Correspondence
normal prenatal diagnosis. We received no no-
tice from any delivering physician or neonatolo-
gist of the birth of an affected infant. Because
there were no known infants with trisomy 13 or
18, we calculated a negative predictive value of
100%. One baby was born with trisomy 21, which
resulted in a negative predictive value of greater
than 99.9%.
CharlesM. Strom, M.D., Ph.D.
MeganD. Maxwell, M.D.
Renius Owen, Ph.D.
Quest Diagnostics Nichols Institute
San Juan Capistrano, CA
charles.m.strom@questdiagnostics.com
TARGT Gene Therapy Platform for Correction
of Anemia in End-Stage Renal Disease
To the Editor: Anemia in patients with end-
stage renal disease who are undergoing hemodi-
alysis is caused, in part, by a reduction in or
absence of secretion of endogenous erythropoi-
etin. Recombinant human erythropoietin given
intravenously has been associated with an in-
creased risk of cardiovascular complications and
death, which is considered to be related, at least
in part, to the high peak levels of plasma eryth-
ropoietin that are achieved after intravenous
administration.1,2 Transduced autologous restor-
ative gene therapy (TARGT), a new autologous
protein-delivery platform designed to provide
sustained secretion of proteins such as recombi-
nant human erythropoietin, uses a small autolo-
gous dermal explant or explants (see Fig. S1 in
Supplementary Appendix, available with the full
text of this letter at NEJM.org) in which several
biopsy samples of a given patients dermis are
transduced ex vivo with the use of a helper-de-
pendent adenovirus system that is engineered to
carry a gene in this case EPO, which encodes
the human erythropoietin protein. The resulting
delivery system is referred to as TARGTEPO.
We performed studies of TARGTEPO in vitro
and in vivo (in SCID [severe combined immuno-
deficient] mice) and measured the secretion of
erythropoietin. By assessing the rate of erythro-
poietin secretion into the media and the amount
of erythropoietin secreted, we estimated the
dose of recombinant erythropoietin that would
n engl j med 376;2nejm.org January 12, 2017
The New England Journal of Medicine
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Copyright 2017 Massachusetts Medical Society. All rights reserved.
Supported by Quest Diagnostics, which provided salary sup-
port and editorial assistance in the preparation of this letter.
Disclosure forms provided by the authors are available with
the full text of this letter at NEJM.org.
1. Bianchi DW, Parker RL, Wentworth J, et al. DNA sequencing
versus standard prenatal aneuploidy screening. N Engl J Med
2014;370:799-808.
2. Wang JC, Sahoo T, Schonberg S, et al. Discordant noninva-
sive prenatal testing and cytogenetic results: a study of 109 con-
secutive cases. Genet Med 2015;17:234-6.
3. Snyder MW, Simmons LE, Kitzman JO, et al. Copy-number
variation and false positive prenatal aneuploidy screening re-
sults. N Engl J Med 2015;372:1639-45.
DOI: 10.1056/NEJMc1604205
typically be needed for each patient. We then
washed the TARGTEPO units extensively to remove
residual culture media and implanted the appro-
priate number subcutaneously into each patient.3
We found that erythropoietin secreted from
TARGTEPO units had an isoform pattern similar
to that of secreted endogenous erythropoietin.
After we performed the in vitro and in vivo
studies to determine individual initial doses,
we administered TARGTEPO in a phase 12, open-
label, multicenter, single-group, dose-escalat-
ing clinical study (ClinicalTrials.gov number,
NCT02117427) to treat anemia in 10 patients with
end-stage renal disease who were undergoing
hemodialysis. Patients were assigned to one of
two dose groups low dose (18 to 25 IU per
kilogram of body weight per day, 6 patients) and
higher dose (35 to 45 IU per kilogram per day,
4 patients). The primary outcomes were clinical
safety, pharmacokinetic properties, and time
within the target hemoglobin range (9 to 12 g
per deciliter) in accordance with clinical guide-
lines for patients with end-stage renal disease.
The study protocol is available at NEJM.org.
All patients who underwent TARGTEPO im-
plantation had an initial increase in plasma
levels of erythropoietin, and plasma levels of
hemoglobin remained above or within the de-
sired range for most patients (Fig. 1). The plas-
ma levels of erythropoietin remained stable
within the physiologic range. In 7 of the 10 pa-
 The n e w e ng l a n d j o u r na l of m e dic i n e

A
35
Patient 1

Mean Plasma Erythropoietin

30 Patient 2
25 Patient 3
Patient 4
(mIU/ml) 20
Patient 5
15 Patient 6
Patient 8
10
Patient 9
5 Patient 10
0
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19
Months after Implantation

B
14
Mean Plasma Hemoglobin (g/dl)

Patient 1
13 Patient 2
Patient 3
12
Patient 4
11 Patient 5
Patient 6
10 Patient 8
Patient 9
9
Patient 10
0
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19
Months after Implantation

Figure 1. Levels of Erythropoietin and Hemoglobin after Implantation in Patients with End-Stage Renal Disease.
Shown is the monthly plasma concentration of erythropoietin (Panel A) and hemoglobin (Panel B) after TARGTEPO
implantation in 9 of the 10 patients (the erythropoietin concentration in Patient 7 stayed above baseline for only 2
weeks). The gray area in Panel B indicates the desired hemoglobin range according to clinical guidelines4 (the darker
gray area indicates the most desirable range; the lighter gray area [between 11 and 12 g per deciliter] indicates an
adequate range that does not require intervention).

