1. No category

From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
1991 77: 792-798
Loss of interferon antibodies during prolonged continuous interferonalpha 2a therapy in hairy cell leukemia
RG Steis, JW 2d Smith, WJ Urba, DJ Venzon, DL Longo, R Barney, LM Evans, LM Itri and CH
Ewel
Updated information and services can be found at:
http://www.bloodjournal.org/content/77/4/792.full.html
Articles on similar topics can be found in the following Blood collections
Information about reproducing this article in parts or in its entirety may be found online at:
http://www.bloodjournal.org/site/misc/rights.xhtml#repub_requests
Information about ordering reprints may be found online at:
http://www.bloodjournal.org/site/misc/rights.xhtml#reprints
Information about subscriptions and ASH membership may be found online at:
http://www.bloodjournal.org/site/subscriptions/index.xhtml
Blood (print ISSN 0006-4971, online ISSN 1528-0020), is published weekly by the American
Society of Hematology, 2021 L St, NW, Suite 900, Washington DC 20036.
Copyright 2011 by The American Society of Hematology; all rights reserved.
From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
Loss of Interferon Antibodies During Prolonged Continuous Interferon-a2a
Therapy in Hairy Cell Leukemia
By Ronald G. Steis, John W. Smith II, Walter J. Urba, David J. Venzon, Dan L. Longo, Ruth Barney, Lydia M. Evans,
Loretta M. Itri. and Cynthia H. Ewe1
Although highly active in hairy cell leukemia (HCL), interferons (IFN) are not curative in this disease; current data
indicate that prolonged IFN therapy will be necessary t o
control disease in the majority of patients. We previously
observed acquired IFN resistance in association with neutralizing IFN-a2a antibodies in small numbers of patients with
HCL. This finding suggests that the requisite long-term
therapy may be compromised if there is an increasing incidence over time of neutralizing antibodies. We performed a
follow-up study of IFN antibodies in our patients receiving
continuous IFN therapy. All 16 patients who were previously
antibody negative remained so. Surprisingly, all nine patients
who previously had non-neutralizing IFN antibodies became
antibody negative after a median of 14.5 months. Moreover,
3 of 10 patients who had neutralizing antibodies became
antibody negative and five had only non-neutralizing antibod-
ies a median of 10 months from the time neutralizing
antibody had first been detected. Only two patients had
persisting neutralizing antibodies. Inhibition of neopterin
synthesis, inhibition of generation of 2‘, 5’ oligoadenylate
synthetase activity, and inability t o detect IFN in serum after
subcutaneous injection of IFN-a2a was observed only in the
one patient tested with neutralizing IFN antibodies confirming that these antibodies have functional significance in
vivo. We conclude that, although neutralizing IFN antibodies
inhibit the effectiveness of IFN in vivo, these antibodies are
produced only transiently during long-term therapy. The
long-term effectiveness of this drug will not likely be affected
in most patients by neutralizing antibody.
This is a US government work. There are no restrictions on
its use.
I
if not all, patients will require repetitive courses of therapy
or chronic maintenance therapy to maintain normal peripheral blood (PB) counts.
We have been studying the long-term effectiveness of
IFN in 56 patients with HCL. In contrast to other studies in
which there was a predetermined duration of IFN administration and in light of inevitable disease progression after
cessation of therapy, we opted to administer IFN continuously until either disease progression or unacceptable toxic
effects developed. The results of this study will be published
separately.8During the course of this study, we observed a
31% incidence of IFN-neutralizing antibody formation and
the development of acquired IFN resistance in approximately one-third of the patients who had neutralizing
antibodies? The antibodies were specific for IFN-a2a;
there was no cross-reactivity with partially purified natural
IFN. Resistance to the antitumor effects of IFN was not
observed in the absence of neutralizing antibodies. Because
prolonged use of IFN will be necessary for the long-term
management of this disease, the development of neutralizing IFN antibodies and IFN resistance is a potentially
significant problem that may affect an increasing proportion of patients over time. To further study this issue and to
study possible alternative therapies for patients who develop antibody-related IFN resistance, we determined the
effect of IFN antibodies on IFN-induced changes in serum
neopterin levels and PB mononuclear cell Z‘, 5‘ oligoadenylate synthetase activity. Such analyses were performed
following doses of either IFN-a2a (the IFN species these
patients had been receiving) or human lymphoblastoid IFN
to investigate the specificity of neutralizing antibodies in
vivo. We also reevaluated serum specimens from our
patients late in the course of IFN therapy for the presence
of IFN antibodies.
