anti-CD20 Monoclonal Antibody Treatment Research Articles

Prediction of graft prolongation by mixed lymphocyte culture following anti-CD4 monoclonal antibody treatment among different donor-recipient combinations.

This study was conducted to examine whether mixed lymphocyte culture (MLC) could be used as a predictor of the efficacy of anti-CD4 monoclonal antibody (MAb) immunosuppression in vivo in a mouse model. C57BL/10 or BALB/c hearts were transplanted into C3H.He recipients. Anti-CD4 MAb administration prolonged graft survival, but there was a clear difference between the two donor-recipient combinations studied, the median survival time (MST) being >100 days in the C57BL/10 --> C3H group, and 17 days in the BALB/c --> C3H group. Anti-CD8 MAb prolonged the survival of C57BL/10 hearts slightly to a MST of 22 days, but the BALB/c hearts were rejected at control rates. Combining anti-CD4 and anti-CD8 antibody therapy prolonged the survival of C57BL/10 hearts indefinitely, but had little effect on the survival of BALB/c grafts, achieving an MST of only 24 days. Next, MLCs were performed in the presence and absence of the MAbs and compared with the graft survival data. The inhibition rates in the MLC, being the C3H lymph node cell responder, correlated well with graft survival. When three kinds of C3H responder cells, namely lymph node (LN) cells, T cells, and CD4+ cells, were examined to determine which was the most suitable for predicting graft survival, the MLC results showed that the responses of LN cells correlated most closely with graft outcome. In conclusion, MLC using LN cells as the responder is a useful tool for predicting allograft survival induced by anti-CD4 MAb therapy.

Prediction of Graft Prolongation by Mixed Lymphocyte Culture Following Anti-CD4 Monoclonal Antibody Treatment Among Different Donor-Recipient Combinations

Prediction of Graft Prolongation by Mixed Lymphocyte Culture Following Anti-CD4 Monoclonal Antibody Treatment Among Different Donor-Recipient Combinations

A Synthetic CD4-CDR3 Peptide Analog Enhances Bone Marrow Engraftment Across Major Histocompatibility Barriers

AbstractThe efficacy of a synthetic peptide analog mimicking the CDR3-D1 domain of the CD4 molecule was investigated in murine models of allogeneic bone marrow engraftment after transplantation across major histocompatibility complex (MHC) barriers. A single dose of a CD4-CDR3 peptide analog was administered at the time of marrow transplantation to three different allogeneic mouse strain combinations after appropriate sublethal total body irradiation: (1) B10.BR → C57BL/6J (B6), a full allogeneic MHC difference; (2) (B6xDBA/2)F1 → (B6xCBA)F1 , a haploidentical MHC combination; and (3) B6.C-H2bm12 → B6-Ly5.2, involving only a MHC class II difference. Donor-host chimerism was assessed after 1 and 2 months posttransplantation by flow cytometric analysis of spleen and/or lymph node cells. Peptide-treated animals in all three strain combinations exhibited significantly enhanced donor lymphoid engraftment, which was similarly reflected in the total lymphocyte compartment and its T-cell (CD4+, CD8+) and B-cell subsets. In addition, peptide-treated mice in the haploidentical and MHC class II-mismatched strain combinations exhibited prolonged tolerance of both donor and syngeneic host-type tail skin grafts while rejecting third-party allogeneic grafts, thus supporting the reconstitution of immunocompetence in these chimeras. Lymphocytes from the peptide-treated haploidentical chimeric mice also displayed donor-specific tolerance upon stimulation in a one-way mixed lymphocyte reaction. In a 6-day colony-forming unit–granulocyte-macrophage (CFU-GM) assay to quantitate the level of hematopoietic cell engraftment in both the haploidentical and class II-disparate strain combinations, bone marrow cells from the peptide-treated mice exhibited significant increases in CFU-GM compared with the saline-treated control groups. Finally, early multiple treatments with the peptide after transplantation significantly enhanced donor chimerism in donor-presensitized recipient mice across the MHC class II barrier and proved to be significantly more effective than anti-CD4 monoclonal antibody treatment. These results indicate that the structure-based CD4-CDR3 peptide analog may represent a valuable approach to the inhibition of graft rejection after MHC-mismatched bone marrow transplantation.

