C H A P T E R

6
Biologics: Delivery options and
formulation strategies
Ridahunlang Nongkhlawa, Parameswar Patraa, Akash Chavrasiyaa,
Nirmal Jayabalana, Sachin Dubeyb
a
Department of Pharmacy, Birla Institute of Technology and Science, Pilani, Hyderabad, India
b
Formulation, Analytical and Drug Product Development, Glenmark Pharmaceuticals, La Chaux de
Fonds, Switzerland

1 Introduction include antibodies, interleukins, and protein-,

The uniqueness of biologic products has
paved the way for treating various types of dis-
eases wherein it targets a particular defected
gene or protein. About three decades ago, bio-
logics entered the market and the commercial
success they have achieved has proved their ben-
efits in patient care. Almost >300 biologics have
been approved since then, used to treat millions
of people, and about 900 biologics are under
development. The importance of biologics in
today’s world continues to increase, but many
challenges still need to be addressed during their
development. While ensuring the therapeutic
effectiveness of biologics, there is a growing
interest from many researchers in identifying
ways to overcome these challenges. Biologics are
pharmaceutical products manufactured or
extracted through a biological process (involving
biotechnology methods) from living organisms
rather than chemical synthesis. Types of biologics
peptide-, and vaccine-based products (Fig. 6.1
classifies different types of biologics). Recently,
biologics have rapidly emerged as an important
class of pharmaceuticals due to several advan-
tages. Biologics are in clinical use for several
life-threatening and rare diseases such as cancer,
diabetes, anemia, rheumatoid arthritis, multiple
sclerosis, etc. Biologics are considerably more
complex and expensive than a small molecule
pharmaceutical product. Nearly 30% of all drugs
approved by the U.S. Food and Drug Administra-
tion (FDA) in 2015–18 were biologics [1]. In the
near future, biologics-based products are
expected to attain a significant market share
among various pharmaceuticals.
Biologics have further got great impetus by the
advent of biosimilars. A biosimilar is a biological
product that is highly similar to and has no clini-
cally meaningful differences from an existing
FDA-approved reference product. In simpler
terms, biosimilars are medicines that are highly

Drug Delivery Aspects 115 # 2020 Elsevier Inc. All rights reserved.
https://doi.org/10.1016/B978-0-12-821222-6.00006-3
116 6. Biologics: Delivery options and formulation strategies

Biologics

Vaccines Blood Allergenics Gene Human Proteins and

components therapy tissue Peptides

FIG. 6.1 Types of biologics.

similar and clinically equivalent to complex bio-
logics medicines approved for use against serious
life-threatening diseases in the fields of immunol-
ogy, gastroenterology, and oncology. In terms of
product, biosimilars are divided into recombinant
nonglycosylated protein, recombinant glycosy-
lated protein, and recombinant peptides.
The biosimilar market is expected to reach
>23 billion USD by 2023. >60 biosimilar prod-
ucts are now in the pipeline in therapeutic areas
such as oncology, immunology, and diabetes.
There are many blockbuster biologic products
of pharmaceuticals companies, such as Remi-
cade, Rituxan, and Herceptin, and other patents
have expired in the recent past, and many will
expire in the upcoming decade. This creates
new opportunities for many pharmaceutical
companies to launch their product and enter
the biologics market. The various importances
of biologics by virtue of sales are summarized
in Table 6.1; they are becoming more and more
significant. Currently, the majority of biologics
are delivered either by intravenous infusion or
by subcutaneous injections; alternative delivery
options for biologics have been explored to some
extent, but are largely elusive in the current sit-
uation. The main aim of this chapter is to pro-
vide an update on the current status with
alternative routes and to highlight the possibili-
ties with these routes.

TABLE 6.1 Revenue from some of the top products in the USA.
Product Biologics Sales in
name (active) Manufacturer Indication/Disease 2018

Humira Adalimumab AbbVie Inc. Rheumatoid arthritis, plaque psoriasis, Crohn’s $20.2 b
disease
Revlimid Lenalidomide Celgene Multiple myeloma $ 9.2 b
Enbrel Etanercept Pfizer/Amgen Rheumatoid arthritis $ 7.3 b
Eylea Aflibercept Bayer Wet macular degeneration $ 6.5 b
Avastin Bevacizumab Roche Metastatic cancers $ 6.4 b
Rituxan Rituximab Roche Non-Hodgkin’s lymphoma $ 6.4 b

Herceptin Trastuzumab Roche Breast cancer $ 6.4 b

Remicade Infliximab Johnson & Johnson Rheumatoid arthritis $ 6.3 b
Keytruda Pembrolizumab Merck & Co. Metastatic melanoma $ 6.1 b
Xarelto Rivaroxban Johnson & Oral anticoagulant $ 6.1 b
Johnson/Bayer
 2 Intravenous infusion of biopharmaceuticals
2 Intravenous infusion of
biopharmaceuticals
Intravenous administration is one of the com-
mon delivery options for biopharmaceuticals—
this route of administration allows maximum
bioavailability, and for certain indications (e.g.,
oncology) where patients are in hospital, this
is often the most convenient option. Intravenous
administration is usually used when rapid onset
of action is required (e.g., emergency medicine
and anesthesia) or the drug is unable to be
administered orally because of some inherent
physicochemical properties of the drug mole-
cule. Intravenous administration is advanta-
geous over other routes for biopharmaceutical
administration as it bypasses the absorption bar-
riers and other metabolic pathways. By this
route, high-concentration biopharmaceuticals
can be administered rapidly to the relevant
organ, which may also reduce toxic effects.
Drugs in the form of suspensions or oily solu-
tions cannot be given intravenously. IV adminis-
tration bypasses many of the absorption
barriers, efflux pumps, and metabolic mecha-
nisms. Drugs administered via the intravenous
route reach the brain directly, with onset occur-
ring within 20–40 s. IV infusions may be used to
achieve a constant level of drug in the blood-
stream. Multiple doses can be administered
through an intravenous infusion, with a high
degree of flexibility and control. Biologic prod-
ucts such as infliximab, abatacept, rituximab,
tocilizumab, vedolizumab, etc. commonly used
for various clinical indications like hyperten-
sion, rheumatoid arthritis, Crohn’s disease,
ulcerative colitis, psoriasis, psoriatic arthritis,
etc. are administered via the IV route.
Intravenous administration can be achieved
by two ways: bolus injections or continuous
infusion. With a bolus injection, complete med-
ication is given in a small period of time—for
example, in <10 min. The bolus injection pos-
sesses several characteristics such as being easy
117
to perform, rapid action, short duration of con-
trast effect, etc. The second approach to admin-
ister the drug using this approach is by infusion;
an intravenous infusion is given at a slower rate
over an extended period of time without a suit-
able diluent. An IV infusion possesses several
characteristics like extended drug release in
the blood, consistent contrast effect, easy to
adjust dose as per clinical requirements during
treatment, requires infusion kit, infusion titra-
tion needed, etc. The drug is delivered immedi-
ately via the IV route and therefore rapid onset
of drug effects is observed. Dose adjustment as
per patient need can be done easily by IV admin-
istration. Drug candidates having low bioavail-
ability can be administered suitably by the IV
route with 100% bioavailability (Table 6.2).
Products meant for IV administration should
be sterile, free of pyrogens, buffered to physio-
logical pH, isotonic, stable, isohydric, and iso-
viscous. Generally for the drugs which are
unstable in the GIT environment are not suitable
for administration through the oral route as this
route of administration has lower bioavail-
ability. Therefore, IV route is preferred in such
scenarios. Among several advantages, there
are some limitations with this route of
administration—the patient needs to be hospi-
talized for IV administration, which incurs
patient discomfort, high treatment cost, etc.
Any wrong medication, dosing, or contamina-
tion may result in serious side effects or hyper-
sensitivity reactions, which could be fatal.
Within IV administration, the use of
advanced delivery options like stealth systems,
slower release, etc. are largely limited to small
peptides and vaccines. The most likely reason
for this is their size and potential instability dur-
ing processes (e.g., use of organic solvents)
required for consistent manufacturing of these
advanced delivery systems. At the other end,
advancements in discovery are pushing the
limits with gene therapy, RNA-based delivery,
and even CAR-T based advance treatments,
TABLE 6.2 List of biologics for intravenous use approved by the FDA.
Brand name Indications Target Year of approval Company name

Reopro (abciximab) Cardiac ischemic complications GPIIb/IIIa receptor 12/22/1994 (U.S.) Janssen Biologics B.V.

Herceptin (trastuzumab) Breast cancer, HER2-positive gastric HER-2 09/25/1998 (U.S.) Roche
cancer

Remicade (infliximab) Psoriatic arthritis TNF-α 08/24/1998 (U.S.) Johnson & Johnson
Tysabri (natalizumab) Multiple sclerosis (MS) and Crohn’s Integrin receptor 11/23/2004 (U.S.) Biogen
disease (CD) antagonist

Nutropin Depot (somatropin) GH treatment IGF-I 12/22/1999 (U.S.) Genentech, Inc.

Nivestym (filgrastim) Neutropenia, acute myeloid leukemia G-CSF 07/20/2018 (U.S.) Pfizer Inc
(AML)
Retacrit (epoetin alfa) Chronic kidney disease (CKD) Hemoglobin 05/15/2018 (U.S.) Hospira, Inc.a Pfizer
Ixifi (infliximab) Crohn’s disease, ulcerative colitis TNF 12/13/2017 (U.S.) Pfizer Inc.
Mvasi (bevacizumab) Metastatic colorectal cancer VEGF 09/14/2017 (U.S.) Amgen
Renflexis (infliximab) Crohn’s disease, ulcerative colitis TNF 04/21/2017 (U.S.) Merck & Co. Inc.
Inflectra (infliximab) Pediatric Crohn’s disease, ulcerative TNF 04/05/2016 (U.S.) CELLTRION Inc
colitis
Rituxan (rituximab) Non-Hodgkin’s lymphoma (NHL) CD20 11/26/1997 (U.S.) Genentech
Simulect (basiliximab) Prophylaxis of acute organ rejection CD20 05/12/1998 (U.S.) Novartis Pharmaceuticals
Corporation
Mylotarg (gemtuzumab Ozogamicin) Acute myeloid leukemia (AML) CD33 05/17/200 (U.S.) Pfizer Inc
Erbitux (cetuximab) Metastatic colorectal carcinoma EGFR 02/12/2004 (U.S.) Eli Lilly and Company

Truxima (rituximab) Non-Hodgkin’s lymphoma (NHL) CD20 11/28/2018 (U.S.) TEVA Pharmaceuticals
Lemtrada (alemtuzumab) Multiple sclerosis (MS) CD52 05/07/2001 (U.S.) Genzyme Corporation
Zevalin (ibritumomab tiuxetan) B-cell non-Hodgkin’s lymphoma (NHL) CD20 02/19/2002 (U.S.) Spectrum Pharmaceuticals Inc.
Herzuma (trastuzumab) Metastatic breast cancer HER2 12/14/2018 (U.S.) Teva Pharmaceuticals
Andexxa (coagulation factor Xa Factor Xa 05/03/2018 (U.S.) Portola Pharmaceuticals Inc.
(recombinant)
Cathflo activase Restoration of function to central venous t-PA 11/13/1987 (U.S.) Genentech
system
Ogivri (trastuzumab) Breast cancer HER2 12/01/2017 (U.S.) Mylan Pharmaceuticals Inc.