NTERFERON a (IFNa) has been used as an effective
agent in the treatment of hairy cell leukemia (HCL)
since its activity in this disease was established by Quesada
et a1 in 1983.’ A number of
have shown that the
majority of patients will have a clinically meaningful improvement in peripheral blood counts with IFN doses that
have minimal toxic effects. This therapy has become the
treatment of choice for HCL patients who progress after
splenectomy and is indicated for that subset of newly
diagnosed patients who are unlikely to benefit from splenectomy.’ Studies are ongoing to determine if IFN should
replace splenectomy as initial therapy for other patient
subsets.‘
Despite this impressive activity in HCL, IFN is not
curative in this disease. Only rarely do patients have
well-documented complete remissions and disease progression is the rule following even up to 18 months of continuous therapy.’ The preponderance of data suggest that most,
From the Biological Response Modifiers Program, Clinical Research
Branch, and Program Resources, Inc, National Cancer InstituteFrederick Cancer Research and Development Center (NCI-FCRDC),
Frederick; Frederick Memorial Hospital, Frederick; Biostatistic and
Data Management Section, Clinical Oncology Program, Bethesda,
MD; and Clinical Investigation, Hofmann-LaRoche, Inc, Nutley, NJ.
Submitted June 11, 1990; accepted October 17, 1990.
Supported in part with Federalfunds from the Department of Health
and Human Services under contract number NOl-CO-74102. The
content of this publication does not necessarily reflect the views or
policies of the Department of Health and Human Services, nor does
mention of trade names, commercial products, or organizations imply
endorsement by the U.S.Government.
Address reprint requests to Ronald G . Steis, MD, NCI-FCRDC
(BRMP), FMH Cancer Treatment Center, Clinical Research Branch,
502 W Seventh St, Suite No. 3, Frederick, MD 21 701.
The publication costs of this article were defrayed in part by page
charge payment. This article must therefore be hereby marked
“advertisement” in accordance with 18 U.S.C. section 1734 solely to
indicate this fact.
This is a US government work. There are no restrictions on its use.
0006-4971I9117704-0008$0.0010
792
PATIENTS AND METHODS
Patients. Before receiving IFN therapy, the diagnosis of HCL
was established in all patients using previously reported criteria.“
All patients were required to have one or more PB cytopenias
Blood, Vol77, No 4 (February 15). 1991: pp 792-798
From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
793
LOSS OF INTERFERON ANTIBODIES
(hemoglobin less than 12 g/dL, platelets less than lW,WO/mm3,or
granulocytes less than 1,500/mm3).
All patients were treated according to the same protocol as
outlined previo~sly.~~'~
Each initially received 3 million units of
recombinant I F N - d a (rIFN-u2a) (Roferon-A; Hoffmann-LaRoche, Nutley, NJ) daily by subcutaneous injection for approximately 6 months. The dose in responding patients was then
reduced to 3 million units administered thrice weekly. Maintenance therapy at this dosage was continued indefinitely as long as
responses were maintained with acceptable side effects. Several
patients who did not respond initially or who stopped responding
were given higher doses of IFN for variable periods of time.
The in vivo effects of IFN-a2a antibodies were determined by
serially measuring serum levels of IFN and neopterin and PB
mononuclear cell-associated 2', 5' oligoadenylate synthetase (2-5
A) activity in patients following a single dose of either IFN-cr2a or
human lymphoblastoid IFN. Patients who were still receiving
maintenance therapy discontinued IFN treatments 2 weeks before
this evaluation. Baseline blood samples were obtained three times
for measurement of these parameters. Patients then received 9
million units of I F N - d a subcutaneously. Serial blood samples
were obtained for IFN levels at 0, 1, 2, 3, 4, 6, 8, 10, 12, 24, and 48
hours after the IFN injection and for neopterin levels and 2-5 A
activity at 0, 4, 8, 24, and 48 hours after IFN injection. One week
later, baseline blood samples were obtained again and patients
were then administered 9 million units of human lymphoblastoid
IFN (WELLFERON, Burroughs Wellcome Research, Triangle
Park, NC) subcutaneously. Serial blood samples were again obtained as previously described for measurement of the same
parameters. Patients were not given acetaminophen for fever but
were offered meperidine for chills following the administration of
IFN. Vital signs were obtained serially before and after IFN
administration.
Both studies were approved by the Investigational Review
Boards of the Frederick Cancer Research and Development
Center and the Clinical Oncology Program of the National Cancer
Institute. All patients gave written informed consent.
