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Treatment of rheumatoid arthritis with monoclonal CD4 antibody M-T151. Clinical results and immunopharmacologic effects in an open study including repeated administration

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525
TREATMENT OF RHEUMATOID ARTHRITIS WITH
MONOCLONAL CD4 ANTIBODY M-T151
Clinical Results and Immunopharmacologic Effects in an Open Study,
Including Repeated Administration
CHRISTIAN REITER, BAHRAM KAKAVAND, ERNST PETER RIEBER,
MANFRED SCHATTENKIRCHNER, GERT RIETHMULLER, and KLAUS KRUGER
Recent experimental and clinical data point to
the T helper lymphocyte subset as playing a central role
in the pathogenesis of rheumatoid arthritis (RA). Thus,
a therapeutic strategy aimed specifically at the CD4 T
cell subset is warranted. We treated patients with active
RA for 7 days with a daily dose of 20 mg of CD4
monoclonal antibody M-T151, administered intravenously over 30 minutes. There were no negative side
effects. According to changes in the combined parameters of Ritchie articular index, pain assessment, grip
strength, and morning stiffness, 6 patients had a good
response. Clinical improvement was greatest approximately 2 weeks after termination of the therapy and
lasted from 4 weeks to 6 months. Of the serologic
parameters of inflammation, only the C-reactive protein
level improved in the patients with a favorable response.
Close immunologic monitoring revealed a transient,
selective depletion of CD4+ T cells after each infusion.
During the entire treatment period, residual circulating
CD4+ cells were found to be coated with CD4 antibody,
whereas free antibody was detected in the serum only
for approximately 8 hours after each infusion. ImmediFrom the Institute for Immunology and the Department of
Rheumatology, Medizinische Poliklinik, University of Munich, Ge:rmany.
Christian Reiter, MD: Institute for Immunology; Bahram
Kakavand: Institute for Immunology; Ernst Peter Rieber, MD:
Professor of Immunology, Institute for Immunology; Manfred
Schattenkirchner, MD: Professor of Medicine and Head, Department of Rheumatology, Medizinische Poliklinik; Gert Riethmiiller,
MD: Professor of Medicine and Head, Institute for Immunology;
Klaus Kriiger, MD: Department of Rheumatology , Medizinische
Poliklinik.
Address reprint requests to Gert Riethmiiller, MD, Institut
fur Immunologie, Goethestr. 31, D-8000 Miinchen 2, FRG.
Submitted for publication May 4, 1990; accepted in revised
form November 6, 1990.
Arthritis and Rheumatism, Vol. 34, No. 5 (May 1991)
ately after infusion, soluble CD4 antigen appeared in the
serum. In addition to the cell-bound CD4 antibody,
complement components could be detected on the surface of the remaining CD4+ cells. The proliferative
response of peripheral blood mononuclear cells to purified protein derivative was significantly diminished 4
weeks after cessation of antibody treatment. Six patients
showed a weak antibody response to mouse immunoglobulin. In 4 of the responders who received a second
course of therapy (2 of them as outpatients), a therapeutic effect was noted that was similar to that after the
first course. Only 1patient, who had low titers of serum
IgE anti-mouse Ig antibodies, showed a mild anaphylactic reaction at the end of the second course of therapy.
Treatment of RA with the monoclonal CD4 antibody
M-T151 seems to be a promising alternative, although
the optimal dose and the regimen of administration are
still to be defined.
Several independent lines of evidence indicate
T lymphocyte dysregulation as a major factor in the
pathogenesis of rheumatoid arthritis (RA). Therefore,
in the search for a selective and pathogenetically
oriented treatment of RA, therapeutic modulation of T
lymphocyte subpopulations seems a logical approach.
Several procedures in which total lymphocyte depletion has been used as a therapeutic regimen, such as
thoracic duct drainage (l), total lymphoid irradiation,
and lymphapheresis ( 2 4 , have proved to be either too
hazardous or ineffective. Use of monoclonal antibodies (MAb) directed at a particular T cell subpopulation
or receptors on T cells may represent a more selective
approach to the treatment of RA. Because CD4:CD8 T
cell ratios in peripheral blood have been shown to be
elevated during active phases of the disease (7) and
REITER ET AL
526
Table 1. Characteristics of the study population before study entry*
Disease
duration Functional
Patient Age (years)
class
NSAIDs
33
10
I11
6
7
I1
3
59
22
111
4
58
10
I1
5
57
10
I11
6
57
17
I1
7
68
12
I1
8
49
25
I1
9
68
8
I11
+
+
+
+
+
+
+
+
+
10
49
10
111
+
1
2
6
Steroids, dosage
Prednisolone, 10
mg/day
Methylprednisolone,
8 mg/day
Fluocortolone, 12.5
mglday
Fluocortolone, 10
mglday
Fluocortolone, 12.5
mg/day
Fluocortolone, 10
mg/day
Fluocortolone, 10
mg/daY
Fluocortolone, 10
mg/day
Fluocortolone, 10
mg/day
Methylprednisolone,
10 mg/day
Previous DMARDs and
immunosuppressive agents
CQ, gold, DP, AZA, CSA
CQ, gold, SSZ, AZA
CQ, gold, SSZ, AZA, LA
CQ. gold, SSZ, AZA
CQ, gold, DP, AZA
CQ, gold, DP, AZA
CQ, gold, DP, AZA, IFNy
CQ, gold, SSZ, AZA, LA
CQ, gold, SSZ, AZA
CQ, gold, DP, SSZ, AZA
* Functional class was determined according to Steinbrocker’s criteria (22). NSAIDs = nonsteroidal
antiinflammatory drugs; DMARDs = disease-modifyingantirheumatic drugs; CQ = chloroquine; gold
= gold salts; DP = D-penicillamine;AZA = azathioprine; CSA = cyclosporin A; SSZ = sulfasalazine;
IFNy = interferon-y; LA = lymphapheresis.
because CD4+ T cells have been found to be the
predominant lymphocyte subpopulation infiltrating the
synovial membrane of affected joints (8), a therapy
targeted at this particular subpopulation seemed to be
warranted. Moreover, recent reports demonstrate that
experimental autoimmune diseases in rodents (9,lO)
and primates (1 1) can be effectively controlled by the
administration of monoclonal antibodies to CD4.