Notes: Nonexhaustive list, mainly to show examples of different categories. Information has been obtained for different products from www.accessdata.fda.gov (accessed on June 22,
2019).
 3 Subcutaneous injection of biopharmaceuticals
and the most convenient option for delivering
these therapeutics is either by a direct local injec-
tion or more commonly through intravenous
administration.
3 Subcutaneous injection of
biopharmaceuticals
The second most commonly used and clini-
cally feasible delivery route for biologicals is
by subcutaneous injection. The success of insu-
lin through subcutaneous injection has given
way to many more products being developed
through this route. Several studies are under-
way to develop products to administer through
the subcutaneous (SC) route. The importance
gained by the SC route is because of the ease
of administration compared to the IV route,
which requires trained personnel to administer
the drug [2–4] (Table 6.3). In the SC route, the
drug can be self-administered, which increases
patient compliance tremendously. The uptake
of drugs injected through the SC route occurs
by two ways: blood capillaries and lymphatic
vessels. Drug molecules that are >16 kDa are
taken up by lymphatic vessels, while smaller
ones (<16 kDa) enter the systemic circulation
directly [5].
Major pharmaceutical companies and the
healthcare industry have diverted their atten-
tion toward safe and effective SC delivered pro-
teins. SC injections are a widely administered
route for biotherapeutics despite the huge
knowledge gap in SC biotherapeutic absorption.
Catabolic first-pass clearance after SC adminis-
tration is found to be one of the main contribu-
tors for many limitation of SC administered
biotherapeutics, but unfortunately catabolic
process in the lymphatic pathways is poorly
explored. The main advantage to this route lies
in its cost-effectiveness and self-administration.
SC delivery is more preferred among lower
molecular weight biologics, as rapid uptake
through the vascular network is observed. Some
119
of the success stories of SC delivery include
insulin (diabetes), bortezomib (oncology), defer-
oxamine (blood poisoning), and lanreotide (hor-
mone inhibition).
A major challenge to SC injection is the small
volume of injection (<1.5 mL) requiring protein
concentration >100mg which leads to an increase
in viscosity as well as causing manufacturing
difficulties for industries [6–8]. Injection volumes
>2.5mL are associated with pain in injection,
leakage, tissue distortion, and tissue back
pressure. Higher SC injection volumes have also
been evaluated using permeation enhancers.
The volume barrier within the ECM is created
by GAG hyaluron together with collagen. Hyal-
uronidase cleavage of hyaluron enables delivery
of biologics through the SC route by increasing
the tissue surface area and reducing tissue back
pressure. This leads to fast administration of SC
injection volumes >2.5mL with a reduced indu-
ration and leakage at the injection site. In spite
of the known potential of hyaluronidase, ways
to purify it from human body tissue need to be
established to overcome the limitations of immu-
nogenicity [9] or an alternative supply means are
required to be established. Surface charge, mole-
cule shape, solvent viscosity, pH, ionic strength,
temperature, and shear rate [8] are some variables
that affect the viscosity of a protein solution. The
development of high-concentration formulation
(due to high dose) of monoclonal antibodies poses
great challenges due to the chances of aggrega-
tion, viscosity, subvisible and visible particles,
as well as the method to concentrate the
proteins. The translation of SC animal data to
humans has not been clearly established.
Relatively very few data report the mechanism
of SC absorption of biologics in different
species. The role of lymph and blood capillaries
in systemic absorption, cross-species differences
in hypodermis morphology and physiology, drug
formulation, stability of the molecule, the site of
injection, the depth of injection, as well as the
molecular properties of the biologics themselves
are major parameters affecting the absorption
TABLE 6.3 List of biologics for the SC route approved by the FDA.
Date of
Brand name (drug name) Indications Target approval Company name

Udenyca (pegfilgrastim) Reduction in the duration of neutropenia Leukocyte growth factor 11/02/2018 Coherus
(U.S.) Bioscience

Fulphila (pegfilgrastim) Neutropenia in adult patients Leukocyte growth factor 06/04/2018 Mylan
(U.S.)

Hyrimoz (adalimumab) Rheumatoid arthritis, juvenile arthritis, Crohn’s disease, TNF-α 10/30/2018 Sandoz
ankylosing spondylitis, psoriasis (U.S.)
Tegsedi (inotersen) Polyneuropathy of hereditary transthyretin-mediated Human transthyretin 10/05/2018 Ionis
amyloidosis in adults (TTR) protein (U.S.) Pharmaceuticals
Emgality (galcanezumab) Migraine Calcitonin gene-related 09/27/2018 Eli Lily
peptide (CGRP) (U.S.)
Ajovytm (fremanezumab) Migraine Calcitonin gene-related 09/14/2018 Celltrion
peptide (CGRP (U.S.)
Takhzyrotm (lanadelumab) Prevent attacks of hereditary angioedema (HAE) Plasma kallikrein 8/23/2018 Dyax Corp.
(U.S.)
Nivestym (filgrastim) Acute leukemia, neutropenia Leukocytes growth factor 07/18/2018 Pfizer
(U.S.)
Palynziq (pegvaliase) Phenylketonuria Phenylalanine- 05/24/2018 Biomarin
metabolizing enzyme (U.S.) Pharmaceuticals
Aimovigtm (erenumab) Migraine Calcitonin gene-related 05/17/2018 Amgen
peptide receptor (U.S.)
Retacrit (epoetin alfa) Chronic kidney disease Progenitor stem cells 05/17/2018 Pfizer
(U.S.)

Crysvita (burosumab) X-linked hypophosphatemia (XLH) Fibroblast growth factor 04/17/2018 Ultragenyx
23 (FGF23) (U.S.)

Ilumya (tildrakizumab) Plaque psoriasis Interleukin-23 03/21/2018 Merck Sharp

(U.S.)
Admelog (insulin lispro Types I and II diabetes mellitus Glucagon-like peptide 1 12/05/2017 Sanofi Aventis
injection) (GLP-1) receptor (U.S.)
Ozempic (semaglutide) Glycemic control in adults with type 2 diabetes mellitus Glucagon-like peptide 1 12/05/2107 Novo Nordisk
(GLP-1) receptor (U.S.)
Hemlibra (emicizumab) Hemophilia A with factor VIII Factor VIII 11/16/2017 Genetech
(U.S.)
Fasenra (benralizumab) Asthma Interleukin-5 receptor 11/14/2017 Medimmune/
alpha (U.S.) Astra zenneca
Fiasp (insulin) Glycemic control GLP-1 09/29/2017 Novo Nordisk
(U.S.)
Cyltezo (adalimumab) Rheumatoid arthritis, juvenile arthritis, Crohn’s disease, TNF-α 08/25/2017 Boehringer
ankylosing spondylitis, psoriasis (U.S.) Ingelheim
Pharmaceuticals
Tremfya (guselkumab) Psoriasis Interleukin 23 07/13/2017 Janssen Biotech
(U.S.)
Rituxan hycela (rituximab Chronic lymphocytic leukemia CD20, Hyaluridase 06/23/2017 Genentech
and hyaluronidase human) (U.S.)

Haegarda (C1 esterase Hereditary angioedema C1 Esterase 06/22/2107 CSL Behring

inhibitor subcutaneous) (U.S.)

Kevzara (sarilumab) Rheumatoid arthritis Interleukin 6 05/22/2017 Sanofi &

(U.S.) Regeneron
Dupixent (dupilumab) Severe eczema Interleukin-4 receptor 03/28/2017 Regeneron
alpha (U.S.)
Siliq (brodalumab) Plaque psoriasis Interleukin 17 receptor 02/15/2017 Valeant Pharma
(U.S.)
Basaglar (insulin glargine Type II diabetes GLP-1 12/16/2016 Eli Lilly
injection) (U.S.)
Xultophy (insulin degludec Diabetes mellitus type II GLP-1 11/21/2016 Novo Nordisk
(U.S.)
Soliqua 100/33 (insulin Diabetes mellitus type II GLP-1 11/21/2016 Sanofi
glargine and lixisenatide (U.S.)
injection)
Amjevita (adalimumab) Rheumatoid arthritis, juvenile idiopathic arthritis, Crohn’s TNF-α 09/23/2016 Amgen
disease, ulcerative colitis, alkylosing spondylitis, plaque (U.S.)
psoriasis

Continued
TABLE 6.3 List of biologics for the SC route approved by the FDA—cont’d
Date of
Brand name (drug name) Indications Target approval Company name

Cuvitru, immune globulin Primary immune deficiencies Immunoglobulin 09/14/2016 Shire

(U.S.)
Adlyxin (lixisenatide) Type II diabetes mellitus GLP-1 07/28/2016 Sanofi
(U.S.)
Zinbryta (daclizumab) Multiple sclerosis Interleukin 2 receptor 05/27/2016 Biogen
(U.S.)
Taltz (ixekizumab) Plaque psoriasis Interleukin 17A 03/22/2016 Eli Lilly
(U.S.)

Amjevita (Adalimumab) Rheumatoid arthritis, juvenile arthritis, Crohn’s disease, TNF-α 09/23/2016 Amgen
ankylosing spondylitis, psoriasis (U.S.)

Erelzi (etanercept) Rheumatoid arthritis, juvenile arthritis, ankylosing TNF-α 08/30/2016 Sandoz
spondylitis, psoriasis (U.S.)
Basaglar (insulin glargine Type II diabetes mellitus GLP-1 12/16/2015 Eli Lilly
injection)
Nucala (mepolizumab) Asthma Interleukin 5 11/04/2015 GlaxoSmithKline
(U.S.)
Strensiq (asfotase alfa) Perinatal, juvenile and infantile hypophosphatasia Alkaline phosphatase 10/23/2015 Alexion
(U.S.)
Tresiba (insulin degludec Diabetes mellitus GLP-1 10/16/2015 Novo Nordisk
injection) (U.S.)
Ryzodeg 70/30 (insulin Diabetes mellitus GLP-1 10/16/2015 Novo Nordisk
degludec and insulin aspart (U.S.)
injection)

Repatha (evolocumab) Treatment for low density level cholesterol (LDL) PCSK9 (proprotein 08/27/2015 Amgen
convertase subtilisin kexin (U.S.)
type 9)
Praluenttm (alirocumab) Treatment for low density level cholesterol (LDL) PCSK9 (proprotein 07/24/2015 Sanofi
convertase subtilisin kexin (U.S.)
type 9)
Neupogen (filgrastim) Neutropenia Leukocyte growth factor 03/06/2015 Novartis
(U.S.)
Toujeo (insulin glargine Diabetes mellitus GLP-1 02/25/2015 Sanofi
injection) (U.S.)
Cosentyxtm (secukinumab) Plaque psoriasis Interleukin 17 01/23/2015 Novartis
(U.S.)
Zarxio (filgrastim-Sndz) Incidence of infection‚ as manifested by febrile neutropenia‚ in Leukocytes growth factor 03/06/2015 Sandoz
patients with nonmyeloid malignancies (U.S.)
Trulicity (dulaglutide) Diabetes mellitus GLP-1 09/18/2014 Eli Lilly
(U.S.)
Hyqvia [immune globulin Immune globulin with a recombinant human hyaluronidase Immune globulin and 09/12/2014 Baxter
infusion 10% (human) with hyaluronidase (U.S.)
recombinant]
Plegridy (peginterferon Multiple sclerosis Interferon beta 08/15/2014 Biogen
beta-1a) (U.S.)

Tanzeum (albiglutide) Diabetes mellitus type II GLP-1 04/15/2014 GlaxoSmithKline

(U.S.)