IFN antibody assays. Serum samples were screened for the
presence of antibodies against IFN-a2a by a solid-phase enzyme
immunoassay (EIA) as described by Itri et al." Specimens testing
positive in the EIA were assessed for their ability to neutralize the
antiviral effects of rIFN-a2a or natural IFN-a in vitro in an
antibody neutralization bioassay (ANB)."Neutralizing antibodies
were considered to be present if the specimen neutralized the viral
protective effects of IFN in bovine kidney cells exposed to vesicular
stomatitis virus (VSV). Results are expressed as neutralization
titers, ie, that dilution of serum that reduced the effective concentration of IFN from approximately 20 U/mL to 1 U/mL. The
antibody titer, expressed as IFN-neutralizing U/mL (INU/mL), is
equivalent to the number of units of IFN neutralized by 1mL of the
patient's serum; under the assay conditions described, it is defined
as the neutralization titer times 20.
Serum IFN levels. IFN levels were determined by inhibition of
the cytopathic effect of VSV on the human amnion WISH cell line
(ATCC, Rockville, MD) as described previously.'* Serial threefold
dilutions of serum samples were added to WISH cell monolayers
and incubated overnight at 37°C. VSV was added and cell viability
was graded from 0 (no kill) to 4 (total kill) after 24 hours. The
endpoint IFN titer was defined as the last serum dilution to show
50% protection of the monolayer. IFN units were defined as 3N
where 3 is the base dilution and N equals the endpoint IFN titer.
Neopterin determinations. Neopterin levels, which are expected
to increase following the administration of IFN," were measured
by a commercial radioimmunoassay (Neopterin-RIAcid, Henning,
Berlin, Germany). Briefly, sera were incubated with an anti-
neopterin antibody followed by addition of an 'ZSI-neopterintracer.
Serum neopterin concentrations were calculated from the percent
drop in 1Z51-binding
when compared with control.
2'-5'-oligoadenylate synthetase (2-5 A ) assay. 2'-5'-oligoadenylate synthetase activity was measured as described by Merritt
et al.14Ficoll-hypaque separated PB mononuclear cells were lysed
and incubated with poly rI.rC agarose beads to allow adherence of
the 2-5 A enzyme. Beads were then incubated for 20 hours with
'H-adenosine triphosphate (ATP) buffer solution followed by
digestion with bacterial alkaline phosphatase. Five-microliter aliquots were spotted onto diethyl aminoethyl (DEAE)-cellulose
paper disks, washed, eluted with 0.3 mol/L KCI, and counted.
Specific activity was defined as picomoles ATP incorporated per
10s cells per hour. One unit of enzyme activity was defined as the
incorporation of 1pmol/L ATP/hour at 37°C.
RESULTS
IFN antibodies. Between April 1984 and December
1985, 56 patients with HCL were entered on study and
began treatments with IFN-a2a. Of these, 35 subsequently
were studied to determine the in vivo effects of IFN in the
presence or absence of IFN antibodies. Thirty-two of these
35 patients were receiving IFN up to 2 weeks before study
entry while three patients had discontinued IFN 6, 15, and
17 months before study entry. IFN was stopped in these
three patients because of acquired IFN resistance (one
patient) or because of what were felt to be IFN-related side
effects (two patients). IFN antibody analyses of these 35
patients (Table 1) showed that many patients who previously had IFN antibodies became antibody negative during
continuing IFN administration. For example, of the 10
patients who previously had neutralizing antibody, three
became antibody negative, five continued to have antibody
that no longer had neutralizing activity, and only two
continued to have neutralizing antibody. One of these latter
two patients had developed IFN resistance when neutralizing antibodies developed' and discontinued IFN 17 months
before participation in this study. However, at the time he
entered this study he had become neutralizing-antibody
negative.
The loss of IFN antibodies observed in our study population despite continued exposure to IFN prompted us to
evaluate the antibody status of the 21 remaining patients
who had not entered this second study. Serum specimens
were available from 16 of these patients. Of three patients
who previously had non-neutralizing IFN antibodies, one
Table 1. Antibody Status of Patients Receiving Continuous IFN-a2a
Therapy for HCL
~
No. PatientstPrior
Antibody Status
Current Antibody
status*
Median (range)
Duration of
Antibody
Positivity (mo)
16 Antibody negative
9 Non-neutralizing
10 Neutralizing
16 Antibody negative
-
~
9 Antibody Negative
3 Antibody negative
5 Non-neutralizing
2 Neutralizing
14.5 (2-48)
10 (2-34)
*"Current antibody status" refers to the patients' antibody status at
the time of entry onto the study or, if IFN had been stopped, at the time
IFN was discontinued.
