani siltä käsittää toimittaneeni
tuota naurettavaa osaa, josta mainitsit. Etpä ole itsekään
halveksinut…
— Minä itse, — kertoi kuningas, vilkkaasti keskeyttäen, — olen

nähnyt näiden herrain salatieteilijäin temppuja sekä kuullellut heidän
ennustuksiansa, se on totta; mutta mitä minä silloin olen ajatellut, ja
mistä syystä se on tapahtunut… niin, yhdentekevä, — keskeytti hän
vähän ajan kuluttua, — minä pidän näitä herroja enemmän salaisina
kapinoitsijoina kuin salaisten tietojen omistajina, asiamiehinä, jotka
sanovat meille, hallitsijoille, kentiesi terveellisiä totuuksia…
jonkinlaisina rahvaan tribuneina, jotka ainoastansa tällä tiellä
rohkenevat valtaistuimen ääressä ilmaista ajatuksiansa. —
Kuninkaan tulee mikäli mahdollista tietää kaikki, ja tässäkin kohtaa
olen kokenut tunkea hämäryyden läpi. Mitä siis on ladeltu omalla
suostumuksellani, sitä olen suvainnut, mutta jos joku heistä omalla
luvallansa tuo kuuluviin varoituksia sekä neuvoja, pidän sen
loukkauksena, ja Ulfvenklou on rohjennut lähettää minulle
kirjoituksen, jossa hän varoittaa minua Maaliskuulta ja etenkin, että
minun tulee olla varoillani punaisiin puetuilta henkilöiltä. (41)
Kustaa-kuningas vaikeni. Tämä kirje oli nähtävästi tehnyt tukalan

vaikutuksen hänen vilkkaasen mieleensä, vaikutuksen, jonka
uudistatamista hän tahtoi välttää ja sai häntä haluamaan sen
alkuunpanijan kartoittamista. Muutaman silmänräpäyksen kuluttua
jatkoi kuningas:
— Nyt tiedät, miksi toivon sinun lakkaavasi seurustelemasta

Ulfvenklou'n kanssa; hän on seikkailija… minä pyydän sinua
pitämään häntä sellaisena. Mitä hän sinun kauttasi toivoo
saavuttavansa, en tiedä, mutta olenpa aivan vakuutettu hänen
vieneen sinulta kauniit rahat ja että sinä, jos tahdot pitää silmäsi
auki, jos tahdot pitää häntä sinä, jona hän on pidettävä, piankin tulet
huomaamaan sanojani totuuden.
Kustaa-kuningas tarttui veljellisellä suosiolla herttuan käsivarteen.

Tuolla sujuvalla puhetaidolla, jonka luonto hänelle niin auliisti oli
lahjoittanut, lasketteli hän pian leikkiä päivän tapahtumista, etenkin
niistä, jotka koskivat omaa hovi piiriä, ja lähestyi täten erästä
nelisnurkkaista, kuninkaallisen huvilan toisella sivustalla olevaa
nurmikkoa, jossa joukko herroja ja naisia olivat "leskisillä" tahi
muuten katselivat noita kansallisia leikkejä, jotka kuuluivat tämän
päiväiseen ohjelmaan.
— Naiset ovat vastaanväittämättä kepeät jaloiltansa, — lausui

Kustaa-kuningas. — Minusta on mieluista nähdä valkoisten
hameiden liehuvan nurmikolla… kreivitär Höpken muistuttaa
metsäkauriista.
Kaarlo-herttuan silmät kääntyivät noihin leikitteleviin henkilöihin.

— Luulenpa Fredrik'in aikovan ahdistaa hengen neiti
Rudenschöld'stä… hän on todellakin rehoittava kaunotar, ja Fredrik
näyttää ihastuneelta, — lausui Kaarlo-herttua, jonka silmät olivat
ikäänkuin tarttuneet kiinni Magdalenan pieniin jalkoihin, jotka hänen
juostessansa vilahtelivat hänen runsaspoimuisen hameensa
laskoksien alta.
Ja Fredrik-herttua oli todellakin ihastunut. Pienellä kaappauksella

onnistui hänen saada kiinni tuota läähättävää tyttöä kädestä:
— Sainpa teidät sittekin kiinni, — lausui hän katsellen tulisilla

silmäyksillä Magdalenan hehkuvia kasvoja — seisottehan te, paroni
Armfelt, aivan meidän tiellämme! Eikö teillä ole halua olla
vuorostanne leskenä?
— Ei sinä eikä tänä. Kuninkaan suosikin vastaus oli jokseenkin

lyhyt, ja se silmäys jonka hän loi herttuaan oli kaikkea muuta paitsi
herttainen.
Fredrik kääntyi Magdalenaan hymyillen: —Nyt emme enään eriä

toisistamme, neiti Rudenschöld, suostutteko siihen?
— Koetan tehdä parastani, — lausui Magdalena hilpeästi, luoden

salaisen silmäyksen Armfelt'iin, joka nyt oli lähestynyt sitä ryhmää,
missä kuningas oli.
— Gösta-hyväni, — lausui Kustaa III, joka hiukka tämän jälkeen