Mircera (methoxy Anemia associated with chronic renal failure Erythropoiesis- 11/14/2014 Vifor
polyethylene glycol-epoetin stimulating agent (ESA) (U.S.)
beta)
Herceptin SC (trastuzumab) Breast cancer Human epidermal growth 02/10/2014 Genentech
factor receptor 2 (HER2) (U.S.)
Kynamro (mipomersen Antilipidemic Apo-B Lipoprotein 01/17/2013 Genzyme
sodium) (U.S.) Corporation
Gattex (teduglutide [rDNA Short bowel syndrome GLP-2 12/21/2013 NPS Pharma
origin]) (U.S.)
Benlysta (belimumab) Positive systemic lupus erythematosus B-lymphocyte stimulator 03/09/2011 Human Genome
(BLyS)-specific (U.S.) Sciences
Hizentra, immune globulin Primary immunodeficiency Immune globulin 03/04/2010 CSL Behring
(U.S.)
Prolia (denosumab) Postmenopausal with osteoporosis, to increase bone mass in Antireceptor activator of 06/01/2010 Amgen Inc.
men with osteoporosis at high risk for fracture, nuclear factor kappa-B (U.S.)
glucocorticoid-induced osteoporosis in men and women at ligand (RANKL)
high risk for fracture

Continued
TABLE 6.3 List of biologics for the SC route approved by the FDA—cont’d
Date of
Brand name (drug name) Indications Target approval Company name

Stelara (ustekinumab) Plaque psoriasis Interleukin 12 and 23 09/25/2009 Janssen Biotech

(U.S.)

Extavia (inteneron beta-l b) Multiple sclerosis Interferon beta 08/15/2009 Novartis

(U.S.)

Ilaris (canakinumab) Cryopyrin Associated periodic syndrome Interleukin-1β 06/17/2009 Novartis

(U.S.)
Pegasys (peginterferon alfa- Chronic hepatitis C, hairy cell leukemia, chronic IFN-α-2a 08/28/2009 Genentech
2a) myelogenous leukemia (U.S.)
Simponi (golimumab) Rheumatoid arthritis, ankylosing spondylitis, psoriasis TNF-α 04/24/2009 Janssen Biotech
(U.S.)
Cimzia (certolizumab Refractory Crohn’s disease TNF-α 04/23/2008 Bayer Schering
pegol) (U.S.)
Arcalysttm (rilonacept) Cryopyrin-associated periodic syndromes (CAPS) disorders Interleukin-1 02/27/2008 Regeneron
(U.S.)
Accretropin (somatropin) Pediatric growth failure Somatropin 01/24/2008 Cangene
(U.S.)
Valtropin Growth deficiencies Somatropin 04/19/2008 LG Lifesciences
(U.S.)
Omnitrope (somatropin Growth hormone deficiencies Human growth hormone 05/30/2006 Sandoz/
[rDNA origin]) (U.S.) Novartis
Zostavax (Zoster vaccine Herpes Zoster – 05/25/2006 Merck
live) (U.S.)

Vivaglobin Primary immunodeficiency Immune globulin 01/09/2006 ZLB Behring

(U.S.)

Iplex (mecasermin rinfabate Growth deficiency Insulin like growth factor 1 12/12/2005 Insmed Inc.
[rDNA origin] injection) (U.S.)
Hylenex recombinant To enhance delivery of local anesthesia Hyaluronidase 12/05/2005 Halozyme
(hyaluronidase human (U.S.) Therapeutics
injection)
Hydase (hyaluronidase To enhance delivery of local anesthesia Hyaluronidase 10/25/2005 Prima Pharm
injection) (U.S.)
Increlex (mecasermin Growth deficiency Insulin like growth factor 1 08/31/2005 Tercica Inc.
[rDNA origin] injection) (U.S.)
Levemir (insulin detemir Diabetes mellitus Insulin receptors 06/17/2005 Novo Nordisk
[rDNA origin] injection) (U.S.)
Orencia (abatacept) Rheumatoid arthritis CTLA-4/Fc fusion 12/23/2005 Bristol-Myers
(U.S.) Squibb
Amphadase (hyaluronidase To enhance delivery of local anesthesia Hyaluronidase 10/24/2004 Amphastar
injection) (U.S.)
Fuzeon (enfuvirtide) for HIV-1 infection HIV fusion inhibitor 10/15/2004 Hoffman
Injection (U.S.) La-Roche
Luveris (lutropin alfa for Infertility treatment Stimulation of follicular 05/24/2004 Serono Inc.
injection) development (U.S.)
Vitrase (hyaluronidase To enhance delivery of local anesthesia Hyaluronidase 05/04/2004 ISTA Pharm
injection) (U.S.)
Apidra (insulin glulisine Diabetes mellitus GLP-1 04/16/2004 Aventis
[rDNA origin] injection) (U.S.)

Xolair (omalizumab) Asthma IgE 06/20/2003 Genetech

(U.S.)

Iprivask (desirudin for Deep vein thrombosis Thrombin inhibitor 04/03/2003 Aventis
injection) (U.S.)
Humira (adalimumab) Rheumatoid Arthritis TNF-α 12/30/2002 Abbott
injection (U.S.)
Forteo (teriparatide) Osteoporosis Parathyroid hormone 11/26/2002 Eli Lilly
(PTH) (U.S.)
Rebif (interferon beta-1a) Multiple sclerosis – 03/07/2002 Serono
(U.S.)
Neulasta (pegfilgrastim) Nonmyeloid malignancies Leukocyte growth factor 01/31/2002 Amgen
injection (U.S.)
Pegasys (peginterferon alfa- Chronic hepatitis C, hepatitis B PEGylated IFN-α-2b 10/16/2002 Schering
2b) (U.S.) Corporation

Continued
TABLE 6.3 List of biologics for the SC route approved by the FDA—cont’d
Date of
Brand name (drug name) Indications Target approval Company name

PEG-intron (Peginterferon Chronic hepatitis C PEGylated IFN-α-2b 08/07/2001 Schering

Alfa-2b (U.S.) Corporation

Kineret (anakinra) Rheumatoid arthritis Interleukin-1 11/14/2001 Amgen

(U.S.)

Lantus (insulin glargine
injection)
Actimmune (interferon
gamma-1b)
Enbrel (etanercept)
Avonex (interferon beta-1a) Multiple sclerosis
Betaseron (interferon beta-
1b)
Humulin (insulin human)
Glycemic control in adults and children with type 1 diabetes GLP-1 04/20/2000 Sanofi Aventis
mellitus and in adults with type 2 diabetes mellitus (U.S.)
Chronic granulomatous disease, osteopetrosis IFN-γ-1b 25/02/1999 Horizon Pharma
(U.S.)
Rheumatoid arthritis, juvenile arthritis, ankylosing TNF-α 11/02/1998 Amgen
spondylitis, psoriasis (U.S.)
IFN-β-1a 27/05/1996 Biogen Idec
(U.S.)
Multiple sclerosis IFN-β-1b 07/23/1993 Bayer Health
(U.S.) care pharm
Glycemic control in adults and children with type 1 and type 2 GLP-1 10/28/1982 Eli Lily
diabetes mellitus (U.S.)

Notes: Nonexhaustive list, mainly to show examples of different categories. Information have been obtained for different products from www.accessdata.fda.gov (accessed on
June 22, 2019).
 4 Targeted localized delivery of biologics
process [10–12]. In addition, the anatomy and
physiology, and lymphatic absorption, may differ
among species. Hence, it is important to control
and maintain standard experimental protocols
to reduce sources of variability [13].
The injection site and injection rate play an
important role in the drug absorption after SC
injection due to various factors. Namely, the
thickness of hypodermis differs across various
sites of the body and from person to person
[14]. The lymphatic absorption and lymph
node uptake of proteins is also determined
by the site of injection. Differences in SC
uptake are generally attributed to differences
in blood flow to those areas and/or to regional
variations in lymph flow [12]. Apart from
these pressure gradients, lymph movement,
variable lymph flow, and blood flow affect
the drug absorption after an SC injection
[15]. However, the role of the injection site is
mostly unexplored. Another important factor
affecting the uptake from the SC site is molec-
ular size. Lymphatic uptake is usually around
10–100 nm [16]. Reports suggest that 100 nm
molecules/liposomes have been found to be
trapped at the site of injection, resulting in
decreased uptake [17] (Fig. 6.2).
4 Targeted localized delivery of biologics
The above two delivery routes, IV infusions
and SC injections, cover approximately 95% of
biological delivery. Nevertheless, several stud-
ies have been performed for alternative nonstan-
dard routes of administration from a delivery of
biologicals perspective. The majority of these
alternative routes focus on localized delivery,
rather than targeting systemic delivery. Local-
ized drug delivery is one delivery option
wherein a drug is administered specifically to
a particular organ to reduce the unwanted sys-
temic exposure, thereby avoiding the side effects
and to increase the therapeutic concentration in
127
the particular organ (Table 6.4). There are persis-
tent efforts in this direction, and Table 6.5 sum-
marizes all the ongoing efforts along with their
development stages. In the upcoming subsec-
tions, organ-specific delivery options will be dis-
cussed in detail.
4.1 Brain targeting
In the next 20 years there will be an increase in
global drug development for senior citizens and
patients with central nervous system (CNS) dis-
orders. Unfortunately, the drug development
for brain diseases has a poor success rate. The
aim of brain drug delivery is to treat brain disor-
ders by passing through the tight junctions
called the blood-brain barrier (BBB). The BBB
is a dynamic diffusion barrier which occurs
between the endothelial lining and the cerebral
microvasculature, made up of special tight junc-
tions to protect the brain from unwanted and
harmful substances entering via the blood-
stream [19, 20]. The areas of BBB researched
until now show that it is simply not a barrier that
blocks the drugs, but a complex, well-
coordinated, ever-adapting interface which
facilitates the communication between the
CNS and blood [19]. Biologics like proteins, pep-
tides, and monoclonal antibodies are macromol-
ecules which have greater resistance to reach the
target as they do not cross the BBB. Apart from
this, it has been identified that the presence of
enzymes that could lead to inactivation of the
drug pose a major hindrance after passing
through the BBB. There is a need to understand
these target junctions’ barriers and transporters
in order to promote the permeation of drugs
through the brain to reach the target [21].
Strategies have been put forward to improve
the therapeutic efficacy of the products at the site
of action. One of the reports suggests re-
engineering the therapeutic protein to get
through the BBB with the help of a molecular
128 6. Biologics: Delivery options and formulation strategies

Figure legend: Skin

architecture and
cellular components

= Blood vessels

= Lymphatics

= Collagen fiber

= Elastin fiber

= Fibroblast

= Adipocyte

= Macrophage

Administered
= MAb

= Langerhan’s
cell

Activated
=
Langerhan’s
cell

Dermal
= dendritic cell

Activated
= dermal
dendritic cell

= T-cell

FIG. 6.2 Challenges and opportunities for subcutaneous delivery of biologics [18].