From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
794
had become antibody negative and two continued to have
non-neutralizing antibodies. Eight patients previously had
neutralizing antibodies; two became antibody negative, two
continued to have antibody but without IFN-neutralizing
activity and four continued to have neutralizing antibodies.
All five antibody-negative patients remained antibody negative. Thus, in total, all 21 patients who were antibody
negative previously remained antibody negative, 10 of 12
patients with non-neutralizing antibodies became antibody
negative, and 12 of 18 patients with neutralizing antibodies
became either antibody negative (five patients) or continued to have antibody that no longer had neutralizing
activity (seven patients). Of the remaining six patients with
persisting neutralizing IFN antibodies, five had either de
novo (one patient) or acquired (four patients) IFN resistance and remained neutralizing-antibody positive at least
until IFN therapy was discontinued. One patient with
persisting neutralizing antibody continues on IFN therapy.
The median duration of antibody positivity among the 35
patients who entered our study with neutralizing or nonneutralizing antibodies was 10 months (range 2 to 34
months) and 14.5 months (range 2 to 48 months), respectively. The two patients who continued to have neutralizing
antibodies have had them for 7 and 35 months. Those five
patients who converted from neutralizing to non-neutralizing antibodies had neutralizing antibodies for a median of
14 months (range 2 to 18 months). The median duration of
IFN therapy among those patients who never developed
detectable IFN antibodies was 46 months.
The specificity of the antibody response to IFN was
tested in vitro in a viral protection assay using either
IFN-a2a or partially purified natural IFN. All neutralizing
IFN antibodies detected in our patients were specific for
IFN-a2a. No inhibition of the viral protective effects of
partially purified natural IFN was observed.
In vivo effects of IFN antibodies. Subcutaneous administration of both IFN-a2a and human lymphoblastoid IFN
brought about consistent and reproducible increases in
serum levels of neopterin, IFN, and PB mononuclear
cell-associated levels of 2-5 A in patients without antibodies
and in those with non-neutralizing antibodies (Fig 1).
Non-neutralizing IFN antibodies had no statistically significant effect (by the Wilcoxon rank sum test) on the increase
over baseline of neopterin or 2-5 A levels when compared
with what was observed in these same patients following
human lymphoblastoid IFN administration or compared
with the antibody-negative group following IFN-a2a administration. Serum IFN levels increased to a significantly
greater degree following human lymphoblastoid IFN than
following IFN-a2a administration in both the antibodynegative and non-neutralizing antibody groups. In the one
patient with neutralizing IFN antibodies, serum IFN was
not detectable and there was no or very little increase in
neopterin and 2-5 A levels following subcutaneous injection
of IFN-a2a. However, following subcutaneous injection of
the same number of units of human lymphoblastoid IFN,
changes in IFN, neopterin, and 2-5 A levels were similar to
STEIS ET AL
Neutralizing Ab Positive
=
-2.
E
1
400
-
300
- -Interferon
:
b
=
IFNp2a
500
200-
-50
-40
-30
-0- Neopterln
-20
..e..2 ’ 5 ’ - A
c
100r 4 - 2
0
2
4
6
8
-+
-.E
i
30
-25
5
.-
-20
5
0
-15
f
0
-10
5
ul
-10
-5
0
-0
N
10
Non-Neutralizing Ab Positive
IFN,uZa
I
1 4 - 2
0
2
6
4
IO
8
Antibody Negative
IFNunl
IFNa2a
100
I
I
160
-2.
E
30
E
e
c
:.
40
b
-c
c
20
10
-2
2
4
6
8
1
0
Day Relative to Start of Therapy
Fig 1. Changes in IFN, neopterin, and 2-5 A levels in a representative patient from each group following subcutaneous administration
of either IFN-da or human lymphoblastoid IFN. Nine million units of
IFN-u2a was administered on day 0 and the same amount of human
lymphoblastoid IFN was administered on day 7.
those observed in patients with either no antibody or only
non-neutralizing antibody.
Body temperature changes were monitored for 12 hours
following each IFN injection (Fig 2). Given the limited
number of patients who continued to have neutralizing IFN
antibodies, statistical comparisons of the febrile response to
the IFNs is not possible. However, the one patient with
neutralizing IFN-a2a antibodies had only a transient temperature increase following IFN-a2a injection that was of
shorter duration than patients with either no antibody or
only non-neutralizing antibodies. However, this same patient had as high and as prolonged a temperature increase
following human lymphoblastoid IFN injection as the other
patients. Although patients with non-neutralizing antibod-
From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
795
LOSS OF INTERFERON ANTIBODIES
40
- "'1
0
Ab Positive
Ab Positive
Neutralizing
Non-Neu t ralizing
3gl
Ab Negative
3g,61
I
p
n='
39
'(
n-6
39
II
39
n924
3 4
'
36"
o
i
'
z
'
s
'
4
'
~
.