Synergism of CTLA4-IG and anti-CD4 monoclonal antibody treatment in a rat kidney transplant model

Synergism of CTLA4-IG and anti-CD4 monoclonal antibody treatment in a rat kidney transplant model

Indefinite survival of neural xenografts induced with anti-CD4 monoclonal antibodies

Xenografts of neural tissue are usually rapidly rejected when transplanted into the central nervous system of adult recipient animals. This study has examined the cell mediated immune response to both concordant (between closely related species) and discordant (between distantly related species) neural xenografts in the mouse, and has investigated the role of the CD4 + and CD8 + T lymphocyte subsets in this process using monoclonal antibodies specific for the CD4 and CD8 cell surface glycoproteins. We have established that: (1) in this model system concordant neural xenograft rejection occurs within 15–30 days; however, xenograft survival can be dramatically prolonged with CD4 +, but not CD8 +, T lymphocyte depletion; (2) the administration of two successive courses of a high dose of anti-CD4 monoclonal antibody treatment results in indefinite concordant neural xenograft survival; (3) the mechanism by which the high dose anti-CD4 monoclonal antibody therapy appears to function involves the depletion of intrathymic CD4 + cells; (4) anti-CD4 monoclonal antibody treatment enhances discordant neural xenograft survival, to beyond 60 days in many cases. These results demonstrate that CD4 + T lymphocytes are of central importance in the immune response to both concordant and discordant neural xenoantigens. Thus the use of anti-CD4 monoclonal antibody therapy is an effective strategy to prolong significantly the survival of xenogeneic neural transplants. Furthermore this treatment caused no obvious deleterious side-effects. These findings have implications for future cross-species studies in experimental neurobiology and, possibly, in clinical neural transplantation.

Two levels of help for B cell alloantibody production.

We have examined whether T cell stimulation by direct or indirect pathways contributes to alloantibody production by B cells after major histocompatibility complex (MHC)-disparate skin graft rejection in mice. Experiments were performed using normal mice, MHC class II-deficient mice, MHC class II-deficient mice with an intact peripheral CD4+ cell population (due to expression of class II antigens only on thymic epithelium), mice lacking the cytoplasmic tail of their MHC class II antigens, and mice depleted of CD4+ cells by anti-CD4 monoclonal antibody treatment. Depletion of recipient CD4+ cells reduced alloantibody production to barely detectable levels. Absence of donor MHC class II antigens did not affect the production of either immunoglobulin (Ig)M or IgG antibodies directed at class I alloantigens. Absence of recipient MHC class II antigens, however, led to production of only IgM but not IgG antibodies, even if the recipients had an intact CD4+ cell population. Absence of the cytoplasmic tail of the recipient's MHC class II antigens led to the production of slightly reduced amounts of IgG antibody. These findings indicate that (a) CD4+ cells are essential helper cells for B cell alloantibody production; (b) production of IgM alloantibody can occur with help from CD4+ cells, which recognize either donor class II antigens or modified recipient class II antigens; (c) isotype switching from IgM to IgG alloantibody requires help from CD4+ cells activated by antigens presented by recipient MHC class II molecules; and (d) the cytoplasmic domain of the recipient MHC class II molecules may be involved in the mechanism that leads to isotype switching by B cells. Thus, there are two levels of CD4-mediated help available for B cells responding to alloantigens: one (involving a noncognate interaction) can produce B cell activation, and a second (involving a cognate interaction) is required for differentiation and IgG alloantibody production.

Manipulation of xenogeneic skin and/or renal graft survival in the rat-mouse concordant combination by portal vein pretransplant transfusion

We have examined whether portal venous pretransplant transfusion, which has been shown to produce prolongation of rodent vascularized (small intestine, kidney) and nonvascularized (skin) allografts, in the absence of other nonspecific immunosuppression, can produce similar graft prolongation in animals receiving vascularized or nonvascularized xeno- (not allo-) grafts. Rat kidney or skin grafts were transplanted into mice after portal venous pretreatment with rat or mouse spleen cells. Animals in some groups received additional immunosuppressive regimens including drug therapy (methotrexate, cyclosporin A) or monoclonal antibody treatment (anti-CD4, anti-CD8). Animal survival and serum creatinine was followed daily, and lymphoproliferation, cytokine production (including cytokine mRNA in grafted mice) and anti-xenograft antibody production was measured at distinct time points postgrafting. Both portal venous pretransplant transfusion and anti-CD4 monoclonal antibody treatment led to increased graft survival. However, unlike the rodent allograft model, graft survival in these animals was not simply explained by altered Th1/Th2 ratios. Other mechanism(s), possibly including xenoantibody production, are likely of importance in the regulation of xenograft rejection.