Trojan horse technology. This is a peptide mono-
clonal antibody molecule which, when fused
together with the biologic molecule and intro-
duced into the brain, binds with the endogenous
transporter present in the BBB. This mechanism
of binding helps the transport of the drug
through these barriers in the brain [21]. Another
approach of delivering the therapeutic drug were
reported by Klyachko et al. in 2017 using immune
cells, macrophages and monocytes, etc., which
TABLE 6.4 List of biologics approved for localized delivery.
Date of
approved
Drug name (brand name) Indications Target by FDA ROA Company name

Brain drug delivery

Brineura (cerliponase alfa) Slows the loss of ambulation in Lysosomal 04/27/ Intraventricular BioMarin
symptomatic pediatric patients 3 years of N-terminal 2017
age and older with late infantile neuronal tripeptidyl
ceroid lipofuscinosis type 2 (CLN2), also peptidase
known as tripeptidyl peptidase 1 (TPP1)
deficiency
Spinraza (nusinersen) Treatment of spinal muscular atrophy Motor 12/23/ Intrathecal Biogen, Inc.
(SMA) in pediatric and adult patients neuron-2 2016
(SMN2)-
directed
antisense
oligonucleotide

Pulmonary drug delivery

Affreza (insulin human) Improve glycemic control in adult patients Glucagon like 07/27/ Inhalation Mankind Corporation
with diabetes mellitus peptide-GLP-1 2014
Exubera (insulin human, rDNA) Types 1 and 2 diabetes Glucagon like 01/27/ Oral inhalation Pfizer
peptide-GLP-1 2006

Pulmozyme (dornase alfa) Cystic fibrosis (CF) patients to improve DNAse 12/30/ Inhalation Genentech
pulmonary function 1993 solution

Transdermal drug delivery

Fluzone intradermal quadrivalent Prevention of influenza disease caused by Influenza 05/09/ For intradermal Sanofi Pasteur
(influenza vaccine) influenza A subtype viruses and type A and B 2011 use only
B viruses contained in the vaccine
Oral drug delivery
Orallair (sweet vernal, orchard, Grass pollen-induced allergic rhinitis with or Eye as target 04/01/ Sublingual Greer Labs
perennial rye, timothy, and 7 without conjunctivitis organ 2014
Kentucky blue grass mixed
pollens allergen extract)

Continued
TABLE 6.4 List of biologics approved for localized delivery—cont’d
Date of
approved
Drug name (brand name) Indications Target by FDA ROA Company name

Live, monovalent, human Prevention of rotavirus gastroenteritis Rotavirus 04/03/ Oral GlaxoSmithKline
attenuated rotavirus stain caused by G1 and non-G1 types (G3, G4, 2008 suspension
(rotavirus vaccine, live, oral) and G9)
Ocular drug delivery

Oxervate (cenegermin) Neurotrophic keratitis High affinity 08/22/ Topical Dompe Pharmaceuticals
nerve growth 2018 ophthalmic use
factor
Luxturna (voretigene Vision loss due to confirmed biallelic Biallelic RPE65 12/19/ Subretinal Spark Therapeutics
neparvovec) RPE65-mediated inherited retinal disease, 2017 injection
Eylea (Aflibercept) Diabetic retinopathy in patients with Vascular 07/29/ Intravitreal Regeneron
diabetic macular edema (DME) endothelial 2014 injection Pharmaceuticals
growth factor
(VEGF)

Jetrea (ocriplasmin) Symptomatic vitreomacular adhesion Alpha-2 10/17/ Intravitreal Thrombo Genics Inc.
antiplasmin 2012 injection
Lucentis (ranibizumab) Diabetic macular edema Vascular 08/10/ Intravitreal Genentech
endothelial 2012 injection
growth factor
(VEGF)
Macugen (pegaptanib) Macular degeneration VEGF 12/17/ Intravitreal Pfizer
2004 injection

Data collected from FDA website and product information leaflets.

TABLE 6.5 List of biologics under clinical trials for localized delivery.
Drug name (brand name) Indications Target Status Delivery approaches Company name

Brain delivery
Glial cell line-derived neurotrophic Parkinson’s disease r-metHuGDNF Phase II Convection enhanced North Bristol NHS
factor delivery Trust

Pulmonary delivery
Human insulin (Dance 501) Pharmacokinetic and Beta-cells Phase I, II Inhaler Dance Biopharm
pharmacodynamic profiles of Dance
501 in healthy subjects without
diabetes but with mild to moderate
asthma or COPD
Recombinant modified vaccinia Tuberculosis vaccine Mycobacterium Phase I Aerosol Oxford university
virus Ankara expressing antigen tuberculosis
85A (MVA85A)

PUR003 Flu vaccine Not listed Phase I Nebulizer Pulmatrix

Recombinant replication-deficient Tuberculosis vaccine Human Phase I Aerosol McMaster
human adenoviral tuberculosis adenovirus University
vaccine containing
immunodominant antigen Ag85A
(Ad5Ag85A)
Transdermal drug delivery
Teriparatide (MicroCor PTH) Osteoporosis Human Phase II Dissolving MicroCor PTH
parathyroid microneedles (1–34) (Corium)
hormone
(increase bone
growth)
Abaloparatide (baloparatide-TD) Osteoporosis in postmenopausal Human Phase II Transdermal Radius Health
women parathyroid microneedle patch
hormone
(increase bone
growth)
Insulin (icronJet600) Determining pharmacokinetics and Beta-cells Early Phase Hollow microneedles Nanopass
pharmacodynamics of insulin I

Continued
TABLE 6.5 List of biologics under clinical trials for localized delivery—cont’d
Drug name (brand name) Indications Target Status Delivery approaches Company name

C19-A3 GNP (proinsulin) Immunotherapy for diabetes Beta-cells Phase I Hollow microneedles Nanopass
MicronJet600
Glucagon (ZP-glucagon) Hypoglycemic Glucagon Phase I Transdermal patch Zosano
Gonadotropin releasing hormone Infertility Gonadotropin Phase II Iontophoretic patch Ferring
releasing Pharmaceuticals
hormone
(GnRH)
Mixture of peptides from islet Induce or restore immunological Phase I NanoPass MicronJet Kings College
autoantigens multipeptide tolerance to β-cells technology London
Inactivated polio vaccine Polio vaccine Phase II NanoPass MicronJet Nanopass
600 microneedle
device
Single multivalent peanut Phase I NanoPass MicronJet Astellas
lysosomal associated membrane technology
protein DNA plasmid (ASP0892)

Recombinant hepatitis B vaccine Hepatitis B vaccine Liver virus, Phase II/III NanoPass MicronJet
(hepatitis B virus vaccine) hepatitic technology
B virus
Oral drug delivery
Octreotide (mycapssa) Acromegaly Growth Phase III Capsule using the Chiasma
hormone proprietary
technology platform
transient permeability
enhancer
Semaglutide (NN9924) Type 2 diabetes Glucagon like >25 trials; Tablet with Novo Nordisk
peptide Phase III/I absorption-enhancing
receptor excipients
Leuprolide (ovarest) GnRH Phase II
 Pharmacokinetic and Peptelligence: Enteris Biopharm
pharmacodynamic profiles in healthy improved solubility Inc.
female volunteers and absorption of
peptides for oral
delivery

Salmon calcitonin Postmenopausal osteoporosis in Calcitonin Phase III/II Tablet Tarsa therapeutics
women receptor
Interferon-α Idiopathic pulmonary fibrosis Viral RNA Phase II Oral lozenge Texas Tech
University Health
Sciences Center/
Amarillo
Biosciences, Inc.
Anti-CD3 monoclonal antibody Hepatitis C CD-3 Phase II Oral delivery Inspira medical
Ocular drug delivery
AAVCAGsCD59 AMD Vascular Phase II/III Intravitreal injection Hemera
endothelial Biosciences
growth factor

Data collected from clinical trial website (https://clinicaltrials.gov).