'
'
'
8
7
e
'
.
'
9
'
m
36,6
SI
'
n
w
L
o
i
a
3
4
Hours
6
e.
7
a o i o n w
36.6
9
.1
.
.
'
'
'
o
i
z
'
s
Hours
'
4
'
6
'
'
8
7
'
a
'
o
'
'
m
n
u
'
Hours
Fig 2. Body temperature changesfollowing subcutaneous administrationof 9 million units of either IFN-a2a (H)or human lymphoblastoid IFN
(0).
Curves show median temperatures for each patient group. Standard deviations for each point were usually less than 0.2% and are not
included in the graphs for clarity. The discrepancy in the number of patients with non-neutralizingantibodies between this figure and Table 1 is
because one patient had neutralizingantibody when IFN-&a was discontinued but had a non-neutralizing antibody when he entered our second
study. Four patients in the antibody-negative group were excluded from this analysis because two did not complete the study and two
inadvertentlyreceived acetaminophen.
ies showed a trend toward lower temperatures following
IFN-a2a than after human lymphoblastoid IFN, the difference was not statistically significant.
Relationship of IFN antibody status and clinical outcome.
The presence of neutralizing IFN antibodies has previously
been correlated with acquired resistance to the antileukemic effects of IFN in patients with HCL.' Despite substantial inhibition of IFN-induced changes in body temperature
and in 2-5 A and neopterin levels, our one remaining
patient with IFN-neutralizing antibodies has continued to
maintain a partial response to IFN therapy with sustained
improvements in PB counts and bone marrow (BM) hairy
cell infiltration. However, he is the only patient receiving
continuous IFN-a2a therapy who has a progressively increasing soluble interleukin-2 (IL-2) receptor level (data not
shown). Levels of the soluble IL-2 receptor have previously
been correlated with disease status in HCL.15316
The increasing soluble IL-2 receptor level in this patient with neutralizing IFN antibodies suggests that disease progression may be
occurring despite continued IFN-a2a administration.
The clinical relevance of loss of neutralizing IFN antibodies is illustrated in the clinical course of the patient shown
in Fig 3. This patient developed resistance to IFN at
approximately day 100 of therapy. At about the same time,
the IFN dose was reduced from 3 million units daily to 3
million units thrice weekly. Decreasing granulocyte and
platelet counts did not improve when IFN was again
administered at 3 million units daily but blood count
improvement was observed following dose escalation to 6
million units daily. However, when she began treatments at
this dose neutralizing antibodies were no longer detectable.
Subsequent IFN-dose reductions to 3 million units daily
and 3 million units thrice weekly, which previously were
ineffective in the presence of antibody, were now effective
at maintaining normal PB counts in the absence of antibody. Thus, antibody-mediated IFN resistance was overcome by increasing the IFN dose and the effective dose of
IFN subsequently decreased to the "standard" dose level (3
million units thrice weekly) when neutralizing antibodies
no longer were detectable.
DISCUSSION
The use of IFNa in the treatment of malignant and
infectious diseases is increasing. Although currently approved for use in HCL and in acquired immunodeficiency
syndrome (AIDS)-associated Kaposi's sarcoma, recombinant forms of IFNa are also significantly active in indolent
non-Hodgkin's lymphoma^,'^ chronic myelogenous leukemia (CML),18.'9
and chronic active hepatitis.'" One study has
also suggested that IFNa in combination with 5-fluorouracil
is highly active in previously untreated patients with colorectal carcinoma." In all of these diseases, complete remissions are uncommon and, where evaluated, disease progression following cessation of therapy is the r ~ l e . ~ ,The
*~,~
development of IFN-neutralizing antibodies, observed with
both commercially available forms of IFNa, and the correlation of the presence of these antibodies with acquired
disease resistance to IFNy,,Z*1,Z5
raises the possibility that the
prolonged IFN therapy anticipated for these diseases might
not be possible. Increasing proportions of patients might be
expected to develop antibodies to IFN during long-term
therapy, rendering IFN ineffective. A greater understanding of the effects of these antibodies on easily measurable in
vivo parameters of IFN activity might allow rational therapeutic alterations in patients with antibody-mediated IFN
From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
796
STEIS ET AL
IFN Dose
EIA
ANB
-I
3 MUqd
:
.3MUqd+3MUtiw
+
+
+
+
:
+ + -
6 MUqd
3 MUqd-3
t
-
I
4
MUtiw-
(150)/600)(1200)/1200)(1200]i3001
I II
III I
I
6,000
600
5,000
,500
4,000
,400
%
3,000
300
4
I
8
.