Reports of CD4 MAb treatment in humans with
RA have recently been published (12-16), and the
results are promising. CD4, an integral membrane
glycoprotein on T helper lymphocytes, is intracellularly associated with the tyrosine kinase p56ICk(17) and
seems to interact as a coreceptor with the T cell
antigen-receptor complex (18). Because of its binding
affinity for class I1 major histocompatibility complex
molecules, CD4 is deemed to stabilize the cellular
contact of T cells with class 11-positive antigenpresenting cells (19). From experimental work, there is
no clear indication which mechanisms are operative in
producing the therapeutic effect, and on which parameters the therapeutic efficiency of a CD4 MAb can be
predicted. It is likely that CD4 antibodies block natural
cell-cell interactions, mediate new contact to Fc
receptor-bearing cells, and modulate the state of activation of CD4+ cells. In vivo, CD4 MAb may alter the
recirculation of CD4 cells or may eliminate them by
direct cytolytic action or via opsonization. Thus, a
further study using a higher dose of CD4 MAb than
was previously used was initiated with the intention of
studying more thoroughly the induced clinical and
immunopharmacologic effects.
PATIENTS AND METHODS
Patients. Ten patients with active RA, as defined by
the criteria of the American Rheumatism Association (20),
were selected for the study. Disease was considered severe
and active if at least 3 of the following 4 inclusion criteria
were present: 1) Ritchie articular index (21) >lo, 2) duration
of morning stiffness 2 4 5 minutes, 3) erythrocyte sedimentation rate (ESR) >30 mm in the first hour, and 4) patient’s
assessment of pain >50 mm on a 100-mm visual analog scale.
Exclusion criteria were: age 270, functional class IV according to Steinbrocker’s criteria (22), leukopenia (<3,000/ml),
intercurrent infection, severe concomitant medical illness, preexisting allergy to murine MAb (excluded by skin testing).
All patients had undergone several therapeutic regimens
with disease-modifying antirheumatic drugs (DMARDs) and
with immunosuppressive agents (Table 1).
MAb treatment. To avoid any interference with previous therapy, a 1-month washout period was recommended. Medication was restricted to a constant dose of
prednisolone (maximum dosage 10 mg per day, a dosage
that, evidently, did not control the disease activity) from 4
ANTI-CD4 THERAPY FOR RA
weeks before until 4 weeks after the course of CD4 MAb
treatment. Only paracetamol was permitted as an analgesic.
None of the patients was taking a DMARD at the time of
recruitment for study, but all were taking corticosteroids at
a daily dosage of 2 1 0 mg of prednisolone equivalent.
Patients were hospitalized during MAb treatment,
and 20 mg of CD4 MAb M-TI51 was infused intravenously,
over a period of 30 minutes, every morning for 7 days. Two
of the patients received a second course of treatment on an
outpatient basis.
Therapeutic MAb M-T151. M-T151 (23) is a mouse
IgG2a monoclonal antibody that binds to an epitope formed
by the second, and influenced by the first, immunoglobulinlike domain of the human CD4 antigen (24,25). It efficiently
blocks antigen-induced Ca2’ mobilization in T cell clones
(26) and can suppress the response of peripheral blood
mononuclear cells (PBMC) to soluble antigen or allogeneic
lymphocytes. The hybridoma was propagated as ascites
tumor, and Ig was purified from ascites fluid by Pevicon
C-870 block electrophoresis (Fluka, Neu-Ulm, FRG) and
Sephacryl S300 gel chromatography, then adjusted to 5
mg/ml in 0.9% NaCl, and frozen in sterile aliquots. The
antibody preparation met the German Association of Immunology recommendations for the production and testing of
monoclonal antibodies for therapeutic trials (27).
Clinical assessment. Immediately prior to treatment
and on days 7, 21, and 35, and at monthly intervals up to 6
months, a general clinical examination was performed on the
patients. The following parameters were assessed each time:
Ritchie articular index, grip strength, duration of morning
stiffness, and patient’s assessment of pain. Because we
could not expect to induce a complete remission of this
extremely refractive RA by a 1-week course of treatment,
we used the following definitions of overall clinical response:
(a) good response was defined as at least 20% improvement
in all parameters or at least 35% improvement in 3 parameters, (b) moderate response was defined as at least 20%
improvement in 3 parameters or at least 35% improvement in
2 parameters, (c) no response was defined as all other cases.
Laboratory assessment. The following laboratory parameters were analyzed at the same time as the clinical
assessments: ESR, C-reactive protein (CRP), serum iron,
serum creatinine, transaminase levels, alkaline phosphatase,
serum protein electrophoresis, quantitative immunoglobulins, and rheumatoid factor (by nephelometry). During the
7-day treatment course, the hemoglobin level, platelet count,
and white blood cell count (including differential cell count)
were analyzed daily.
Immunologic assessment. At the same times as routine laboratory assessments were done, plasma was obtained
and PBMC were prepared from heparinized blood by Ficoll
density gradient centrifugation. These samples were stored
frozen. For kinetics studies, blood was also drawn into tubes
containing 10 mM EDTA, at 1 hour, 3 hours, 8 hours, and 24
hours after the first MAb infusion. PBMC were separated
and complement fixation was determined on fresh PBMC
(within hours of venipuncture). All other assays were done
with frozen specimens.
Analysis of lymphocyte subpopulations. In vivo MAblabeled cells were stained with fluorescein isothiocyanate
(FITC)-conjugated F(ab’), rabbit anti-mouse Ig (Dakopatts,
527
Hamburg, FRG) as second antibody. Lymphocyte subpopulations were determined using fluorochrome-conjugated
MAb or biotinylated antibodies and avidin conjugated with
FITC or phycoerythrin (PE). Analysis of cells was done with
a FACScan flow cytometer (Becton Dickinson, Mountain
View, CA).
Complement fixation. To determine whether the infused CD4 MAb induced complement fixation on the cell
surface, PBMC were separated from EDTA-treated blood
and were labeled with rabbit antisera to human complement
components Cls, C3b, C3d, C5, C9, and C4bp (Calbiochem
Behring, Frankfurt, FRG) that had been preabsorbed with
immobilized M-T151. After washing the cells, FITCconjugated swine anti-rabbit Ig (Dakopatts) absorbed with
mouse serum and human Ig was added. The cells were then
incubated with biotinylated CD4 MAb M-T3 10, which binds
independently of M-T151, followed by PE-conjugated
avidin.
Concentration of free MAb in plasma. Plasma samples were serially diluted, and 40 pl was mixed with 2 X lo’
PBMC from a healthy donor. After incubation for 30 minutes
on ice, the cells were washed twice, stained with FITCconjugated F(ab’), rabbit anti-mouse Ig, washed again, and
fixed in phosphate buffered saline (PBS) containing 1%
paraformaldehyde. The mean fluorescence intensity of positive cells was determined, and the MAb concentration was
calculated by comparison of the titration curves for test
samples against that for a standard dilution of M-Tl51.