Reduction of synovial inflammation after anti‐cd4 monoclonal antibody treatment in early rheumatoid arthritis

To study the effect of chimeric anti-CD4 monoclonal antibody (MAb) therapy on synovial inflammation, in order to interpret the clinical experience with anti-CD4 treatment. The immunohistologic features of synovial biopsy specimens before and 4 weeks after anti-CD4 MAb (cM-T412) therapy were studied in patients with rheumatoid arthritis. The patients received intravenous doses of either placebo (n = 1) or 10 mg (n = 4), 25 mg (n = 2), or 50 mg (n = 1) of cM-T412 daily for 5 consecutive days. Although the patients did not experience clinical improvement, significant decreases in the number of circulating CD4+ cells, the degree of synovial inflammatory infiltration, and the mean scores for expression of adhesion molecules were found in the 7 patients 4 weeks after receiving cM-T412. The scores for infiltration with CD4+ and other inflammatory cells were particularly reduced following treatment with either 25 mg or 50 mg cM-T412. Cytokines, such as interleukin-1 beta and tumor necrosis factor alpha, could still be detected in the synovial tissue after treatment. The decline in the numbers of inflammatory cells and adhesion molecules in synovial tissue after CD4+ cell depletion supports the view that CD4+ T cells orchestrate local cellular infiltration. The lack of clinical effect of anti-CD4 therapy might be explained by an insufficient decrease in the number of synovial CD4+ cells and by the persistence of cytokines. Determination of whether more adequate dosing would lead to a clinical improvement must await further study.

Effect of Long-Term Anti-CD4 or Anti-CD8 Treatment on the Development of lpr CD4 − CD8 − Double Negative T Cells and of the Autoimmune Syndrome in MRL- lpr / lpr Mice

We have determined the effect of anti-CD4 or anti-CD8 monoclonal antibody (mAb) treatment from birth on the generation of the lpr CD4- CD8- double-negative (DN) T cell subset and on the development of lupus-like autoimmune syndrome in MRL-lpr/lpr mice. Both anti-CD4 and anti-CD8 mAb treatments resulted in a marked inhibition of lymph-adenopathy, whereas the development of the lpr DN T cells and of the lupus-like autoimmune syndrome strikingly differed in these two groups of mice. The treatment with anti-CD8 mAb almost completely blocked the appearance of the lpr DN T cells without any significant effect on the development of lupus-like autoimmune syndrome in MRL-lpr/lpr mice. In contrast, mice treated with anti-CD4 mAb failed to develop a lupus-like syndrome, while they still developed the lpr DN T cell subset, the predominant population in their lymph nodes, although absolute numbers were markedly diminished. Our results support the idea that CD8+ T cells are a major source of the lpr DN T cells, and that the lpr DN T cells play a minor, if any, role in the pathogenesis of lupus-like autoimmune syndrome in MRL-lpr/lpr mice.

Effects of Early and Late Treatment with Anti-CD4 Monoclonal Antibody on Autoimmune Disease in MRL/MP- Ipr/ Ipr Mice

MRL/Mp- Ipr/ Ipr (MRL/Ipr) mice spontaneously develop a systemic autoimmune disease, characterized by vasculitis, lymphadenopathy, glomerulortephritis, and autoantibody formation, with target organ inflammatory lesions composed largely of CD4 + (helper) T cells. Previous reports have demonstrated that anti-CD4 monoclonal antibody (mAb) treatment of MRL/Ipr mice from 1 to 5 months of age resulted in a dramatic reduction in both the frequency and the severity of autoimmune disease. In order to investigate the effects of early, short-course and late, short-course anti-CD4 mAb therapy on the autoimmune disease in MRL/Ipr mice, groups of 12 to 15 animals were treated with weekly intraperitoneal injections according to one of four regimens: (i) anti-CD4 mAb from age 1 to 5 months (continuous treatment); (ii) anti-CD4 mAb from age 1 to 3 months (early treatment); (iii) anti-CD4 mAb from age 3 to 5 months (late treatment); and (iv) either normal saline or rat immunoglobulin (control treatment). Continuous treatment resulted in a dramatic reduction of both frequency and severity of the autoimmune disease, as demonstrated histologically and serologically. Early treatment also resulted in a significant reduction in autoimmune disease, while late treatment had little effect. Glomerulonephritis was detected in none of the animals in the continuously treated group ( P < 0.05), 38% of those in the early-treated group ( P = < 0.05), 92% of the late-treated group, and 100% of controls. The titer of antinuclear antibodies, of anti-dsDNA antibodies, and total immunoglobulin levels were all significantly reduced in the continuous-treatment and early-treatment groups, but not in the late-treatment group. Murine antibodies to rat anti-CD4 mAb were present in the late-treatment group. These results indicate that early short-course anti-CD4 mAb treatment of MRL/Ipr mice is effective in ameliorating the autoimmune disease in this model, while late-treatment is ineffective, probably due to the induction of antibody directed against anti-CD4 mAb itself.