(*
V
1
c
E
LI
:
1
1
1
8
3
2
200
2,000
-4
0
1,000
loo
51
9
2
$
0
0
100 200 300 400 500 600 700 800 900 1000 1100 1200 1300 1400 1500
Day of Therapy
resistance. Therefore, we studied the in vivo effects of IFN
antibodies in a population of patients with HCL, some of
whom had developed antibodies to IFN-a2a, to determine
the in vivo effects of these antibodies and to determine if
serologically distinct IFN species could overcome antibodyrelated resistance to IFN-a2a.
We first determined the incidence of IFN antibodies. To
our surprise, the majority of patients who were previously
antibody positive had become antibody negative despite
continuous exposure to IFN-a2a. The loss of serum IFN
antibodies was seen in patients with a wide range of titers
(300 to 6400 INU/mL), not only in those with the lowest
values. The assay used in this study had a lower limit of
sensitivity of 100 INU/mL and the lowest neutralizing
antibody titer previously observed in our patients was 300
INU/mL. Furthermore, all patients in our study had discontinued IFN treatments 2 weeks before study entry. Loss of
IFN antibodies was, therefore, not due to recent IFN
administration, which can acutely lower antibody titers."
Alterations in the immunogenicity of the IFN-a2a administered to our patients also does not explain loss of IFN
antibodies because the processes used in IFN-a2a manufacture have not changed (Itri, L., personal communication,
March 1990). Thus, either immunologic tolerance to IFNa2a developed in these patients or the IFN antibodies were
produced by short-lived B cells without the development of
immunologic memory.
Loss of IFN antibodies has clinical relevance in some
patients. The acquired IFN resistance of the patient described in Fig 3 may have been overcome by an increase in
IFN dose. But an increase in IFN dose to 6 million units
daily was made at the same time that neutralizing IFN
antibodies became undetectable. Thus, responsiveness to
IFN may have been restored because of loss of IFN
antibodies rather than to the higher IFN dose. Certainly,
6
Fig 3. Serial granulocyte and
platelet counts in a patient receiving continuous IFN-a2a for
HCL. IFN doses and results of
both EIA and ANB for IFN antibodies are shown across the top.
Numbers across the top refer to
neutralizing antibody titers.
the continued response to IFN-a2a at a dose of 3 million
units three times weekly must have been due to loss of IFN
antibodies because disease progression had occurred at this
IFN dose when neutralizing antibodies were present. Therefore, if patients develop IFN antibodies and IFN resistance,
higher IFN doses may overcome the resistance but may be
required for only a short period of time (in our patient, less
than 10 months). Serial measurements of IFN antibodies
may be helpful in guiding the management of such patients.
Resistance to IFN is not universal among patients with
HCL who develop neutralizing IFN antibodies. In one
study, only 6 of 16 patients with neutralizing IFN antibodies
experienced disease progression despite continued IFN
administration.' This observation may be explained by the
indolent nature of this disease. In the Cancer and Acute
Leukemia Group B study of IFN-a2b in HCL: disease
progression requiring reinstitution of therapy occurred a
median of approximately 2 years following cessation of
either 12 or 18 months of therapy. Therefore, in patients
initially responsive to IFN who develop antibody-mediated
IFN resistance, a median of as long as 2 years might elapse
before IFN resistance becomes evident clinically, even
among patients with antibody titers sufficiently high to
neutralize all IFN administered. The results shown in Fig 1
are consistent with this interpretation in that acute changes
in levels of neopterin and 2-5 A induced by IFN were nearly
completely blocked by neutralizing IFN antibodies. The
antibodies were sufficiently potent to completely block the
appearance of IFN-a2a in the serum following subcutaneous administration. Despite these inhibitory effects of the