134 6. Biologics: Delivery options and formulation strategies
have the capability to infiltrate into the brain
during inflammation and were shown to be
successful in a mouse model to deliver the thera-
peutic neurotropic factors and other enzymes
[22]. Manipulation of transporters, secretory
functions, extracellular pathways, and adsorp-
tive transcytosis are examples of promising
approaches for drug development [23]. Fur-
thermore, the application of nanocarriers like
polymeric nanoparticles, liposomes, Dendrimers,
micelles, etc. could be a useful approach in
delivering biologicals into the brain [24]. Over
the last two decades, the use of viral vectors like
lentivirus, herpes simplex virus, and
adeno-associated virus has proven to be one of
the options in brain delivery. In addition, exo-
somes, which are extracellular small vesicles pro-
duced by the cells, could offer more advantages
over nanoparticles because of their nonimmuno-
genicity in nature. These have been used to
deliver proteins and nucleic acids with the aim
to cross the barriers in the brain [25]. Delivering
drugs through another route like the intranasal
route, which could bypass the BBB and reach
the brain directly, have also been explored and
could be one of the options in directing the
delivery of biologics in future [26].
In relation to this, most of the biological drugs
have been approved by the FDA (Food and Drug
Administration) like natalizumab for the treat-
ment of multiple sclerosis and bevacizumab
for brain cancer, but the commercial success of
these drugs is low because of poor permeation
through the tight junctions (BBB) in the brain.
Due to the major challenges in crossing the
BBB and various other reasons, very few bio-
logics have been approved by the FDA for treat-
ing brain disorders. However, these drugs
require direct injection into the cerebrospinal
fluid to exert therapeutic efficacy. Intrathecal
ziconotide (2.6 kDa) for chronic pain, intrathecal
nusinersen (
7 kDa) for spinal muscular atro-
phy (SMA), and intraventricular cerliponase alfa
(59 kDa) are some biologics approved by the
FDA for CNS diseases [27]. A few biologics for
multiple sclerosis have emerged as clinical suc-
cesses in treating the disease. DNAse or deoxy-
ribonucleases is another biologic that has been
identified by researchers as playing an impor-
tant role in the apoptotic process. These bio-
logics are the enzymes which consist of
DNAse 1 and DNAse 2. These enzymes cleave
the DNA at a molecular level for various appli-
cations like RNA isolation, DNA fragmentation,
etc. Reports suggest end-stage Alzheimer’s dis-
ease could be improved by injecting DNAse 1
into the brain [28].
4.2 Pulmonary delivery
Pulmonary drug delivery appears to be the
easy route of administration as this allows the
delivery of drugs directly into the lungs and
hence drug deposition through this route is
more efficient due to the large absorptive sur-
face area (between 70 and 100 m2) and the thin-
ness of the alveolar epithelium (between 0.5
and 1.0 μm) [29]. Due to the limited presence
of drug metabolizing enzymes in the lungs,
delivery of biologics does not pose a hindrance
like other routes. Pulmonary drug delivery can
be self-administered by patients. Although this
route offers many advantages, many challenges
associated with the physicochemical properties
of biologics and delivery systems like the insta-
bility of biologics, size of particles, and shape
and delivery mechanism need to be addressed.
Lungs have a complex defense mechanism to
pass inhaled drug particles out of them and
to eliminate these once deposited. All these fac-
tors have a greater impact on delivering bio-
logics through this route. Many other
approaches have been taken into consideration
in developing products that can overcome these
challenges.
Studies have reported that only particle sizes
ranging from 1 to 5 μm are suitable for pulmo-
nary delivery, and the shape of particles should
also be taken into consideration. Interestingly,
 4 Targeted localized delivery of biologics
nonspherical shapes are mostly preferred to
avoid the particles getting captured by macro-
phages [30]. Maintaining the size of particles
for pulmonary drug delivery is key to overcome
the challenges. The size has to be optimized
properly because the large particles get depos-
ited in the central surface of airways with more
drugs per unit surface area, whereas with small
particles, the drug gets deposited in the periph-
ery of the airways with less amount of drug per
unit surface area. Micronization for controlling
particle size is one option to deal with this chal-
lenge [31]. Other approaches like developing
particulate nanocarrier systems such as micro-
particles, nanoparticles [32], and liposomes have
been widely used and provide benefits in
improving delivery of drugs [33]. Many other
approaches have been taken into consideration
in developing products that can overcome these
challenges. The design of the device to deliver
the drugs during inhalation also has a greater
impact in deciding the bioavailability of the
drug. The most available devices for inhalation
at present are the pressurized meter dose
inhalers (pMDIs), nebulizers, and dry powder
inhalers (DPIs). Each of these has its advantages
and disadvantages [34]. There is a need to
design an ideal device that can be used to deliver
drugs in an ideal manner [35].
The pipelines of biologics for pulmonary
delivery remain a smaller portion in the market
due to the abovementioned challenges. Cur-
rently biologics approved by the FDA for pul-
monary drug delivery system are dornase
alfa, pulmozyme, exubera (insulin—withdrawn
2007), and afrezza [36] (Table 6.4). Among all
the biologics, DNAse is one of the first promis-
ing proteins that was approved by the FDA in
1993 (Pulmozyme, Genentech, Inc.). It works
by hydrolyzing the DNA strand or breaking
down the strands of DNA in the airway tract
and clears the sputum to improve the complica-
tions of diseases. DNAse has played an impor-
tant role as a protein therapeutic which is
applicable clinically at the time of writing for
135
pulmonary drug delivery [37] to treat the most
commonly affected diseases like cystic fibrosis
(CF). CF is a disease that results in abnormal
functioning of the lungs due to respiratory tract
infections, which results in high concentrations
of DNA from degenerating polymorphonuclear
leukocytes in lungs. Due to a high concentra-
tion of DNA, the viscosity of the sputum in
the airway tract increases and hence results in
various respiratory inflammatory responses.
Pulmozyme has proved preclinically as well
as clinically to have more advantages over tra-
ditional inhalants, such as improved lung depo-
sition, decreased rate of administration, and
improved stability of the protein. All these
advantages could reduce patient compliance,
which is usually faced with the traditional
inhalants for treating such types of diseases.
Various studies have been done in order to
investigate the effect of DNAse as a therapeutic
protein to treat these types of diseases. Yang
et al. investigated inhalable antibiotic delivery
using a dry powder co-delivering recombinant
deoxyribonuclease and ciprofloxacin for treat-
ment of cystic fibrosis [38]. The study was
based on co-delivering ciprofloxacin and
DNAse through a single particulate system,
and it was suggested that this single particulate
system could be one of the strategies to improve
the delivery of antibiotics as the DNAse
enhanced the penetration activity of antibiotics
and thereby cleared off the high concentration
of sputum viscosity in cystic fibrosis patients.
This study was done to compare the efficiency
of antibiotics alone and in combination with
DNAse. The study showed that the effective-
ness of this particulate system was able to kill
the bacteria more efficiently when compared
to ciprofloxacin alone, because DNAse
improved the penetration of the antibiotics to
diffuse into the airway tract [39]. The delivery
route is still in the exploratory phase and
requires significant development before it can
be considered a standard delivery option for
biopharmaceuticals.
136 6. Biologics: Delivery options and formulation strategies
4.3 Transdermal delivery
One of the potentially ideal routes to deliver
therapeutic proteins and peptides is transder-
mal drug delivery. This route offers several
advantages like noninvasiveness and constant
delivery of drugs, hence avoiding frequent
interventions. The degradation of therapeutic
proteins can be avoided as this route has lim-
ited proteolytic enzymes which can degrade
the proteins compared to other routes, and
hence the bioavailability of proteins and pep-
tides can be improved. It is also convenient
for patients as it can be self-administered by
placing the patch on the skin, which is not
painful. Physiologically, it is well known that
skin is the largest organ in the body, and it acts
as an excellent natural barrier, preventing
most drug and foreign particles from entering
the body. This mechanism is governed by the
presence of the outermost layer of the skin
called the stratum corneum, which is about
10–15 um thick and consists of several brick-
type structures called corneocytes organized
in a bilayer lipid form. Therefore, only the
small molecules (approximately 500 kDa) that
have these physicochemical properties can
pass through this limiting barrier, whereas
the large macromolecules cannot diffuse
through it [40].
The success of transdermal drug delivery is
limited to the properties of the skin barrier,
mainly the presence of the stratum corneum,
which restricts the hydrophilic macromolecules
from entering the skin. Considering the
challenges faced by macromolecules, many strat-
egies and approaches have been developed to
address the problem by various physical and
chemical enhancement techniques. Some com-
monly used physical techniques are micropora-
tion, iontophoresis [41, 42], electroporation, and
sonophoresis [43, 44]. Similarly, chemical enhan-
cement techniques involve the use of permeation
enhancers like alcohols, surfactants, sulfoxides,
etc. in designing a formulation which could be
used in physical methods to deliver across skin
barriers [45]. The application of nanocarriers such
as liposomes, micelles, nanoparticles, etc. has
been widely used across the globe. In addition,
the prodrug approach is also an option that could
improve the permeation process—for example,
the conjugation of acyl derivatives with INF-α
improves the permeation rate up to 2.5- to
5-fold [46]. Although skin has a limited number
of proteolytic enzymes when compared to other
routes, protease inhibitors are being used for pep-
tide formulations so that the delivery process
could be enhanced much more [45]. Combina-
tions of multiple transdermal techniques have
further advanced the delivery of biologics with
promising results in recent years [47–49].
>20 biologics have been reported and inves-
tigated for delivery through the skin. Currently
>10 biologics are under development in clinical
trials (phases II and III), and >15 products are
based on the microneedle system. Most biologic
products for transdermal drug delivery are
proteins, peptides, and vaccines. Some of the
products tested are available in hydrogel
formulation—for example, single-chain anti-
TNF-α antibody DLX105 hydrogel, for which
phase II trials (NCT01936337) have been com-
pleted. This study was investigated to demon-
strate the safety, efficacy, and tolerability of
DLX105 hydrogel to treat mild to moderate pso-
riasis vulgaris. Many strategies were tested pre-
clinically using novel peptides for delivering
biologics across the skin using various mecha-
nisms. The delivery of biologics like small-
interfering RNA (siRNA), glyceraldehyde-3-
phosphate dehydrogenase (GAPDH),
interleukin-10, and insulin were delivered suc-
cessfully using pore-forming peptides such as
magainin, cell-penetrating peptides, peptides
that create transient openings in the skin, skin-
penetrating peptides, and peptides with protein
transduction domains. Details of the undergo-
ing clinical trials product for transdermal drug
delivery are listed in Table 6.5 [50].
 4 Targeted localized delivery of biologics
4.4 Ocular delivery
In the present scenario, treatments for various
ocular diseases have proved successful using
biologic products. Many of the biologic products
used for ocular treatment are monoclonal anti-
bodies, proteins, and peptides. However, the
blood ocular barriers like the blood-aqueous
barrier (BAB) and blood-retinal barrier (BRB)
of the eye pose a great challenge for the delivery
of large molecular weight biologics to treat both
anterior and posterior segment eye diseases. To
overcome the BRB, direct intravitreal injections
of monoclonal antibodies like anti-vascular
endothelial growth factor (anti-VEGF) are the
standard of care for treating many posterior seg-
ment eye diseases like age related macular
degeneration (ARMD), diabetic retinopathy
(DR), diabetic macular edema (DME), etc. How-
ever, the clinical success of these therapeutics is
challenged by a higher frequency of injection
[51]. In addition, treatment with such types of
biologics pose a challenge because of poor per-
meability across the retinal barriers, poor avail-
ability due to their large molecular size, stability,
and additionally the dynamic clearance mecha-
nism of the biologics from the eye. There is an
unmet medical need for these challenges to
improve patient compliance. There has been a
growing interest and enormous attention for
researchers to overcome these challenges by
developing several novel technologies like a sus-
tained drug delivery system using nanotechnol-
ogy for delivering biologics [52].
Many approaches have been studied on how
to overcome the various ocular barriers, and to
minimize the clearance mechanism. The nano-
technology at the time of writing is one strategy
that could respond to the challenges using both
physical and chemical methods. The enhance-
ment of permeation by different nanocarriers
like nanoparticles, micelles, and liposomes
was shown to have potential by many
researchers in delivering drugs to different ocu-
lar tissues [51].
137
DNAse is also one of the biological products
that has been used in the treatment of dry eye
diseases. Dry eye disease is a condition where
the eye lacks moisture and lubrication on the
ocular surface, which could lead to visual dis-
comfort, irritation, etc. A few studies have
reported that severe dry eye disease results
from inflammation due to decreases in tear
film nucleases (e.g., DNAse). Treatment of
dry eyes with DNAse1 eye drops has been
studied in clinical trials. The first eye drop of
DNAse (Pulmozyme 0.1%) administered four
times a day was studied topically by a Tibre-
wal and group. They determined the reduction
of extracellular DNA in dry eye patients by
using DNAse eye drops. Results showed that
DNAse eye drops could reduce the excessive
extracellular DNA from dry eye patients and
improved the dry eye symptoms and inflam-
mation as well [53].
Monoclonal antibodies and their subunits
(e.g., bevacizumab and ranibizumab) have
found a use in the treatment of ophthalmological
conditions. Many others have also shown
positive effects and are likely to be used in
future. Additional care has to be taken with
the ophthalmic delivery route mainly in terms
of skilled injection procedure as well as particle
control.
4.5 Oral delivery
The oral route of administration offers advan-
tages in delivering proteins and peptides
because of its ease of drug administration for
patients of all ages, and so the demand is high
due to patient convenience. Hence, oral drug
delivery of biologics remains a “holy grail” at
the time of writing. Unfortunately, the success
of this route for therapeutics is very poor
because of acidic environment and enzymatic
degradation due to the presence of intestinal
enzymes in the intestinal gut; on the other hand,
permeability of these agents is low due to high
138 6. Biologics: Delivery options and formulation strategies
molecular weight, which leads to low bioavail-
ability. The high demand for oral delivery of
proteins and peptides has led many researchers
to put great effort into developing an oral drug
delivery system that can overcome these
challenges.
Different strategies to overcome the chal-
lenges of oral drug delivery are: (i) use of
mucoadhesive polymers to increase contact time
of the drug with the physiological membrane
[54]; (ii) compounds that can disrupt the mem-
brane barrier and promote the transient opening
of the epithelial tissues [55]; (iii) materials that
can inhibit the degrading proteolytic enzymes
in the gut [56] and materials that can promote
dissociation of protein (e.g., cyclodextrins) [57].
All these nanotechnology-based mediated forms
of oral delivery have not yet improved beyond
the preclinical level. Although various
approaches are claimed and being investigated
by many researchers, many unaddressed chal-
lenges remain to be considered to make oral
drug delivery of biologics a reality.
Clinically, it has been reported that several
biologics are being investigated for oral delivery
across the globe. Commercially, there are two
peptides which are available in the market: (1)
Linzess (Iron Wood Pharmaceuticals) and Tru-
lance (Synergy Pharmaceuticals). Trulance was
approved in January 2017 for treating chronic idi-
opathic constipation (CIC). Interestingly, in Jan-
uary 2018, Trulance was approved for the
treatment of another disease called irritable
bowel syndrome with constipation (IBS-C) [58].
A few peptides are undergoing clinical trials
(Phase III), e.g., octreotide for acromegaly, sema-
glutide for diabetes, insulin for diabetes, salmon
calcitonin for osteoporosis, and desmopressin for
diabetes. In addition, about four vaccines have
been approved clinically (against vibrio, cholera,
typhoid and rotavirus). The biologics for oral
delivery that are undergoing clinical trials are
listed in Table 6.5.
5 Formulation strategies, degradation
routes, and role of excipients
Biologics (peptide, proteins, mAbs, and vac-
cines) are naturally less stable than small mole-
cules, and formulation development is often
challenging due to their complex structure.
Complex molecular structure, lack of well-
defined analytical tools, and multiple degrada-
tion mechanisms create major problems for the
formulation development of biologics and vac-
cines. Because of the limited availability of drug
substance for initial screening and development,
formulation scientists perform forced degrada-
tion studies using limited analytical techniques,
which help in developing optimal formulation
and provide insight in understanding degrada-
tion pathways of active molecules.
Formulation development is one of the most
important aspects of drug development, as
proper formulation ensures appropriate shelf life
and safety to patients. For biologics, the
formulation development is generally divided
into three interconnected stages: preformulation,
formulation, and process development. The
preformulation stage involves biochemical analy-
sis, amino acid sequencing, and biophysical
characterization of large molecules in the pres-
ence of pH, ionic strength, along with develop-
ment of stability indicating assay. This study
also helps researchers to understand degradation
pathways and therefore provides an approach to
select appropriate excipients of the required
characteristics. A thorough understanding of
macromolecular behavior can be obtained using
high-throughput formulation approaches to
explore changes in structure (e.g., secondary,
tertiary) and function (e.g., activity, potency,
binding wherever possible) in the presence of
stress conditions (e.g., pH, temperature, freeze-
thaw, drying, sheer). In the case of liquid formu-
lations, if the initial strategy fails to meet the
target product profile, it is recommended to
 5 Formulation strategies, degradation routes, and role of excipients
develop a lyophilized product using a suitable
lyophilization process. Application of quality
by design (QbD) in formulation development is
also useful to obtain a stable product with the
desired target profile. Through extensive stability
studies (both real-time and accelerated) and in
vivo animal model testing (when applicable), a
potential formulation can be selected for preclin-
ical and clinical study. To minimize the cost and
complexity of clinical study, it is recommended
that process and formulation optimization
should occur at an early stage.
Peptide, protein, and other biologically
derived macromolecules have rapidly emerged
as a major class of pharmaceuticals because of
their multiple advantages. Excipients are phar-
macologically inactive substances formulated
along with the active pharmaceutical ingredi-
ents (APIs) to impart specific physicochemical
properties to pharmaceutical products. Excipi-
ents have a defined functional role in pharma-
ceutical products like maintaining pH,
osmolality, solubility and bioavailability
enhancement, antioxidant effect, emulsifying
action, stabilization, etc. The stability of the fin-
ished product depends on the selection of the
excipients, their concentrations and the interac-
tion between drug-excipients and excipients-
excipients interaction. It is important during
the early stage development phase that the
excipients are carefully screened and optimized
based on the interactions mentioned above.
Owing to their clinical and commercial suc-
cess, biologics are the fastest growing class and
have become dominant over other conventional
therapy. Protein-based macromolecules now
constitute a major proportion of therapeutic
imperative for the treatment of various diseases.
But due to the fragile nature and complex struc-
ture of proteins, it is very challenging to maintain
integrity during processing, administration, and
distribution of these products. Two types of
degradation process are responsible for the
139
instability of protein molecular structure: physi-
cal instability and chemical instability (Table 6.6).
Various excipients like buffers, salts, sugars, and
surfactants are designed to improve the stability
of protein-based biologics. These excipients are
used to optimize the environment surrounding
the protein to maintain its conformational and
colloidal stability, reduce interactions with
neighboring proteins, and block interactions
with container surfaces. This section will provide
information about majorly used excipients along
with their role and functionality (Table 6.6).
Prevention against these degradation routes
requires careful selection of stabilizing excipi-
ents. The choice of the excipients is based on
the compatibility, functionality, and variability
of critical material characteristics within a cer-
tain acceptable range for development of a par-
ticular drug product. Selected excipients’
concentrations and characteristics can modulate
the finished product performance (e.g., bioavail-
ability and stability). Excipients used for the
product formulation should preferably be multi-
compendial [European Pharmacopeia (EP),
Japanese Pharmacopeia (JP), United States Phar-
macopeia (USP), etc.] so that the product can be
easily registered across the globe with the same
version of excipient. Currently, approximately
1000 excipients of >40 functional categories
are used in marketed pharmaceutical products.
Conventional excipients are simple in structure,
well known, pharmacologically inert, and of
natural origin such as sugar, minerals, and
wheat, but in recent years, many more novel
and highly complex excipients have been devel-
oped and evolved for specific usage, especially
for novel drug delivery systems like lipids, poly-
mers, dextrans, cyclodextrins, polyethylene gly-
col, etc. Excipients play a vital role in the
formulation development of both small and
macromolecule pharmaceutical products. The
type and level of excipients’ usage depends on
various factors like type of API, dosage form,
140 6. Biologics: Delivery options and formulation strategies
TABLE 6.6 Common chemical degradation pathway of amino acid sequences.
Reaction
Deamidation
Peptide bond hydrolysis
β-Elimination
Disulfide bond reshuffling
Oxidation
Thiol-disulfide exchange
Racemization
Hydrolysis
route of administration, indication, target popu-
lation, etc. As per the growing status of protein,
peptide therapeutics, and vaccines develop-
ment, the stabilization of biologics during pro-
cessing and storage poses a significant
challenge for product development scientists.
Biologics and vaccines are highly unstable in
nature and heavily prone to degradation by
physical and chemical (oxidation, hydrolysis,
deamination, photo-oxidation, and thio-
oxidation) mechanisms. Excipients play a signif-
icant role in the development of stable biologics
product and thereby the selection and use of
appropriate excipients is very important at the
product development stage. Excipients in bio-
logics products can be used for different func-
tionality, such as: (a) controlling pH and
tonicity; (b) enhancing the solubility of the active
molecule; (c) preventing its aggregation and
degradation; (d) enhancing the process and sta-
bility of the active molecule; (e) maintaining its
conformation; and (f ) other functions like anti-
oxidants, preservatives, bulking agents, etc.
The understanding of drug-excipient interac-
tions is also critical in the rational design of for-
mulations to stabilize protein-based therapeutic
drugs and vaccines. The complexity of formula-
tion development is often more pronounced for
the development of high-concentration anti-
body formulations due to additional constraints
of viscosity, analytical characterization at high
Corresponding amino acid involved
Asparagine and glutamine
Primarily at aspartic
Cystine
Cystine
Cystine and methionine
Cystine and cysteine
Aspartic acid and glycine
Aspartic acid, serine and threonine
concentration, and the desire to formulate an
isotonic formulation while maintaining the ratio
of excipient to antibody to provide the stability.
Commonly used excipients in biologics and vac-
cine formulation are summarized in Table 6.7.
Regulatory acceptance of excipients is also
critical for their selection and inclusion in the
final formulation. Regulatory agencies across
the globe, maintain information about approved
excipients which are already in use with various
commercial products. USFDA maintains the list
of approved excipients in a database (inactive
ingredient database; IID) with their dosage form,
route of administration and concentration. In
Japan, Japanese Pharmaceutical Excipients Dic-
tionary (JPED) provides compilation of all excip-
ients for which there is a precedence of use in
drug products, route of administration, and
patient exposure. It includes monographs from
the JP or Japanese Pharmaceutical Excipients
Council (JPEC) as well as all nonmonograph
excipients that have been previously used.
5.1 Surfactants
Surfactant are surface active agents, which
reduces surface tension of liquids or interfacial
tension between a multiple-phase system (usu-
ally a two-phase system) by adsorbing on the
surface/interface. The role of surfactants in the
 5 Formulation strategies, degradation routes, and role of excipients 141