Stimulation by mitogens and antigens. PBMC (5 x
lo4) were suspended in 200 pl of RPMI 1640 plus 10% fetal
calf serum and stimulated with either 1% phytohemagglutinin (Difco, Detroit, MI), 1 pg/ml concanavalin A (Difco), 5
pg/ml tetanus toxoid, 5 pg/ml purified protein derivative
(PPD; Behringwerke, Marburg, FRG), or 2 x lo’ irradiated
PBMC from 2 allogeneic donors. Unstimulated and mitogenstimulated cultures were pulsed on day 3 and antigenstimulated cultures were pulsed on day 6 with 2 pCi of
’H-thymidine per well for 16 hours. The radioactivity in the
cultures was determined with a Beckman L1801 liquid
scintillation counter, and the mean counts per minute were
calculated from triplicate cultures. For quantitative comparison, the values were normalized according to the following
formula:
Pt =
cpmt
cpm,
+ cpm2 + . . . + cpm,
n
where P, = relative proliferation capacity at time t, cpm, =
mean cpm in an individual patient at time t, and n = total
number of samples tested for an individual patient.
Soluble CD4. A sandwich enzyme-linked immunosorbent assay (ELISA) was established to measure soluble CD4
in plasma. Microtiter plates (Immunoplate I; Nunc, Wiesbaden, FRG) were coated overnight with 2 p g h l of CD4 MAb
M-T310 (IgGl) in carbonate buffer, pH 9.6. After washing
the plates in PBS-0.05% Tween 20, additional free sites were
blocked with 1% human serum albumin (HSA) in PBS. Serial
dilutions of patient plasma in PBS-1% HSA were added and
cultured overnight. After extensive rinsing, biotinylated
M-T151, which recognizes a different epitope on the CD4
528
molecule, was added at a final concentration of 5 pg/rnl, and
the plates were incubated for 4 hours. Subsequently, the
plates were washed and incubated for 30 minutes with
peroxidase-conjugated avidin (Dakopatts) diluted 1:2,000 in
PBS-l% bovine serum albumin (BSA). As chromogen substrate, 1 mg/ml of D-phenylenediamine in citrate buffer, pH
5.0, and 0.03% H202 was used.
After 10 minutes, the reaction was stopped with the
addition of 0.1M H2S0,, and the optical density was determined in a 2-wavelength ELISA reader. Recombinant soluble CD4 and a detergent lysate of thymocytes served as
positive control standards. One arbitrary unit corresponds to
1 ng/ml of purified recombinant soluble CD4 (kindly provided by Dr. A. Williams, Oxford, UK). To assess the
specificity of the ELISA, different combinations of first and
second CD4 MAb were used, as well as combinations of
MAb directed toward different antigens.
Soluble CD4 complexes. The ELISA was modified for
the detection of complexes formed by soluble CD4 and
infused MAb M-T151. In place of the second CD4 MAb, a
biotinylated mouse IgG2a-specific antibody (Dakopatts) was
used. Further evidence was obtained by absorption of sera
with Sepharose-bound rabbit anti-mouse IgG.
Human anti-mouse Ig antibodies (HAMA). Antibody
formation was determined by a sandwich ELISA, using the
M-T151 antibody for coating the plates and 1% HSA in PBS
for blocking. Serial dilutions of plasma samples were incubated overnight. As a reference sample, diluted plasma
obtained after the second course of MAb treatment in patient
4 was used. This plasma had high titers of all isotypes.
Bound anti-mouse Ig antibodies were detected with peroxidase-conjugated antibodies to human IgA, IgG, and IgM
(Dakopatts) diluted 1:2,000 in PBS-1% BSA and ortho-Dphenylenediamine as substrate. Titration curves were plotted, and titers were determined by comparison with the
standard.
To determine the levels of M-T151-specific IgE
antibodies, the plates were coated with rabbit anti-IgE
antibody (Dakopatts), blocked, and incubated with plasma
samples. Then, P-galactosidase-conjugated M-T151 F(ab’),
fragment (Boehringer, Mannheim, FRG) and chlorophenol
red-PD-galactopyranoside (Boehringer) as substrate were
added to the plates to eliminate cross-reactions with rheumatoid factor-like antibodies binding to the Fc portion of the
Ig molecules.
Anti-M-T151 blocking antibodies. To determine what
antibodies were blocking the binding of M-T151, 40 pl of
serially diluted plasma was preincubated with a suboptimal
concentration (0.1 pg/ml) of biotinylated M-T151 for 20
minutes at room temperature. Then, 2 x 10’ PBMC in 20 pl
of PBS was added and maintained for 20 minutes on ice.
Cells were washed twice, incubated with FITC-conjugated
avidin (Dianova, Hamburg, FRG), washed again, and fixed
for flow cytometry. Inhibition of binding was quantified as
the reduction in mean channel fluorescence of CD4+ cells
compared with that before treatment. To further discriminate between antiidiotypic and antiisotypic antibodies, MAb
M-T310 (CD4, IgGl), OKT3 (CD3, IgGZa), and M-T805
(CD8, IgG2a) were added at 50 d r n l (each) to the preincubation mixture to block the binding of antiisotypic antibodies.
REITER ET AL
T
I
T
reponder
I
time after starting treatment (weeks)
Figure 1. Improvement in grip strength in rheumatoid arthritis
patients treated with CD4 monoclonal antibody M-T151.There was
31% improvement in the entire group of 10 patients and 55% in the
group of 7 responders at 5 weeks after starting treatment (P not
significant). Values are the mean and SD.
Statistical analysis. Clinical and immunologic data are
expressed as the mean and standard deviation. Normally
distributed variables were evaluated by t-test; others were
evaluated by Wilcoxon’s signed rank test.
RESULTS
Treatment with antibody M-T151 was remarkably well tolerated during the first course, at doses of
20 mg. No immediate or long-term adverse side effects
were noted. There were no changes in the laboratory
values that could be ascribed to the antibody.
Clinical response. The extent of the clinical
response was highly variable, but there was a characteristic time course. Clinical efficacy was classified as
“good” in 6 patients and “moderate” in 1 patient.
Complete remissions, according to the preliminary
criteria of the American Rheumatism Association (28),
did not occur.
Figures 1 and 2 show the changes in grip
strength and the Ritchie articular index for all 10
patients. Typically, clinical improvement was first
observed toward the end of the week of CD4 MAb
treatment, reaching an optimum reduction in pain and
duration of morning stiffness 2 weeks after cessation of
the daily MAb infusions. Grip strength reached an
optimum 4 weeks after cessation of MAb treatment
but P values did not become significant, as assessed by
the Wilcoxon signed rank test. Improvement in the
Ritchie articular index was 34% in the responders ( P 5
0.05 day 0 versus day 35) and 20% in the entire group
ANTI-CD4 THERAPY FOR RA
529
*]
50 ::
3.