antibody, this patient is maintaining a partial response to
IFN therapy. That these inhibitory effects are due to
antibody and not to a generalized unresponsiveness to IFN
is confirmed by the normal expected response of neopterin
and 2-5 A levels to human lymphoblastoid IFN, a prepara-
From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
797
LOSS OF INTERFERON ANTIBODIES
tion of IFN that contains many different molecular species
of IFNa, most of which are serologically distinct from
IFN-a2a. These results show that IFN-neutralizing antibodies inhibit IFN responses in vivo. However, the clinical
effects of this inhibition appear to depend upon the natural
history of the disease being treated. The indolent nature of
HCL may permit interference with the antileukemic effects
of IFN to occur for a prolonged period of time without
clinical deleterious effects. It is of interest that the development of IFN-a2b neutralizing antibodies in patients with
CML, a disease generally more rapidly progressive than
HCL, is nearly universally associated with clinical disease
progressionF6
Alterations in the management of patients with disease
responsive to IFN is not required solely because of the
development of neutralizing IFN antibodies. Antibodymediated resistance to IFN may be overcome by increasing
the IFN dose. Even if higher doses are ineffective, patients
who can safely sustain loss of IFN effectiveness for several
months may ultimately lose neutralizing IFN antibodies
with return of IFN effectiveness. Animal studies have
shown that antigens to which tolerance can develop are, in
general, slowly metabolized, presumably resulting in prolonged in vivo exposure.’’ If tolerance induction is the
mechanism by which patients lose IFN antibodies, continuous IFN therapy may be required to induce tolerance and in
this regard may be preferred over intermittent therapy, at
least in patients with HCL. Monitoring of antibody levels
should continue to be a routine part of the clinical evaluation of IFN and other recombinant biologic agents.
REFERENCES
1. Quesada JR, Reuben J, Manning JT, Hersh EM, Gutterman
JU: Alpha interferon for induction of remission in hairy-cell
leukemia. N Engl J Med 310:15,1984
2. Quesada JR, Hersh EM, Manning J, Reuben J, Keating M,
Schnipper E, Itri L, Gutterman JU: Treatment of hairy cell
leukemia with recombinant a-interferon. Blood 68:493,1986
3. Federico M, Chisesi T, Lauria F, Bernasconi C, Capnist G,
Castoldi G, Damasio E, Pagnucco G, Frassoldati A, Resegotti L,
Silingardi V: Human lymphoblastoid interferon as initial therapy in
hairy cell leukaemia: A multicentre study in non-splenectomized
patients. Br J Haematol7254, 1989
4. Golomb HM, Fefer A, Golde DW, Ozer H, Portlock C, Silber
R, Rappeport J, Ratain MJ, Thompson J, Bonnem E, Spiegel R,
Tensen L, Burke JS, Vardiman JW: Report of a multi-institutional
study of 193 patients with hairy cell leukemia treated with
interferon-alfa 2b. Semin Oncol 15:7,1988
5. Ratain MJ, Vardiman JW, Barker CM, Golomb HM: Prognostic variables in hairy cell leukemia after splenectomy as initial
therapy. Cancer 62:2420,1988
6. Steis RG, Longo DL: Update on the treatment of hairy cell
leukemia. Cancer, Principles Pract Oncol2:1, 1988
7. Ratain MJ, Golomb HM, Vardiman JW, Westbrook CA,
Barker C, Hooberman A, Bitter MA, Daly K Relapse after
interferon alfa-2b therapy for hairy-cell leukemia: Analysis of
prognostic variables. J Clin Oncol6:1714,1988
8. Smith I1 JW, Longo DL, Urba WJ, Clark JW,Sznol M,
Creekmore SP, Steis RG: Prolonged, continuous treatment of
hairy cell leukemia (HCL) with recombinant interferon-alfa-2a
(rIFN-a2a). Blood 74:198a, 1989 (abstr)
9. Steis RG, Smith I1 JW, Urba WJ, Clark JW, Itri LM, Evans
LM, Schoenberger C, Longo D L Resistance to recombinant
interferon alfa-2a in hairy-cell leukemia associated with neutralizing anti-interferon antibodies. N Engl J Med 318:1409,1988
10. Foon KA, Maluish AE, Abrams PG, Wrightington S, Stevenson HC, Alarif A, Fer MF, Overton WR, Poole M: Recombinant