TABLE 6.7 Commonly used excipients and their general range in biologics and vaccine formulations.
Category of excipients General range Examples

Buffers 5–100 mM Acetate, succinate, citrate, histidine

Amino acid – Arginine, aspartic acid, glutamic acid, lysine

Stabilizer/bulking agents 1%–10% Lactose, trehalose, dextrose, sucrose, sorbitol

Surfactants
Antimicrobial/preservative
Metals ions/Chelators
Cyclodextrine based
Salts
0.01%–0.1% (w/v) Tween 20, Tween 80, Plurenic F68
– Benzyl alcohol, m-cresol, phenol, 2-phenoxyethanol
– Ca, Zn, EDTA
– Hydrxypropyl β-cyclodextrine
0–300 M NaCl

protein-based formulation is to block the pro-
tein’s interaction with hydrophobic surfaces
(e.g., vial walls or air interfaces), which other-
wise can cause denaturation and aggregation
of proteins. Some biologics products need
surfactants to block the particular subregions
of the protein or to block the protein-protein
interaction that could lead to denaturation.
The surfactants like polysorbate (Polysorbate
80 or Polysorbate 20) and Poloxamer 188
(PO188) are mainly used in protein-based bio-
logics formulation. Even in the presence of these
surfactants, protein drug stability is often insuf-
ficient, particularly because of agitation-induced
aggregation. In recent reports, N-myristoyl
phenylalanine Jeffamide (FM1000) has been
proven to have better interfaces blocking capac-
ity and stabilizes faster than other conventional
surfactants.
5.2 Buffers
The role of buffers in formulation is to main-
tain the pH for optimal solubility and stability
of product during manufacturing, storage, and
reconstitution. The chemical stability of biologics
such as monoclonal antibodies is pH-dependent
and slightly acidic conditions are favorable
for stability of the formulation. The widely used
buffers in biologics products are acetate,
citrate, histidine, succinate, and phosphate
buffers. The self-buffering potential of proteins
becomes more relevant for high-concentration
biologics. Buffer capacity and the possibility of
buffer catalysis are crucial characteristics for
liquid biologics.
5.3 Lyoprotectors (sugar)/bulking agents
The bulking agents are required to provide
a matrix to carry the drug which are normally
present in low quantities. Generally, mannitol,
lactose sucrose, dextran, trehalose, and glycine
are used as bulking agents. Bulking agents
always play a dual role as filler and cryopro-
tectors in lyophilized products. These agents
have the potential to turn entire formulations
into a crystalline (mannitol and glycine) or
amorphous state (sucrose). An appropriate
selection of lyoprotector results in better prod-
uct quality (stability and moisture levels and
reconstitution time) and facilitates freeze-
drying and scaling up to commercial batch.
The concentration of bulking agents utilized
will determine the rationale use (as stabilizer,
eutectic temperature modifier or matrix-
forming agents).
142 6. Biologics: Delivery options and formulation strategies
Aqueous solution-based biologics may pose
stability issues because of hydrolytic degrada-
tion of active molecules. Lyophilization is a tech-
nique used to stabilize pharmaceutical products
by removing water from drug aqueous solution.
Lyophilization is the most frequently used strat-
egy to achieve the desired stability of a finished
product according to its target product profile. It
is also proven to be a superior technique for the
stabilization of thermolabile biologics, espe-
cially vaccines. Lyophilization consists of three
phases: freezing followed by primary and sec-
ondary drying under vacuum. During the freez-
ing step, protein denaturation can occur either in
the freeze-concentrate state or at frozen surface
interfaces. To counter the relevant criticality,
the formulation can be designed using salts
and buffer, which can minimize protein dena-
turation during the lyophilization process. Lyo-
protectant, along with cryoprotectant, is
normally needed for stabilization of biologics.
Lyoprotectant plays a vital role in protection of
the product during the drying phase while cryo-
protectant stabilizes the frozen product during
the freezing phase.
Bulking agents are also added to lyophilized
product to prevent product “blowout” espe-
cially in the case of low concentration product
( 1% solid). They are also useful in increasing
the product collapse temperature and/or to
improve product elegance (aesthetic appearance
of the cake—maintaining proper shape and lack-
ing irregularities). Sucrose, trehalose, lactose,
raffinose, dextran, and hydroxyethyl starch
(HES) are commonly used amorphous bulking
agents while glycine and mannitol are com-
monly used crystalline bulking agents. Excipi-
ents with high glass transition temperatures
(Tg0 ) for amorphous excipients or high eutectic
temperatures (Teu) for crystalline excipients
allow for faster primary drying. However, the
presence of the hydrate form of mannitol and
possibility of glass breakage during manufactur-
ing (due to high fill volume, incorrect freezing
protocol, and/or high concentration) might
limit the selection of mannitol during excipient
screening (Table 6.8).
5.4 Salts
The presence of salt in solution is required to
maintain the optimal ionic strength, which leads
to physical stability of the protein. Salts possess
properties to stabilize proteins by inhibiting
aggregation. Salts also minimize catalytic activ-
ity of phosphate which is present in certain pro-
tein/peptide products. Commonly used slats
are NaCl, LiCl, Tris HCl, etc.
5.5 Antimicrobial preservatives
Antimicrobial preservatives are mainly
required for multidose units to ensure the micro-
biological safety of a solution during use and
storage. The addition of preservatives in bio-
logics can trigger compatibility issues between
protein and antimicrobial agents that need to
be addressed by researchers during product
development activities. Some commonly used
preservatives are benzyl alcohol, metacresol,
phenol, etc.
5.6 Additional formulation considerations
5.6.1 Adjuvants in vaccine formulation
Vaccine formulations often contain an impor-
tant component that may improve the longevity,
breadth, magnitude, and intrinsic immunoge-
nicity to antigens with minimal toxicity and last-
ing immune effects on their own. Strategic
formulations of agonists of the innate immune
system and carriers that selectively present at
the target site of antigen evolve a class of phar-
maceutical “adjuvants,” which significantly
impact immunity resulting from vaccination.
The effects of adjuvants in vaccines could reduce
both the amount of antigen and/or number of
immunizations required to achieve the efficacy.
TABLE 6.8 Various excipients and their role in biologics formulation depicting typical ranges used for different routes of administrations.
Route of Date
Excipients (category) Role Product admin. Concentration approved