Patients
-8-
1
+ 2
-3
+ 4
-+- 5
+6
u
7
+ 8
+9
-*-
0
1
0
5
.
1
~
1
'
1
20
weeks after starling mAb treatment
10
15
'
1
25
'
110
1
30
Figure 2. Improvement in the Ritchie articular index in rheumatoid
arthritis patients treated with CD4 monoclonal antibody (mAb)
M-T151. Ritchie articular indices were reduced by a mean of 20% in
the entire group of 10 patients and 34% in the group of 7 responders
(solid lines) at 5 weeks after starting treatment ( P < 0.05 pretreatment values versus values at 3-9 weeks in all patients, and pretreatment values versus values at 1-17 weeks in the responders, by
Wilcoxon signed rank test). Dashed lines indicate nonresponders.
of patients (P5 0.05 day 0 versus day 35). Improvement in grip strength was 55% in responders (not
significant) and 31% in all patients (not significant) 5
weeks after treatment.
Clinical followup. Followup examinations were
performed every 4 weeks until week 24. Responders
did not receive DMARDs or nonsteroidal antiinflammatory drugs (NSAIDs) during the followup observation period. In all responders, clinical improvement
persisted beyond week 4 (followup was possible only
up to week 8 in 1 patient). Two of the responding
patients still showed a distinct improvement at 8
weeks after the initial course of therapy, but deterioration occurred during weeks 8 and 12, respectively.
In the other 4 responders, the steroid dosage could be
reduced considerably, from the initial 10 mg of preclnisolone equivalent per day, gradually down to 5
mglday in 3 patients, and down to 2.5 mg/day in the
other patient. Their clinical improvement remained
stable for 3 months after the end of therapy, and in 1
patient, improvement persisted for more than 6
months.
In 2 of the 3 patients who were classified as
nonresponders, the clinical course did not change
considerably. In the third nonresponder, a serious
acute deterioration, with development of vasculitis,
occurred during the third month after the 1-week
course of treatment.
Response to second course of therapy. Only
transient clinical improvement could be reinduced by a
second course of CD4 MAb therapy, after a 6-month
interval. The same dose of CD4 MAb infused at the
same rate for the same number of days was administered to 4 patients. Two of them, patients 7 and 8, were
treated as outpatients. Three of these 4 patients had a
good response, similar to that achieved with the initial
course of treatment. The duration of improvement
achieved by the second course appeared to be shorter
than that after the first therapy course, lasting from 3
weeks to 3 months.
Patient 7, who showed only a moderate response, experienced an anaphylactic reaction approximately 5 minutes after initiation of the last infusion
(on the seventh day of treatment). The blood pressure
decreased from 150/90 mm Hg to 100/60 mm Hg, the
heart rate increased from 84 beatdminute to 110
beatdminute, the skin became flushed, and the patient
experienced back pain. Parenteral administration of 5
mg of clemastine and 250 mg of methylprednisolone
12
1
-
0
1
2
3
4
5
time (weeks)
Figure 3. Changes in the levels of C-reactive protein (CRP), as
measured by nephelometry in rheumatoid arthritis patients treated
with CD4 monoclonal antibody M-T151. There was a 36% decrease
in the group of 7 responders (lower graph) at 5 weeks after therapy
(P < 0.1 versus pretreatment levels). The upper graph represents the
group of 3 nonresponders. Values are the mean and SD.
REITER ET AL
530
0
E: 20
x
-g0 15
z
-
% 1.0
z
8
05
0.0
Oh lh 3h 8h 24h
d4 d l
time after starting treatment
d21 d35
Figure 4. Selective depletion of CD4+ cells in rheumatoid arthritis
patients treated with CD4 monoclonal antibody M-T151. Absolute
numbers of cells/pl of blood were calculated from white blood cell
counts, leukocyte differentiation counts, and the relative numbers of
lymphocyte subpopulations, as determined by fluorescenceactivated cell sorter analysis. On days 4 and 7, blood was obtained
immediately before the infusion. The decrease in CD4 cell numbers
was significant at 1 hour, 3 hours, 8 hours, and day 21 (P< 0.05, by
paired t-test). Values are the mean of 14 treatment courses in
10 patients (see Results for details). Bars show the SD for CD4
cells only.
seen, but the intensity of the labeling decreased with
time (Figure 5 ) . The density of the CD4 antigen per cell
was only marginally reduced, as demonstrated with
the non-cross-reactive biotinylated CD4 MAb
M-T3 10. A compensatory increase in the CD4- T cell
subpopulation of the CD3+, CD8- , CD4- phenotype
or of the CD2+, CD16-, CD8-, CD4- phenotype
could not be detected by fluorescence-activated cell
sorter (FACS) analysis. Thus, T lymphocytes that had
completely modulated their CD4 antigen were not
found in the circulation. No drastic change in the
expression of the CD45RA, CD29, CD7, or CD6
Obtained
day 1,
before
MAb-Infusion
M-T310-b10
+ FITt-avldln
FITC-ant1-NIg
v\l
day 1,
1 II after
promptly controlled all symptoms. Thus far, this is the
only such reaction seen in connection with the infusion
of M-T151.
Changes in laboratory parameters of inflammation. The CRP level (Figure 3), as determined by
nephelometry , tended to decrease in parallel with
improvement in the clinical course. The CRP decreased by 36% in the responders (P< 0.1 versus day
0) but by only 2.4% in the nonresponders. The ESR
and rheumatoid factor titers were not affected.
Lymphocyte subpopulations. Upon infusion of
MAb M-T151, a rapid, -50%, decrease in total lymphocyte numbers was observed as a result of the
selective depletion of peripheral blood CD4+ lymphocytes. Within 1 hour, the absolute number of CD4+
cells decreased from a mean & SD level of 1,130 ?
241/pl (at time 0) to 201 -+ 124/pl, but after 24 hours,
the level returned to pretreatment values (1,240 2
221). The absolute numbers of CD8+, CD16+, and
CD20+ (B 1) lymphocytes remained virtually unchanged (Figure 4). An identical pattern of CD4+ T
cell kinetics was seen after each infusion (data not
shown). Two weeks after the last antibody injection
(day 21), a transient decrease in CD4+ lymphocytes
was observed in all patients (Figure 4).