tients who underwent implantation, erythropoi- tions of TARGTs excised from patients or assays
etin and hemoglobin levels were maintained in of antierythropoietin antibodies, of immunoge-
the desired range for a period of at least 5 months nicity to either TARGT or the erythropoietin se-
without the need for injections of recombinant creted from the TARGTEPO.
human erythropoietin. These sustained levels of This early study does not allow us to assess
erythropoietin and hemoglobin after TARGTEPO the risk or benefit of TARGTEPO as compared
implantation were achieved with a maximum with administration of recombinant erythropoi-
plasma level of erythropoietin that was approxi- etin injections. Such an assessment would re-
mately one hundredth the maximum level mea- quire a prospective, randomized trial.
sured in these patients when they received re-
Shany Blum, M.D., Ph.D.
combinant human erythropoietin by injection
Nir Shapir, Ph.D.
during the run-in phase of the study, and with
Reem Miari, M.Sc.
exposure to erythropoietin (measured as the
Benny Lerner, B.Sc.
area under the concentrationtime curve) that
was one tenth that obtained during administration Belly Koren, M.Sc.
of recombinant human erythropoietin by injection. Medgenics Medical Israel
Misgav, Israel
There were no serious adverse events related to blum.shany@gmail.com
TARGTEPO, and no signs, on either histologic sec-

n engl j med 376;2 nejm.org January 12, 2017

The New England Journal of Medicine

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Copyright 2017 Massachusetts Medical Society. All rights reserved.
 Correspondence
KerenDoenyas-Barak,M.D.
ShaiEfrati,M.D.
Assaf-Harofeh Medical Center
Zerifin, Israel
Pablo E.Pergola,M.D.
Renal Associates
San Antonio, TX
DoronSchwartz,M.D.
GilChernin,M.D.
Tel Aviv Medical Center
Tel Aviv, Israel
YoramYagil,M.D., Ph.D.
Barzilai University Medical Center Campus
Ashkelon, Israel
SergeGuzy,Ph.D.
POP-PHARM Pharmacometric Services
Albany, CA
AbrahamNyska,Ph.D.
Sackler School of Medicine
Tel Aviv, Israel
Preserved Renal-Allograft Function and the PD-1 Pathway
Inhibitor Nivolumab
To the Editor: Inhibition of immune check-
points with the use of antibodies targeting pro-
grammed cell death 1 (PD-1) or monoclonal anti-
bodies against cytotoxic T-lymphocyteassociated
antigen 4 (CTLA-4) has been used clinically in
patients with various types of cancer. In the
limited number of reported cases in which these
antibodies have been used in patients who have
undergone kidney transplantation,1 these agents
have been associated with cell-mediated and
antibody-mediated rejection (see Table S2 in the
Supplementary Appendix, available with the full
text of this letter at NEJM.org).
We report on a patient who received a renal
transplant from a living related donor. In this
patient, a regimen of a preemptive glucocorti-
coid and sirolimus (a mammalian target of ra-
pamycin [mTOR] inhibitor) may have prevented
the adverse immune response of nivolumab in
the kidney transplant.
A 70-year-old man with end-stage kidney dis-
ease after bilateral nephrectomies for renal-cell
cancer underwent a kidney transplantation in
2010, with one of six HLA mismatches between
n engl j med 376;2nejm.org January 12, 2017
The New England Journal of Medicine
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Copyright 2017 Massachusetts Medical Society. All rights reserved.
Garry A.Neil,M.D.
Medgenics
Wayne, PA
Supported in part by a grant from the Israel Chief Scientist
Office (programs 52026 and 54586) to Medgenics Medical Israel
and by Medgenics Medical Israel funds.
Disclosure forms provided by the authors are available with
the full text of this letter at NEJM.org.
1. Koulouridis I, Alfayez M, Trikalinos TA, Balk EM, Jaber BL.
Dose of erythropoiesis-stimulating agents and adverse out-
comes in CKD: a metaregression analysis. Am J Kidney Dis
2013;61:44-56.
2. Besarab A, Bolton WK, Browne JK, et al. The effects of nor-
mal as compared with low hematocrit values in patients with
cardiac disease who are receiving hemodialysis and epoetin.
N Engl J Med 1998;339:584-90.
3. Shapir N, Miari R, Blum S, et al. Preclinical and preliminary
clinical evaluation of genetically transduced dermal tissue im-
plants for the sustained secretion of erythropoietin and inter-
feron . Hum Gene Ther Clin Dev 2015;26:216-27.
4. Kidney Disease: Improving Global Outcomes (KDIGO) Ane-
mia Work Group. KDIGO clinical practice guideline for anemia
in chronic kidney disease. Kidney Int Suppl 2012;2:279-335.
DOI: 10.1056/NEJMc1606202
the recipient and the graft. The patient received
basiliximab (an antiinterleukin-2 receptor anti-
body) and glucocorticoid induction followed by
initial immunosuppression that included a gluco-
corticoid, tacrolimus, and mycophenolate mofetil.
In early 2015, the patient received a diagnosis
of microsatellite-stable metastatic adenocarci-
noma of the duodenum with intestinal obstruc-
Table 1. Immunosuppressive Regimen in a Patient Who Had Undergone Kidney
Transplantation.
Timing* Drug and Dosage
1 Wk before Prednisone 40 mg daily
Concurrent Prednisone 20 mg daily; sirolimus target goal,
46 ng per milliliter
1 Wk after Prednisone 20 mg
>2 Wk and 6 mo after Prednisone 10 mg/day; sirolimus target goal,
1012 ng per milliliter
>6 Mo after Glucocorticoid gradually decreased to 5 mg/day;
sirolimus continued to maintain goal of
1012 ng per milliliter
* Timing represents the initiation of the immunosuppressive regimen in relation
to the administration of nivolumab.