Buffer
Sodium phosphate Buffering agent Reopro IV bolus 0.01 M 12/22/1994
(U.S.)
Dibasic sodium phosphate anhydrous Buffering agent Herceptin IV bolus 0.335 mg 09/25/1998
(U.S.)
Monobasic sodium phosphate monohydrate Buffering agent Orencia SC 0.114 mg 12/23/2005
(U.S.)
Monobasic sodium phosphate Buffering agent Remicade IV infusion 2.2 mg 08/24/1998
(U.S.)
Dibasic sodium phosphate Buffering agent Remicade IV infusion 6.1 mg 08/24/1998
(U.S.)

Sodium phosphate, monobasic, monohydrate Buffering agent Tysabri IV infusion 123 mg 11/23/2004
(U.S.)

Sodium phosphate, dibasic, heptahydrate Buffering agent Tysabri IV infusion 17 mg 11/23/2004

(U.S.)
Zinc acetate Buffering agent Nutropin depot IV infusion 1.2 mg 12/22/1999
(U.S.)
Zinc carbonate Buffering agent Nutropin depot IV infusion 0.8 mg 12/22/1999
(U.S.)
Acetate Buffering agent Udenyca (pegfilgrastim) SC 0.35 mg 11/02/2018
(U.S.)
Acetate Buffering agent Nivestym (filgrastim) IV infusion 0.59 mg 07/20/2018
(U.S.)
Acetate Buffering agent Fulphila (pegfilgrastim) SC 0.7 mg 06/04/2018
(U.S.)
Monobasic sodium phosphate Buffering agent Remicade IV infusion 2.2 mg 08/24/1998
(U.S.)
Dibasic sodium phosphate Buffering agent Remicade IV infusion 6.1 mg 08/24/1998
(U.S.)

Continued
TABLE 6.8 Various excipients and their role in biologics formulation depicting typical ranges used for different routes of administrations—cont’d
Route of Date
Excipients (category) Role Product admin. Concentration approved

Sodium phosphate, monobasic, monohydrate Buffering agent Tysabri IV infusion 123 mg 11/23/2004
(U.S.)

Sodium phosphate, dibasic, heptahydrate Buffering agent Tysabri IV infusion 17 mg 11/23/2004

(U.S.)

Sodium phosphate monobasic monohydrate Buffering agent Retacrit (epoetin alfa) IV 1.3 mg 05/15/2018
(U.S.)
Threonine Buffering agent Retacrit (epoetin alfa) IV 0.25 mg 05/15/2018
(U.S.)
Disodium succinate hexahydrate Buffering agent Ixifi (infliximab) IV infusion 12.1 mg 12/13/2017
(U.S.)
Sodium phosphate (monobasic, monohydrate) Buffering agent Mvasi (bevacizumab) IV infusion 23.2 mg 09/14/2017
(U.S.)
Sodium phosphate (dibasic, anhydrous) Buffering agent Mvasi (bevacizumab) IV Infusion 4.8 mg 09/14/2017
(U.S.)
Monobasic sodium phosphate monohydrate Buffering agent Renflexis (infliximab) IV infusion 5.55 mg 04/21/201
(U.S.)
Sodium citrate Buffering agent Erelzi (etanercept) SC 13.52 mg 08/30/2016
(U.S.)
Sodium dihydrogen phosphate monohydrate Buffering agent Inflectra (infliximab) IV infusion 2.2 mg 04/05/2016
(U.S.)
Potassium dihydrogen phosphate Buffering agent Shingrix (zoster vaccine IM 0.54 mg 10/20/2017
recombinant, adjuvanted) (U.S.)

Sodium dihydrogen phosphate dihydrate Buffering agent Shingrix (zoster vaccine IM 0.160 mg 10/20/2017
recombinant, adjuvanted) (U.S.)

Disodium phosphate anhydrous Buffering agent Shingrix (zoster vaccine IM 0.15 mg 10/20/2017
recombinant, adjuvanted) (U.S.)
Sodium phosphate Buffering agent Luxturna (voretigene neparvovec- Intra retinal 10 mM 12/19/2017
rzyl) (U.S.)
Sodium phosphate, dibasic dodecahydrate Buffering agent Heplisav-b (hepatitis b vaccine IM 1.75 mg/mL 11/09/2017
recombinant) (U.S.)
Disodium phosphate anhydrous Buffering agent Shingrix (zoster vaccine IM 0.15 mg 10/20/2017
recombinant, adjuvanted) (U.S.)
Sodium phosphate Buffering agent Luxturna (voretigene neparvovec) Intra retinal 10 mM 12/19/2017
(U.S.)
Sodium phosphate, dibasic dodecahydrate Buffering agent Heplisav-b (hepatitis b vaccine IM 1.75 mg/mL 11/09/2017
recombinan) (U.S.)
Sodium phosphate, monobasic dihydrate Buffering agent Heplisav-b (hepatitis b vaccine IM 0.48 mg/mL 11/09/2017
(recombinant)
Polylactide-coglycolide (PLG) Controlled release Nutropin depot IV infusion 68.9 mg 12/22/1999
polymer (U.S.)
Salts
Sodium chloride Isotonicity Reopro IV 0.15 M 12/22/1994
(U.S.)
Sodium chloride Isotonicity Enbrel SC 120 Mm 11/02/1998
(U.S.)
Sodium chloride Isotonicity Truxima (rituximab) IV infusion 9 mg 11/28/2018
(U.S.)
Sodium Isotonicity Udenyca (pegfilgrastim) SC 0.02 mg 10/30/2018
(U.S.)
Sodium chloride Isotonicity Nivestym (filgrastimaafi) IV infusion 4.93 mg 07/20/2018
(U.S.)
Sodium chloride Isotonicity Erelzi (etanercept) SC 1.5 mg 08/30/2016
(U.S.)

Sodium chloride Isotonicity Shingrix (zoster vaccine IM 4.385 mg 10/20/2017

recombinant, adjuvanted) (U.S.)
Sodium chloride Isotonicity Luxturna (voretigene neparvovec) Intra retinal 180 Mm 12/19/2017
(U.S.)
Sodium chloride Isotonicity Heplisav-b (hepatitis b vaccine IM 9.0 mg/mL 11/09/2017
recombinant) (U.S.)
Sodium chloride Isotonicity Synagis (palivizumab) IM 0.9 mg 06/19/1998
(U.S.)
Potassium chloride Isotonicity Lemtrada (alemtuzumab) IV infusion 0.2 mg

Continued
TABLE 6.8 Various excipients and their role in biologics formulation depicting typical ranges used for different routes of administrations—cont’d
Route of Date
Excipients (category) Role Product admin. Concentration approved

05/07/2001
(U.S.)

Sodium chloride Isotonicity Zevalin (ibritumomab tiuxetan) IV infusion 0.9% 02/19/2002

(U.S.)
Sodium chloride Isotonicity Erbituxtm (cetuximab) IV infusion 8.48 mg/mL 02/12/2004
(U.S.)
Stabilizer
Mannitol Stabilizer Humira SC 16.8 mg 12/31/2002
(U.S.)
Sucrose Stabilizer Cimzia SC 100 mg 04/22/2008
(U.S.)
Sorbital Stabilizer Simponi SC 20.5 mg 04/24/2009
(U.S.)
Sucrose Stabilizer Enbrel SC 1% 11/02/1998
(U.S.)

Α,Α-trehalose Stabilizer Herceptin IV bolus 381.8 mg 09/25/1998

(U.S.)

Dihydrate Stabilizer Herceptin IV bolus 381.8 mg 09/25/1998

(U.S.)
Sucrose Stabilizer Orencia SC 68 mg 12/23/2005
(U.S.)
Sucrose Stabilizer Remicade IV infusion 500 mg 08/24/1998
(U.S.)
Α,Α-trehalose Stabilizer Herzuma (trastuzumab) IV infusion 839 mg 12/14/2018
(U.S.)
Mannitol Stabilizer Hyrimoz (adalimumab) SC 9.6 mg 10/30/2018
(U.S.)
Sorbitol Stabilizer Nivestym (filgrastimaafi) IV infusion 50 mg 07/20/2018
(U.S.)
D-Sorbitol Stabilizer Fulphila (pegfilgrastim) SC 30 mg 06/04/2018
(U.S.)
Sucrose Stabilizer Ixifi (infliximab) IV infusion 250 mg 12/13/2017
(U.S.)
D-Sorbitol Stabilizer Ogivri (trastuzumab) IV fusion 322.6 mg 12/01/2017
(U.S.)
Trehalose dihydrate Stabilizer Mvasi (bevacizumab) IV infusion 240 mg 09/14/2017
Sucrose Stabilizer Renflexis (infliximab) IV infusion 500 mg 04/21/2017
(U.S.)
Sucrose Stabilizer Amjevita (adalimumab) SC 36 mg 09/23/2016
(U.S.)
Sorbitol Stabilizer Zarxio (filgrastim) SC 25 mg 03/06/2015
(U.S.)
Sucrose Stabilizer Inflectra (infliximab) IV infusion 500 mg 04/05/2016
(U.S.)
Sucrose Stabilizer Shingrix (zoster vaccine I M injection 20 mg 10/20/2017
recombinant, adjuvanted) (U.S.)
Sucrose Stabilizer Simulect (basiliximab) IV 10 mg 05/12/1998
(U.S.)
Mannitol Stabilizer Simulect (basiliximab) IV 40 mg 05/12/1998
(U.S.)
Sucrose Stabilizer Andexxa [coagulation factor Xa IV 2% w/v 05/03/2018
(recombinant)] (U.S.)

Mannitol Stabilizer Andexxa [coagulation factor Xa IV 5% w/v 05/03/2018

(recombinant)] (U.S.)
Dextran-based excipients

Dextran 40 Priming fluid Mylotarg (gemtuzumab ozogamicin) IV bolus 41.0 mg 05/17/2000

(U.S.)

Surfactant
Polysorbate 80 Solubilizing agent Reopro IV bolus 0.001% 12/22/1994
(U.S.)

Polysorbate 80 Solubilizing agent Humira SC 0.4 mg 12/31/2002

(U.S.)