Armfelt'in rinnalla meni liinaan päin: — sinä näytät synkältä kuin
ukkoisilma. Jos kellään olisi syytä näyttää sellaisia kasvoja, olisi sitä
minulla.
— Huolien ja murheiden täytyy kuten kaiken muunkin olla teidän
majesteetillenne alamaisia.
- Typerästi sekä sanottu että ajateltu, Gösta! Minun huoleni ja

murheeni ovat väliin sangen lähelle tuppaavia. Tiedätkö miten minun
hyvästi harkitun hankkeen! leskikuningatar Juliana Mariaan nähden
on käynyt? Niin, koko tulos on eräs kohtelias kirje häneltä tavallisilla
ystävyyden vakuutuksilla, — hymyily ilmestyi Kustaa-kuninkaan
huulille, — kuitenkin olemme kaikessa hyvällä kannalla Tanskan
suhteen, enkä minä tietysti muuta enemmän toivokaan kuin rauhaa
naapurieni kanssa.
Kuningas, joka tässä silmänräpäyksessä kokonaan oli vajonnut

omiin ajatuksiinsa, ei huomannut tuota hajamielisyyttä, millä Armfelt
häntä kuulteli, ja hän jatkoi hetkisen vaijettuansa:
— Valtakunnan sisällinen hoito vetää kokonaan puoleensa kaiken

huomioni keskellä huvitusten pyörteitä… toivon, etteivät
naapurivaltakuntain lähettiläät unohda kertoa hallitsijoillensa
huvitusten riistävän minulta milt'ei kaiken aikani. Minä olen erittäinkin
kutsunut tänne venäläisen lähettilään, jotta hän tänään oikein läheltä
saisi nähdä huolettomuuttani ja sokeata kevytmielisyyttäni, joka
saattaa minut unohtamaan kaiken muun. Niin, tämä mies, joka
rahoja jakamalla ja juonittelemisella on herättänyt valtiopäiväin
uppiniskaisuuden minua vastaan… hänen tahtonsapa se määrääkin
päätökset… nyt on minulla syytä rauhanrikkomiseen Venäjän kanssa
ja, jos Jumala suo, ei tämä tule kauan viipymään, vaikka minä nyt
olen menevinäni heidän mukiinsa.
Kuningas, joka astui työhuoneesensa, työnsi syrjään tuon vihriän

silkkiverhon, joka varjosti akkunaa, ja silmäillen ulos jatkoi hän:
Nämät valtiosäädyt, joilta odotin kannatusta ja rahanpulan
poistamista, kuinka no ovat minut pettäneet! Venäjän johdannon alla
oleva aatelisto tahtoo olla muurina minun ja kansan välillä,
rientääksensä minun perikatoni avulla tarkoitustensa perille, mutta
minä tahdon kumminkin ehkäistä Venäjän kunnianhimoa.
— Kansan toivo luottaa siinä kohdassa järkähtämättömästi teidän

majesteettinne…
— Kansa — keskeytti häntä Kustaa-kuningas, ja sisällinen tuska

piirsi uurtojaan hänen kasvoillensa; — kuinka onnetoin minä olenkin!
Kansani rakkaus on menetetty! Mitä on tullut tuosta innostuksesta,
joka minua kaikkiaalla ympäröi, tuosta luottamuksesta, joka oli minun
onnellisuuteni? Kaikki nämät sydämet, jotka ennen olivat täynänsä
suosioa ja rakkautta, ovat nyt kuni jäästä; minulla ei ole enään
mitään toivomista. Oltuaan rakastettu kuten minäkin, ei enään
koskaan voita takaisin niitä sydämiä, jotka ovat katkaisseet niin lujat
siteet. (42)
— Teidän majesteettinne oman ja maamme menestyksen

nimessä… oi, sire, älkäät antautuko näille tuskallisille ajatuksille —
huudahti Armfelt, hämmästyneenä siitä epätoivosta, jonka valtaan
kuningas muutamaksi sekunniksi hervottomasti oli heittäytynyt.
— Mutta niinpä on… kuinka voivatkin he neuvoa kutsumaan

ko'olle valtiopäiviä mielipiteiden ollessa sellaisina… ja Toll, joka
lupasi johtaa valtiopäiviä minun toivomusteni ja maamme tarpeiden
mukaan! Minun omat sukulaisena liittyvät vihollisiini… Kaarlo
imartelee heitä, ja he vuorostansa häntä. Juuri äsken koetin antaa
hänen ajatuksillensa toisen suunnan — ivallinen, milt'ei iloinen hymy
ilmestyi kuninkaan huulille, — ja muutaman päivän kuluttua toivon
hänen heittävän vehkeilemisensä minua kohtaan. Kas tuolla hän