For up to 24 hours after an infusion, residual
circulating CD4+ cells labeled with M-T151 were
MAD-lnf us lon
day 1,
3 h after
MAb- lntus lon
day 1,
8 h after
MAb-lnfus Ion
day 2,
before
MAb-1nf us 1on
day 4,
before
MAb-lnfus Ion
day 8
Iltlonrconca hterlty
Figure 5. Presence of CD4 on peripheral blood lymphocytes (PBL)
from rheumatoid arthritis patients treated with CD4 monoclonal
antibody (MAb) M-T151. M-T151 could be found on all circulating
CD4 cells throughout the treatment period. The reduced staining
intensity at 8 hours and 24 hours after infusion is not due to CD4
modulation, because the staining intensity obtained with CD4 MAb
M-T310 remained unaltered. bio = biotin; FITC = fluorescein
isothiocyanate; MIg = mouse immunoglobulin.
53 1
ANTI-CD4 THERAPY FOR RA
2.0
1.5
4TT
TI
-T
TT
lT
sr
8h 24h
d4
d7
d21 d35
PPD
1.0
0.5
0.0
0
5
10
15
20
hours after mAb injection
25
Figure 6. Concentrations of free, active antibody (with binding
activity), as measured by end-point titration and immunofluorescence analysis of sera from rheumatoid arthritis patients treated
with CD4 monoclonal antibody (mAb) M-Tl5I. Values are the mean
and SD of 12 treatment courses in 10 patients.
8
E
$
1.0
0.5
0.0
2.0
1.5
antigens was found on CD4+ cells by 2-color immunofluorescence analysis. Also, the small proportion
(<4%) of activated T cells expressing class I1 antigens
or the interleukin-2 (IL-2) receptor was not visibly
altered (data not shown).
1TT
1.0
0.5
0.0
Oh
lh
3h
time after starting treatment
t
h
c,
d
rn
g
c,
E:
d
al
0
g
0
rn
...
. . . . I .
......
I
anti-CS-FITC fluorescence intensity
.
"
3
Figure 7. In vivo fixation of complement component C5 on CD4
monoclonal antibody (mAb) M-T15 I-labeled cells from a representative rheumatoid arthritis patient treated with infusions of the
mAb. PE = phycoerythrin; FITC = fluorescein isothiocyanate.
Figure 8. In vitro proliferation of peripheral blood mononuclear
cells (PBMC) from rheumatoid arthritis patients treated with CD4
monoclonal antibody M-T151.Constant numbers of PBMC were
stimulated with tetanus toxoid (TT), Mycobycterium tuberculosis
purified protein derivative (PPD), allogeneic PBMC from 2 donors
(MLC), concanavalin A (Con A), or phytohemagglutinin (PHA). To
reveal treatment-induced changes, values were normalized for individual patients, as indicated in Patients and Methods. As a late effect
the response to PPD, but not to the other stimuli, was significantly
reduced 2 weeks and 4 weeks after cessation of treatment (P< 0.05
at day 35 versus day 0). Values are the mean and SD of 14 treatment
courses in 10 patients.
Serum concentration of M-T151. At 1 hour after
intravenous injection of 20 mg of M-T151, peak concentrations of 4.3 pg/ml (mean ? SD 1.5 f 1.O) of free,
active MAb were found. This level decreased to <5
ng/ml during the following 7 hours (Figure 6). However, the plasma levels of M-T151 varied noticeably in
individual patients.
Complement activation. In vivo binding of the
murine IgG2a MAb to T lymphocytes led to marked
complement fixation on CD4+ cells. In all 6 patients
REITER ET AL
532
lsOl
T
I
I
0
5
I
I
I
10
15
20
hours after mAb injection
I
25
Figure 9. Levels of soluble CD4 in sera from rheumatoid arthritis
patients treated with CD4 monoclonal antibody (mAb) M-T151. A
sandwich enzyme-linked immunosorbent assay was used to determine soluble CD4 levels, and recombinant soluble CD4 was used as
standard. Absorption with immobilized anti-mouse IgG completely
removed soluble CD4, indicating that all soluble CD4 was complexed with M-T151. Values are the mean and SD of 14 treatment
courses in 10 patients.
tested, immunofluorescence analysis with specific rabbit antisera revealed deposition of Cls, C3d, C4d,
C4-binding protein, as well as C5 on circulating T cells
(Figure 7). Only minimal (or no) C9 was detected
between 1 hour and 24 hours after MAb infusion. The
highest concentration of fixed complement components was observed at 3 hours after MAb infusion.
Lymphocyte proliferation. The proliferative response of lymphocytes to various mitogens and antigens was distinctly reduced at 1 hour and up to 8 hours
after MAb infusion. The response returned to pretreatment levels after 24 hours (Figure 8). As a late effect 2
weeks and 4 weeks after cessation of MAb treatment,
the proliferative response to PPD was significantly
reduced (P < 0.05 day 0 versus day 3 9 , while the
response to mitogens was unaltered.
Soluble CD4.A sensitive ELISA was developed
to determine the amount of soluble CD4 glycoprotein
generated after infusion of CD4 antibody. To assure
the sensitivity and specificity of the double-determinant
assay, suitable capture and detector antibodies were
selected from a larger panel of CD4 antibodies. A
control study of 170 sera from healthy blood donors
had demonstrated the feasibility of the assay, under
the premise that the presence of anti-mouse Igreactive antibodies could be excluded or, if present,
had been absorbed with immobilized mouse Ig. A peak
concentration of soluble CD4 could be detected at 3
hours after antibody infusion (Figure 9). The soluble
CD4 activity declined to pretreatment levels within the
ensuing 20 hours. The major part of soluble CD4
appeared to be complexed with the M-T151 antibody
that had been administered, since the soluble CD4
could be removed by absorption with anti-mouse
Ig-Sepharose (data not shown).
Human anti-mouse Ig antibodies. A major problem in clinical studies employing monoclonal antibodies of mouse origin is the generation of human antimouse Ig antibodies. In some patients, low titers of
these HAMA were present before MAb treatment was
initiated. Reactivity increased in 6 patients 2 4 weeks
after CD4 MAb therapy, but in only 3 patients did the
HAMA titer increase more than 4 titer steps. The
anti-Ig antibodies were primarily of IgA and IgG and,
to a lesser extent, IgM isotype. Of the 4 patients who
received a second course of antibody, patients 4 and 7
had a further increase in the HAMA titer, and it was
patient 7 who experienced a mild anaphylactic reaction after the last antibody infusion of the second
course. M-T151-specific IgE was detectable only in
patient 7, and only after the second course of treat-
Table 2. Titer of blocking anti-mouse IgG antibodies in rheumatoid arthritis patients treated with CD4 monoclonal antibody
M-T15 1*
Titer of blocking
antibodies
Patient
1
2
3
Course of
treatment
Total
Idiotype
specific
0
-
-
1
0
1
2
0
6
1
0
1
2
0
1
0
7
1
0
4
5
8
9
10
1
2
0
1
2
0
1
0
1
1
2
1
-
-
3
5
1
2
-
1
5
-
-
2
-
2
1
1
1
-
-
* Values are the highest titer (log, titer) observed before (course 0),
after the first (course l), and after the second (course 2) treatment
course of M-T151. Blocking antibodies were determined by
immunofluorescence-inhibition assay, before and after absorption
with isotype-matched mouse IgG. - = not detectable.