Continued
TABLE 6.8 Various excipients and their role in biologics formulation depicting typical ranges used for different routes of administrations—cont’d
Route of Date
Excipients (category) Role Product admin. Concentration approved

Polysorbate Solubilizing agent Cimzia SC 0.1 mg 04/22/2008

(U.S.)

Polysorbate 80 Solubilizing agent Simponi SC 0.08 mg 04/24/2009

(U.S.)

Poloxamer 188 Solubilizing agent Orencia SC 3.2 mg 12/23/2005

(U.S.)
Polysorbate 20 Solubilizing agent Herceptin IV bolus 1.7 mg 09/25/1998
(U.S.)
Polysorbate 80 Solubilizing agent Cathflo activase IV bolus 0.2 mg 11/13/1987
(U.S.)
Polysorbate 80 Solubilizing agent Tysabri IV infusion 7.24 mg 11/23/2004
(U.S.)
Polysorbate 80 Solubilizing agent Remicade IV infusion 0.5 mg 08/24/1998
(U.S.)
Polysorbate 20 Solubilizing agent Herzuma (trastuzumab) Solubilizing 1.7 mg 12/14/2018
agent (U.S.)
Polysorbate 80 Solubilizing agent Truxima (rituximab) IV infusion 0.7 mg 11/28/2018
(U.S.)
Polysorbate 20 Solubilizing agent Udenyca (pegfilgrastim) SC 0.02 mg 11/02/2018
(U.S.)
Polysorbate 80 Solubilizing agent Nivestym (filgrastim) IV infusion 0.04 mg 07/20/2018
(U.S.)

Polysorbate 80 Solubilizing agent Ixifi (infliximab) IV infusion 0.5 mg 12/13/2017

(U.S.)

Polysorbate 20 Solubilizing agent Mvasi (bevacizumab) IV infusion 1.6 mg 09/14/2017

(U.S.)
Sodium acetate trihydrate Solubilizing agent Cyltezo (adalimumab) SC 2.4 mg 08/25/2017
(U.S.)
Polysorbate 80 Solubilizing agent Amjevita (adalimumab) SC 0.4 mg 09/23/2016
(U.S.)
Polysorbate 80 Solubilizing agent Zarxio (filgrastim) SC 0.02 mg 03/06/2015
(U.S.)
Polysorbate 80 Solubilizing agent Inflectra (infliximab) IV infusion 0.5 mg 04/05/2016
(U.S.)
Polysorbate 80 Solubilizing agent Shingrix (zoster vaccine IM 0.08 mg 10/20/2017
recombinant, adjuvanted) (U.S.)
Poloxamer 188 Solubilizing agent Luxturna (voretigene neparvovec- Intra retinal 0.001% 12/19/2017
rzyl) (U.S.)
Polysorbate 80 Solubilizing agent Heplisav-b [hepatitis b vaccine IM 0.1 mg/mL 11/09/2017
(recombinant) (U.S.)
Polysorbate 80 Solubilizing agent Rituxan (rituximab) IV infusion 0.7 mg 11/26/1997
(U.S.)
Polysorbate 80 Solubilizing agent Lemtrada (alemtuzumab) IV infusion 0.1 mg 05/07/2001
(U.S.)
Polysorbate 80 Solubilizing agent Andexxa (coagulation factor xa IV 0.01% w/v 05/03/2018
(recombinant) (U.S.)

Polysorbate 80 Solubilizing agent Kineret (anakinra) SC 0.70 mg 11/14/2001

(U.S.)

Polysorbate 20 Solubilizing agent Neulasta (pegfilgrastim) SC 0.02 mg 01/31/2002

Antimicrobial/preservative
Benzyl alcohol Preservative Enbrel SC 0.9% 11/02/1998
(U.S.)
Glacial acetic acid Preservative Amjevita (adalimumab-atto) SC 0.24 mg 09/23/2016
(U.S.)

Metacresol Preservative Humulin (insulin human) SC 2.5 Mg 10/28/1982

(U.S.)

Continued
TABLE 6.8 Various excipients and their role in biologics formulation depicting typical ranges used for different routes of administrations—cont’d
Route of Date
Excipients (category) Role Product admin. Concentration approved

Metacresol Preservative Humalog (insulin lispro injection) SC 3.15 Mg 06/14/1996

(U.S.)

m-Cresol Preservative Lantus (insulin glargine injection) SC 2.7 mg 04/20/2000

Metacresol Preservative Novolog (insulin aspart injection) SC 1.72 mg/mL 06/07/2000
(U.S.)
Phenol Preservative Novolog (insulin aspart injection) SC 1.50 mg/mL 06/07/2000
(U.S.)

Glycerin Solvent Humulin (insulin human) SC 16 mg 10/28/1982

(U.S.)

Glycerin Solvent Humalog (insulin lispro injection) SC 16 mg 06/14/1996

(U.S.)
Calcium chloride dihydrate Retacrit (epoetin alfa-epbx) IV 0.01 mg 05/15/2018
(U.S.)
Glycine Solvent Retacrit (epoetin alfa-epbx) IV 7.5 mg 05/15/2018
(U.S.)
Glycine Solvent Simulect (basilixiroab) IV 20 mg 05/12/1998
(U.S.)
Glycine Solvent Synagis (palivizumab) IM 0.1 mg 06/19/1998
(U.S.)
Glycerol 85% Solvent Lantus (insulin glargine injection) SC 20 mg 04/20/2000
Glycerin Solvent Novolog (insulin aspart injection) SC 16 mg/mL 06/07/2000
Amino acid and small acids

L-Histidine
and L-histidine Antioxidant Simponi SC 0.44 Mg 04/24/2009
monohydrochloride monohydrate (U.S.)
L-Arginine hydrochloride Antioxidant Enbrel SC 25 Mm 11/02/1998
(U.S.)
L-Histidine Hcl monohydrate Antioxidant Herceptin IV bolus 9.5 Mg 09/25/1998
(U.S.)
L-Histidine Antioxidant Herceptin IV bolus 6.1 Mg 09/25/1998
(U.S.)
L-Arginine Antioxidant Cathflo IV bolus 77 Mg 11/13/1987
Activase (U.S.)
L-Histidine Hcl Antioxidant Herzuma (trastuzumab) IV infusion 9.5 Mg 12/14/2018
(U.S.)
Adipic acid Antioxidant Hyrimoz (adalimumab) SC 2.69 Mg 10/30/2018
(U.S.)
Leucine Antioxidant Retacrit (epoetin alfa) IV 1 Mg 05/15/2018
(U.S.)
L-Glutamic acid Antioxidant Retacrit (epoetin alfa) IV 0.25 Mg 05/15/2018
(U.S.)
Phenylalanine Antioxidant Retacrit (epoetin alfa) IV 0.5 mg 05/15/2018
(U.S.)
Succinic acid Antioxidant Ixifi (infliximab) IV infusion 0.6 mg 12/13/2017
(U.S.)
L-Histidine hydrochloride monohydrate Antioxidant Ogivri (trastuzumab) IV infusion 9.4 mg 12/01/2017
(U.S.)

Glacial acetic acid Antioxidant Cyltezo (adalimumab) SC 0.13 mg 08/25/2017

(U.S.)

Phenylalanine Antioxidant Retacrit (epoetin alfa) IV 0.5 mg 05/15/2018

(U.S.)
Succinic acid Antioxidant Ixifi (infliximab) IV infusion 0.6 mg 12/13/2017
(U.S.)
L-Histidine hydrochloride monohydrate Antioxidant Ogivri (trastuzumab) IV infusion 9.4 mg 12/01/2017
(U.S.)
Citric acid Antioxidant Kineret (anakinra) SC 1.29 mg 11/14/2001
(U.S.)
Lactic acid Antioxidant Cimzia SC 0.9 mg 04/22/2008
(U.S.)
Citric acid Antioxidant Erelzi (etanercept) SC 0.786 mg 08/30/2016
(U.S.)
Continued
TABLE 6.8 Various excipients and their role in biologics formulation depicting typical ranges used for different routes of administrations—cont’d
Route of Date
Excipients (category) Role Product admin. Concentration approved

Metal ions/chelators/others
Zinc oxide – Humulin (insulin human) SC 0.017 mg 10/28/1982
(U.S.)
Zinc ion – Humalog (insulin lispro injection) SC 0.046 mg 06/14/1996
(U.S.)
Disodium edetate dihydrate – Lemtrada (alemtuzumab) IV infusion 0.0187 mg 05/07/2001
(U.S.)
Disodium EDTA – Kineret (anakinra) SC 0.12 mg 11/14/2001
(U.S.)
Zinc – Lantus (insulin glargine injection) SC 30 μg 04/20/2000
(U.S.)

Zinc – Novolog (insulin aspart injection) SC 19.6 μg/mL 06/07/2000

(U.S.)

Polyethylene glycol Vehicle Ogivri (trastuzumab) IV infusion 94.1 mg 12/01/2017

(U.S.)
Macrogol 3350 Vehicle Ogivri (trastuzumab) IV infusion NA 12/01/2017
(U.S.)
Cholesterol Emulsifying agent Shingrix (zoster vaccine IM 0.25 mg 10/20/2017
recombinant, adjuvanted) (U.S.)
 References
The most commonly used adjuvant formula-
tions include aluminum hydroxide and alumi-
num phosphate for human use because of
their historic safety and efficacy profiles. For
the selection of adjuvants, it is important to
understand the interaction between antigen-
adjuvant and their impact on stability and
immunogenesity. While alum-based adjuvants
are very traditional, quite successful, and very
commonly used, newer adjuvant mechanisms
are being explored, including delivery mecha-
nisms like liposome/virosome-based/inspired
systems.
5.6.2 Stability of biologics, typical shelf life
and storage considerations
Biologics are fragile in nature especially when
compared to small molecules. One of the main
challenges is their stability, mainly in liquid
form. Because of their complex nature, they
are prone to several routes of degradation—
most of the time, more than one degradation
pathway is observed. Typically, biopharmaceu-
ticals are stored and maintained under refriger-
ated conditions, most of them at 5  3 °C. The
liquid forms typically have a shelf life of
24 months while the lyophilized form has a shelf
life of 36–48 months. More and more ready to
use options (e.g., prefilled syringes, autoinjec-
tors) are becoming available, which pose addi-
tional challenges to secure stability during the
shelf life of the product.
5.6.3 Toxicity and immunogenicity of
biologics
Over the journey of biopharmaceutical devel-
opment and especially for monoclonal anti-
bodies, the trend over the last 40 years has been
to get as close to the human proteins sequence
as possible. At first, biopharmaceuticals had
sequences coming from animal origins (e.g.,
mouse), but slowly the sequences have started
to become more humanized and now a large
153
majority are fully human. Newer formats and
sequences are being developed as well, but their
development does involve consideration on
sequences already present/exposed to humans.
The main reason for getting the protein sequence
close to that of humans is to minimize any risk of
immunogenicity, which is the main reason for
toxicities of biopharmaceuticals.
6 Conclusion
Biologics are an important class of molecules
and they continue to gain in significance. Cur-
rently the options to deliver them are largely
limited to intravenous and subcutaneous injec-
tions. While the existing systems have been
employed for several years, there is a need to
understand the operational mechanistics for
these routes, especially for subcutaneous admin-
istration. Improving their efficiency while push-
ing the existing limits will help clinicians on a
broad level. On the other hand, there is a need
to develop drug delivery options for localized
application if not systematic delivery. Clinically
meaningful efforts are being attempted which
hold promise for the future; multiple studies
and different approaches are being developed.
Having these delivery options will open new
therapeutic possibilities with patient ease and
comfort at the center.
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