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Advancing Healthcare Through

Personalized Medicine 2nd Edition

Cancer Immunotherapies Solid Tumors and Hematologic

Alagappan s Clinical Medicine for Dental Students 3rd

Practical Data Analytics for Innovation in Medicine:

Advancing Obesity Solutions Through Investments in the

Translational Bioinformatics and Systems Biology

Vision and Action Reinventing Schools Through

Hot and Cold Theory The Path Towards Personalized

Advancing Therapeutic Development for Pain and Opioid

ISBN 978-3-030-80099-4 ISBN 978-3-030-80100-7 (eBook)

San Mateo, CA, USA Priya Hays

1 “Introduction: Biomedical Innovation and Policy

3 “Patient Narratives: Personalized Medicine in the Field” �������������������� 83

Inherited CRC Syndrome���������������������������������������������������������������������� 212

Summary of PROSPER, SPARTAN, and ARAMIS������������������������������ 295

Study Finds Asciminib Safer and More Effective

Immunotherapy Clinical Trials�������������������������������������������������������������� 353

6 Trends in Precision Oncology and Precision Medicine 2.0�������������������� 419

Personalized Vaccine Approaches���������������������������������������������������������� 470

Pharmacogenetics of Schizophrenia������������������������������������������������������ 516

Evidence for Pharmacogenomic Testing: The Case

Economics of Precision Medicine Driven-Drug

The development of Xalkori, also known as crizotinib, a small molecular inhibitor

© The Author(s), under exclusive license to Springer Nature Switzerland AG 2021 1

the treatment of HER2-positive metastatic breast cancers in September 1998, and on

As the abovementioned examples indicate, by focusing on creating drugs for

survival regardless of neoadjuvant regimen and molecular characteristics, along

In order to ascertain the impact of a biomarker-baesed (personalized) strategy, clinical out-

Fig. 1.1 (a) Umbrella a

Biomarker A Biomarker B Biomarker C

Treatment X Treatment Y Treatment Z

Biomarker A Biomarker B Biomarker C

Treatment X Treatment Y Treatment Z

• A $215 million investment in the 2016 budget

• Treating patients according to their individual genes, lifestyle, and environments

Investments to launch this initiative include funding to the National Institutes of

Data analysis, integration and

Traditional Medicine Stratified Medicine Precision Medicine

Precision medicine research enables development and delivery of the right

effects of personalized medicine on patient clinical outcomes and overall quality of

But we live in a very exciting time

Cheap “omics” and other

1000s of non-cancer drugs that Remarkable

Clinical trial innovations: Favorable regulatory

(1994–2003) of doubling, the rate of increase in biomedical research funding slowed

There is a certain level of dissonance present. With basic science threatened by

genomic-guided therapy. According to Geoffrey Ginsburg, MD, PhD, of the Duke

• Vermurafenib (ZEBORAF®) approved for BRAF V600E–positive melanoma

FoundationOne, a molecular diagnostics company, has a companion diagnostic

Another highly significant development in precision medicine is the use of liquid

— Ruhtinatar kutsuu teitä, Magdalena!

Ja nyt tunsi hän itsensä samojen suojelevien käsivarsien

— Se ei olekaan totta, Magdalena, teitä ei olo kukaan kutsunut;

Magdalena, joka yhä antoi taluttaa itseänsä kauemmaksi, näki

Mennyttä, ijäti mennyttä oli Armfelt hänelle, tahi tuli tämän

— Päätäni vaan vähän huimasi, pyörrytti,— kuiskasi Magdalena,

— Luuletteko te kenenkään huomanneen,., oi, minä en soisi…

— Vihkimisen aikana olin minä teitä lähinnä, ja muiden huomio oli

— Minä voin nyt paremmin…

— Kuten sanoin, minä luulen sangen harvan teitä katselleen ja jos

Katkera tuskan vivahdus ilmestyi Magdalenan huulille. Eikö

Hän nousi äkkiä tuolta pitkältä, kapealta sohvalta, jolle hän

San Mateo, CA, USA Priya Hays

1 “Introduction: Biomedical Innovation and Policy

3 “Patient Narratives: Personalized Medicine in the Field” �� 83

Inherited CRC Syndrome�� 212

Summary of PROSPER, SPARTAN, and ARAMIS�� 295

Immunotherapy Clinical Trials�� 353

6 Trends in Precision Oncology and Precision Medicine 2.0�� 419

Personalized Vaccine Approaches�� 470

Pharmacogenetics of Schizophrenia�� 516