ANTI-CD4 THERAPY FOR RA
ment. Patients 1 and 8 did not develop HAMA, even
after the second course of treatment. FACS analysis
was used to determine whether the induced antimouse Ig antibodies inhibited the binding of M-T151 to
T cells. In 4 patients, low-titer M-T151-blocking antibodies were detected during the first few weeks after
antibody therapy. Only a small proportion of these
HAMA were specific for the M-T151 idiotype, since
most of the blocking activity could be absorbed with
unrelated MAb (Table 2).
DISCUSSION
According to a widely held belief, an imbalance
of T cell activity underlies the pathogenesis of rheumatoid arthritis. Though the cause of this imbalance is
not at all clear, indirect evidence points to the CD4+
lymphocyte subset as being mainly responsible. Of the
known subpopulations within the CD4 subset, it is the
4B4+ (CD29), UCHLl+ (CD45RO) “memory T cell”
subpopulation that appears to be increased during
active phases of RA (29). Increased activity of T cells
may lead to an excessive secretion of growth factors
and lymphokines such as GM-CSF (granulocytemacrophage colony-stimulating factor) and IL-6
(30,31). Because the CD4 glycoprotein is an adhesion
molecule that is required for intercellular contact as
well as for activation of T cells, this molecule may
serve as a suitable target for blocking antibodies. The
immunosuppressive effects of CD4 antibodies in animal models of autoimmune diseases is widely documented (32).
As to the therapeutic efficacy in humans, we
observed distinct and prolonged improvement in at
least 3 clinical parameters in 6 patients and a “moderate” response in 1 patient. Statistical analysis confirmed a tendency toward improvement, and the relief
of clinical symptoms was correlated with an objective
decrease in the CRP values. The inefficacy of the
antibody treatment in 3 patients, who exhibited a
rather high inflammatory activity, may indicate the
possibility that there are phases of the disease which
are not governed by T lymphocytes.
In the patients with a good response, the relief
of clinical symptoms cannot be ascribed to additional
pharmacotherapy, since the standardized dosage of
prednisolone that was allowed (10 mg/day, from 4
weeks before until 5 weeks after the initiation of MAb
therapy) was lower than the dosage taken by any of the
10 patients before entering the study. Similarly, late
effects of former DMARD treatment can also be
533
excluded because of the “washout” period of more
than 4 weeks. During the 6-month followup period, no
DMARDs were given. NSAIDs were given only to
nonresponders and not before day 35. It is noteworthy
that corticosteroid dosages could be further reduced in
4 patients during the posttreatment period.
In view of the variable course of RA, the
observed therapeutic effects in this small group of
patients must be interpreted cautiously. However, the
characteristic change in clinical parameters (as shown
for the Ritchie articular index) after MAb treatment is
evidence against there having been a spontaneous
variation in disease activity. Though patients may
have derived benefit from the hospitalization, the
induction of a longer-lasting improvement by only 1
week of hospitalization seems unlikely. To clarify
whether this is a factor, a double-blind placebocontrolled study of this treatment administered on an
outpatient basis has been initiated. Further studies are
necessary in order to determine the optimal dose and
regimen.
With regard to the immunologic effects of antiCD4 treatment, the selective decrease in circulating
CD4+ cells 1 hour after MAb infusion was the most
impressive event. At this time, fixation of complement
on the residual circulating CD4+ lymphocytes was
clearly demonstrated. Thus, it is reasonable to assume
that opsonization of the anti-CDkoated cells accelerates trapping in the monocyte phagocytic system. It
has been shown in vitro that antilymphocyte MAb can
induce opsonization and adherence of lymphocytes to
complement receptor-positive cells (33), while in vivo,
it has been directly demonstrated that antibodylabeled cells are trapped in lung, liver, and spleen.
Complement-mediated lysis seems to be unlikely; so
far, a lytic effect has only been seen with few selected
MAb-recognizing antigens abundantly expressed on
the cell surface, such as Campath-1 (CDw52) (34), and
for certain combinations of MAb (35). At present, it is
unclear whether the CD4+ cells that reappear after the
initial depletion represent newly formed lymphocytes
recruited from lymphoid organs or whether they represent temporarily immobilized cells having resynthesized CD4 antigen that had been shed.
Determination of levels of free active MAb in
sera revealed almost complete clearing of MAb within
8 hours. This short half-life contrasts with the in vivo
half-life of other murine IgG2a MAb used in tumor
therapy, such as C017-1A, which does not bind to
peripheral blood cells (36), or the CD3 MAb OKT3,
which accumulates in the serum after infusion of only
534
5 mg daily (ref. 37 and Reiter et al: unpublished
observations). On the other hand, MAb BMA031
directed against the T cell antigen receptor-CD3 complex shows kinetics similar to those of M-T151(38). AS
demonstrated with MAb M-T3 10, which recognizes a
different epitope, the CD4 on circulating T cells was
not modulated but was partially occupied by M-T151
at 8 hours and 24 hours after infusion. The number of
circulating CD3+, CD4-/CD8- cells also did not
increase. Shed CD4 may contribute to the neutralization of free MAb, but soluble CD4 bound to M-T151
was detectable only for a short time after MAb infusion, and the concentration was low, since blocking of
M-T3 10 binding activity was not found.
Antibody-induced shedding of T cell antigens
has been previously described in studies concerning
other T cell antigens, such as CD5 (39) and the IL-2
receptor CD25. Whereas some surface molecules,
such as CD8, CD23, and CD25, can be found in soluble
form in healthy individuals (40,41), we could detect
soluble CD4 only after infusion of a CD4 MAb.
Whether soluble CD4 generated during antibody treatment is immunosuppressive remains to be shown.
Recombinant soluble CD4 in a human immunodeficiency virus-neutralizing concentration was not immunosuppressive in vitro (42).
A crucial question is to what extent and how
long-lasting is the M-TI5 1-induced impaired lymphocyte function in vivo. Although a 20-mg dose of
antibody was injected each day for 1 week in this
group of patients, compared with the 10-mg dosage
used in the study published by Walker et a1 (14),
stimulation of T cells by soluble antigens and by
allogeneic cells was still only transiently diminished,
and pretreatment levels were regained within 24 hours.