leukocyte A interferon therapy for advanced hairy cell leukemia:
Therapeutic and immunologic results. Am J Med 80:351,1986
11. Itri LM, Campion M, Dennin RA, Palleroni AV, Gutterman
JU, Groopman JE, Trown PW: Incidence and clinical significance
of neutralizing antibodies in patients receiving recombinant interferon alfa-2a by intramuscular injection. Cancer 59:668, 1987
12. Armstrong JA: Cytopathic effect inhibition assay for interferon: Microculture plate assay, in Pestka S (ed): Methods in
Enzymology, Interferons, vol 78, Part A. San Diego, CA, Academic, 1981, p 381
13. Lang A, Niederwieser D, Huber C, Swetly P, Fuchs D,
Hausen A, Reibnegger G, Wachter H: Treatment with recombinant interferon alpha-2 induces increase of in vivo neopterin
excretion, in Pfleiderer W, Wachter H, Curtius H-Ch (eds):
Biochemical and clinical aspects of pteridines, vol 3. Berlin,
Germany, Walter de Gruyter, 1984, p 251
14. Merritt JA, Borden EC, Ball LA: Measurement of 2’,5’oligoadenylate synthetase in patients receiving interferon-alpha. J
Interferon Res 5:191, 1985
15. Chilosi M, Semenzato G, Cetto G, Ambrosetti A, FioreDonati L, Perona G, Berton G, Lestani M, Scarpa A, Agostini C,
Trentin L, Zambello R, Masciarelli M, Dazzi F, Vinante F,
Caligaris-Cappio F, Pizzolo G: Soluble interleukin-2 receptors in
the sera of patients with hairy cell leukemia: Relationship with the
effect of recombinant a-interferon therapy on clinical parameters
and natural killer in vitro activity. Blood 70:1530, 1987
16. Steis RG, Marcon L, Clark J, Urba W, Longo DL, Nelson
DL, Maluish AE: Serum soluble IL-2 receptor as a tumor marker
in patients with hairy cell leukemia. Blood 71:1304,1988
17. Foon KA, Sherwin SA, Abrams PG, Longo DL, Fer MF,
Stevenson HC, Ochs JJ, Bottino GC, Schoenberger CS, Zeffren J,
Jaffe ES, Oldham R K Treatment of advanced non-Hodgkin’s
lymphoma with recombinant leukocyte A interferon. N Engl J Med
311:1148,1984
18. Talpaz M, McCredie KB, Mavligit GM, Gutterman JU:
Leukocyte interferon-induced myeloid cytoreduction in chronic
myelogenous leukemia. Blood 62:689, 1983
19. Talpaz M, McCredie K, Kantarjian H, Trujillo J, Keating M,
Gutterman J: Chronic myelogenous leukaemia: Haematological
remissions with alpha interferon. Br J Haematol64:87,1986
20. Hoofnagle JH, Jones EA: Therapy of chronic viral hepatitis:
Past, present, and future. Semin Liver Dis 9:231, 1989
21. Wadler S, Schwartz EL, Goldman M, Lyver A, Rader M,
Zimmerman M, Itri L, Weinberg V, Wiernik PH: Fluorouracil and
recombinant alfa-2a-interferon: An active regimen against advanced colorectal carcinoma. J Clin Oncol7:1769, 1989
22. Gisslinger H, Linkesch W, Fritz E, Ludwig H, Chott A,
Radaszkiewicz Th: Long-term interferon therapy for thrombocytosis in myeloproliferative diseases. Lancet 1:634, 1989
23. Davis GL, Balart JdA, Schiff ER, Lindsay K, Bodenheimer
HC, Perrillo RP, Carey W, Jacobson IM, Payne J, Dienstag JL,
VanThiel DH, Tamburro C, Lefkowitch J, Albrecht J, Meschievitz
C, Ortego TJ, Gibas A, Hepatitis Interventional Therapy Group:
Treatment of chronic hepatitis C with recombinant interferon alfa
From www.bloodjournal.org by guest on November 7, 2014. For personal use only.
STElS ET AL
(A multicenter randomized, controlled trial). N Engl J Med
321:1501,1989
24. von Wussow P, Freund M, Block B, Diedrich H, Poliwoda H,
Deicher H: Clinical significance of anti-IFN-a antibody titres
during interferon therapy. Lancet 2:636, 1987
25. Porres JC, Carreno V, Ruiz M, Marron JA, Bartolome J:
Interferon antibodies in patients with chronic HBV infection
treated with recombinant interferon. J Hepatol8:351, 1989
26. Freund M, von Wussow P, Diedrich H, Eisert R, Link H,
Wilke H, Buchholz F, LeBlanc S, Fonatsch C , Deicher H, Poliwoda
H: Recombinant human interferon (IFN) alpha-2b in chronic
myelogenous leukaemia: Dose dependency of response and frequency of neutralizing anti-interferon antibodies. Br J Haematol
72:350, 1989
27. Schwartz RH: Acquisition of immunologic self-tolerance.
Cell 57:1073,1989