However, at day 21 and day 35, in vitro responses to
PPD were significantly decreased. At this time also,
lower numbers of circulating CD4+ T lymphocytes
were found. Hafler et a1 (12) observed a lower level of
both spontaneous and pokeweed mitogen-induced
IgG production for up to 2 weeks after CD4 MAb
treatment.
Because mice treated with CD4 MAb developed a type-specific tolerance to heterologous IgG, a
careful analysis of the anti-mouse Ig response was
performed (43). As outlined in the Results section,
plasma samples from 7 of the RA patients prior to
treatment contained low anti-mouse Ig reactivity, with
a titer of -2-4 above background. Using the same
assay, we have detected low-titer anti-mouse Ig serum
antibodies in -5% of a group of 170 healthy blood
REITER ET AL
donors. In 6 of the 10 RA patients assessed here,
anti-Ig antibody levels were increased 2 weeks after
treatment, but decreased rapidly during the following
weeks, including the antibodies of IgG isotype. Blocking antibodies were detectable in 4 patients, and only
for a short time; only a small portion of the blocking
antibodies was directed against antiidiotypic determinants (submitted for publication).
The presence of anti-mouse Ig antibodies does
not necessarily obviate the therapeutic effect of the
infused antibody. On the contrary, it has been reported by Jonker and den Brok (44) that depletion of
CD4+ cells in a CD4 MAb-treated rhesus monkey
occurred only after the formation of antibodies against
murine Ig. Although all patients selected for a second
course of MAb treatment had developed blocking
antibodies after the first course, these antibodies were
no longer detectable at the time the second course of
treatment was started a few months later. Except for
one IgE-mediated mild anaphylactic reaction in 1
patient after the last infusion of the second course of
treatment, no side effects were seen in these patients.
Nevertheless, replacing the murine Fc portion and the
murine constant regions of the light chains with the
corresponding human peptide segments may considerably improve the therapeutic efficacy and tolerability
of the CD4 antibody.
There was no clinical manifestation of immunosuppression as has been described in patients treated
with CD3 MAb. Whether the remaining unaffected
CD8+ T cells can still provide protection against
infection, particularly against viruses, is still to be
studied. So far, no episodes of herpes simplex or
herpes zoster exacerbations have been observed during or after CD4 antibody treatment.
REFERENCES
Paulus HE, Machleder HI, Levine S, Yu DTY, Mac-
Donald NS: Lymphocyte involvement in rheumatoid
arthritis: studies during thoracic duct drainage. Arthritis
Rheum 20:1249-1262, 1977
Kotzin BL, Strover S, Engleman EG, Calin A, Hope
RT, Kausa GS, Terrell CP, Kaplan HS: Arthritis with
total lymphoid irradiation. N Engl J Med 305:969-976,
1981
Strober S, Tanay A, Field E, Hope RT, Calin A,
Engleman EG, Kotzin B, Brown BW, Kaplan HS:
Efficacy of total lymphoid irradiation in intractable rheumatoid arthritis. Ann Intern Med 102:441-449, 1985
Nusslein HG, Herbst M, Manger BJ, Gramatzki M,
Burmester GR, Fritz H, Sauer R, Kalden JR: Total
ANTI-CD4 THERAPY FOR RA
lymphoid irradiation in patients with refractory rheumatoid arthritis. Arthritis Rheum 28: 1205-1210, 1985
5 . Kriiger K, Demant T, Kellner H , Schattenkirchner M:
Die Lymphozytapherese in der Behandlung der chronischen Polyarthritis: Einfluss auf klinische und laborchemische Parameter sowie auf die Lymphozytenfunktion.
Z Rheumatol46:303-310, 1987
6. Karsh J, Klippel JH, Plotz PH, Decker JL, Wright DG,
Flye MW: Lymphapheresis in rheumatoid arthritis. Arthritis Rheum 24:867-873, 1981
7. Veys EM, Hermanns P, Verbruggen G, Schindler J,
Goldstein G: T lymphocytes in blood and synovial fluid
in rheumatoid arthritis. Lancet I:225-226, 1982
8. Duke 0, Panayi GS, Janossy G, Poulter LW: An immunohistological analysis of lymphocyte subpopulations
and their microenvironment in the synovial membranes
of patients with rheumatoid arthritis using monoclonal
antibodies. Clin Exp Immunol 49:22-30, 1982
9. Ranges GE, Sriram S, Cooper SM: Prevention of type I1
collagen-induced arthritis by in vivo treatment with
anti-L3T4. J Exp Med 162:1105-1 110, 1985
10. Shizuru JA, Taylor-Edwards C, Banks BA, Gregory
AK, Fathman CG: Immunotherapy of the nonobese
diabetic mouse: treatment with an antibody to T-helper
lymphocytes. Science 240365-62,
1988
11. Van Lambalgen R, Jonker M: Experimental allergic
encephalomyelitis in rhesus monkeys. 11. Treatment of
EAE with anti-T lymphocyte subset monoclonal antibodies. Clin Exp Immunol 67:305412, 1987
12. Hafler DA, Ritz J, Schlossman SF, Weiner HL: AntiCD4 and anti-CD2 monoclonal antibody infusions in
subjects with multiple sclerosis. J Immunol 141:131-138,
1988
13. Morel P, Cordier G, Vincent C, Traeger J , Carosella E,
Revillard J-P: Anti-CD4 monoclonal antibody therapy in
humans (abstract), 7th International Congress of Immunology, Gesellschaft fur Immunologie. Stuttgart, Gustav
Fischer Verlag, 1989
14. Walker C, Herzog C, Rieber P, Riethmuller G, Miiller
W, Pichler WJ: Anti-CD4 antibody treatment of patients
with rheumatoid arthritis. 11. Effect of in vivo treatment
on in vitro proliferative response of CD4 cells. J Autoimmunol2:643449, 1989
15. Herzog C, Walker C, Pichler W, Aeschlimann A, Wassmer P, Stockinger H, Knapp W, Rieber P, Muller W:
Monoclonal anti-CD4 in arthritis. Lancet 11:1461-1462,
1987
16. Reiter C, Federle C, Feucht H , Held E, Schattenkirchner M, Riethmuller G, Rieber EP: Immunopharmacological aspects of the treatment with monoclonal CD4
antibodies (abstract). Immunobiology 178: 158, 1988
17. Barber EK, Dasgupta JD, Schlossman SF, Trevillyan
JM, Rudd CE: The CD4 and CD8 antigens are coupled
to a protein-tyrosine kinase ( ~ 5 6 " ~that
) phosphorylates
535
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
the CD3 complex. Proc Natl Acad Sci USA 86:32773281, 1989
Janeway CA: The role of CD4 in T-cell activation:
accessory molecule or co-receptor? Immunol Today
10:234-238, 1989
Doyle C, Strominger JL: Interaction between CD4 and
class I1 MHC molecules mediates cell adhesion. Nature
330:256-259, 1987
Arnett FC, Edworthy SM, Bloch DA, McShane DJ,
Fries JF, Cooper NS, Healey LA, Kaplan SR, Liang
MH, Luthra HS, Medsger TA Jr, Mitchell DM, Neustadt DH, Pinals RS, Schaller JG, Sharp JT, Wilder RL,
Hunder GG: The American Rheumatism Association
1987 revised criteria for the classification of rheumatoid
arthritis. Arthritis Rheum 31:315-324, 1988
Ritchie DM, Boyle JA, McInnos JM, Jasani MK, Dalakos TG, Grieveson P, Buchanan WW: Clinical studies
with an articular index for the assessment of joint
tenderness in patients with rheumatoid arthritis. Q J
Med 147:393406, 1968
Steinbrocker 0, Traeger CH, Batterman RC: Therapeutic criteria in rheumatoid arthritis. JAMA 140:659-662,
1949
Reinherz EL, Haynes EF, Nadler LM, Bernstein ID:
Leukocyte Typing 11. New York, Springer Verlag, 1985
Peterson A, Seed B: Genetic analysis of monoclonal
antibody and HIV binding sites on the human lymphocyte antigen CD4. Cell 54:65-72, 1988
Jameson BA, Rao PE, Kong LI, Hahn BH, Shaw GM,
Hood L E , Kent SBH: Location and chemical synthesis
of a binding site for HIV-1 on the CD4 protein. Science
240: 1335-1339, 1988
Pirron U, Schlunk T , Rieber EP: Interference of CD4
and new T-cell antibodies with the antigen induced
Ca2+-mobilization in T-cells, Leucocyte Typing IV.
Edited by W Knapp, B Dorken, WR Gilks, E P Rieber,
RE Schmidt, H Stein, AEGK von dem Borne. Oxford,
Oxford University Press, 1989
Bicker U, Dippold WG, Emmrich F , Hasskarl H , Heir
W, Herbeck A, Kalden J, Kirchner H, Kleinsorge F,
Knapp W, Kraft V, Kubanek B, Lower J, Meyer Z.
Buschenfelde KH, Muller-Ruchholtz W, Oberhausen E,
Peter H, Rieber EP, Schlumberger HD, SchulteWissermann H, Seiler FR, Stein H, Waller H-D, Wick
G, Wilms H: Empfehlungen fur die Herstellung und
Priifung in vivo applizierbarer monoklonaler Antikorper. Dtsch Med Wochenschr 112:194-198, 1978
Pinals RS, Masi AT, Larsen RA, and the Subcommittee
for Criteria of Remission in Rheumatoid Arthritis of the
American Rheumatism Association Diagnostic and
Therapeutic Criteria Committee: Preliminary criteria for
clinical remission in rheumatoid arthritis. Arthritis
Rheum 24:130&1315, 1981
Emery P, Gentry KC, Mackay IR, Muirden KD, Rowley
M: Deficiency of the suppressor inducer subset of T
536
30.
31.
32.
33.
34.
35.
36.
lymphocytes in rheumatoid arthritis. Arthritis Rheum
30:849-856, 1987
Xu WD, Firestein GS, Taettle R, Kaushansky K, Zvaifler NJ: Cytokines in chronic inflammatory arthritis. 11.
Granulocyte-macrophage colony-stimulating factor in
rheumatoid synovial effusions. J Clin Invest 83:876-882,
1989
Nawata Y,Eugui EM, Lee SW, Allison AC: IL-6 is the
principal factor produced by synovia of patients with
rheumatoid arthiritis that induces B-lymphocytes to
secrete immunoglobulins. Ann NY Acad Sci 557:230238, 1989
Waldmann H: Manipulation of T cell responses with
monoclonal antibodies. Ann Res Immunol 7:407-444,
1989
Sugita K, Majdic 0, Stockinger H, Holter W, Burger R,
Knapp W: Induction of human complement activation
without cytolysis by mouse monoclonal antibodies to
human leukocyte antigens. Transplantation 43 :570-574,
1985
Bindon CI, Hale G, Waldmann H: Importance of antigen
specificity for complement-mediated lysis by monoclonal antibodies. Eur J Immunol 18:1507-1514, 1988
Bindon CI, Hale G, Clark M, Waldmann H: Therapeutic
potential of monoclonal antibodies to the leukocytecommon antigen. Transplantation 40538-544, 1985
Khazaeli MB, Saleh MN, Wheeler RH, Huster WJ,
Holden H, Carrano R, LoBuglio AF: Phase I trial of
multiple large doses of murine monoclonal antibody
C017-1A. 11. Pharmacokinetics and immune response.
JNCI 80~937-942, 1988
REITER ET AL
37. Zwaan FE, Gratama JW, Goldstein G, Jansen J: Treatment of acute graft-versus-host disease with monoclonal
antibody OKT3.PAN. Exp Hematoll2 (suppl15):64-85,
1984
38. Weschka M, Smely S, Dendorfer U, Hillebrand G,
Kurrle R, Land W, Hammer C: Prophylactic use of the
new monoclonal antibody BI 51.013 (BMA031) in clinical kidney transplantation: pharmacokinetics, antibody
response, and cytokine release (abstract), 7th International Congress of Immunology, Gesellschaft fur Immunologie. Stuttgart, Gustav Fischer Verlag, 1989
39. Miller RA, Maloney DG, McKillop J, Levy R: In vitro
effects of murine hybridoma monoclonal antibody in a
patient with T cell leukemia. Blood 58:78-82, 1981
40. Fujimoto J, Levy S, Levy R: Spontaneous release of the
Leu-2 (T8) molecule from human cells. J Exp Med
159:752-766, 1983
41. Nelson DL: Soluble ILZreceptors: analysis in normal
individuals and in certain disease states. Fed Proc 45:
377-400, 1986
42. Hussey RE, Richardson NE, Kowalski M, Brown NR,
Chang H-C, Siliciano RF, Dorfman T, Walker B, Sodroski J, Reinherz EL: A soluble CD4 protein selectively inhibits HIV replication and syncytia formation.
Nature 331:78-81, 1988
43. Benjamin RJ, Waldmann H: Induction of tolerance by
monoclonal antibody therapy. Nature 320:449-451, 1986
44. Jonker M, den Brok JHAM: Idiotype switching of
CD4-specific monoclonal antibodies can prolong the
therapeutic effectiveness in spite of host anti-mouse IgG
antibodies. Eur J Immunol 17: 1547-1553, 1987
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