The effect of anti-CD4 monoclonal antibody treatment on immunopathological changes in psoriatic skin

Recent clinical studies which showed the therapeutic effect of cyclosporin A and of anti-CD4 MoAb emphasized the role of activated CD4 + T cells infiltrating the lesional skin in the pathogenesis of psoriasis. The aim of the present study was to analyze the mode of action of anti-CD4 MoAb in 3 psoriatic patients who experienced an anti-CD4 MoAb-induced clinical improvement maximal 3–4 weeks after the onset of an 8-day therapy. We evaluated the effect of anti-CD4 MoAb treatment on the phenotype of resident and passenger inflammatory skin cells in lesional skin samples. We observed a gradual improvement of 3 out of 4 histopathologic features including parakeratosis, papillomatosis and acanthosis. In the dermis there was no modification in the density of the dermal mononuclear cell infiltrate, which consisted mainly of CD3 +, CD45RO +, TCR αβ +, CD11a +, HLA-DR +T cells with a CD4 CD8 cell ratio of 1.5 1 . Therefore as previously observed for peripheral blood mononuclear cells, the number of CD4 +T cells infiltrating the dermis remained unaffected by the treatment. In contrast, CD4 MoAb treatment was associated with drastic changes in the epidermis. These included a decrease in both CD4 + and CD8 + epidermal T cell infiltrate, diminished numbers of ICAM-1 + and HLA-DR + keratinocytes and restored numbers of CD1a + epidermal Langerhans cells. We conclude from this study that clinical improvement of psoriasis by anti-CD4 MoAb therapy paralleled: (1) a decrease in epidermal T cells, and (2) a down-regulation of keratinocyte activation markers (ICAM-1 and HLA-DR). These results suggest that the observed changes are secondary to down-regulation of inflammatory cytokine production by T cells in situ.

Anti-CD4 monoclonal antibody treatment combined with total lymphoid irradiation and cyclosporin A in hamster-to-rat cardiac transplantation: Analysis of lymphocyte subsets and anti-donor xenoantibodies

Combined treatment with total lymphoid irradiation and cyclosporin A results in prolonged graft survival in concordant xenogeneic cardiac transplantation, but reproducible long-term graft acceptance has proved to be difficult. Anti-CD4 monoclonal antibody treatment has been successful in inhibiting heart graft rejection in allogeneic models. Used as monotherapy in a concordant xenogeneic model for pancreatic islet transplantation, prolonged graft survival has been reported; however, no beneficial effect on primarily vascularized heart grafts was noted. The object of this investigation was to combine these treatment strategies with respect to reproducible long-term hamster heart graft survival in rats, to monitor the effect on lymphocyte subpopulations, and to determine possible anti-donor antibody formation correlated to time of rejection. Graft survival after combined preoperative total lymphoid irradiation and postoperative cyclosporin A + anti-CD4 monoclonal antibody treatment was prolonged from 14 to > 100 days (compared to spontaneous graft survival of three to four days), with long-term graft function in four of 19 recipients. Total white blood counts in the postoperative course were characterized by an unproportional increase of Ig+ cells and an incomplete recovery of CD4+ cells. Flow-cytometric analysis of anti-donor antibodies showed low levels of performed antibodies and increasing amounts of strain-, but not donor-specific antibodies, correlated to the time of rejection. Long-term survivors with functioning grafts at the time of sacrifice had an initially moderate antibody increase with subsequent decrease to baseline levels. Our results indicate that total lymphoid irradiation combined with cyclosporin A and anti-CD4 monoclonal antibodies can prolong graft survival significantly in concordant hamster-to-rat cardiac xenotransplantation. Graft rejection seems to be of a delayed antibody-mediated type, where the inhibition of T cell help for antibody formation may play a significant role.

A chimeric glycoprotein of human respiratory syncytial virus termed FG induces T-cell mediated immunity in mice

Popliteal lymph node cells taken from mice vaccinated with the FG glycoprotein were exposed in vitro to respiratory syncytial virus (RSV) antigens. Proliferation to FG or RSV antigens was blocked with anti-CD4 monoclonal antibody treatment. FG-vaccinated mice developed classical late delayed type hypersensitivity (DTH) reactions when exposed to FG antigen in vivo.

Anti-CD4 antibody treatment of patients with rheumatoid arthritis: I. Effect on clinical course and circulating T cells

Eight patients with arthritis (seven with rheumatoid, one with psoriatic arthritis) were treated for 7 d with a daily injection of 10 mg of mouse monoclonal anti-CD4 antibodies (three with VIT4, five with MT151). With the exception of a short-lasting low-grade fever in one patient, no side effects were observed. Clinical symptoms (morning stiffness, number of swollen joints, pain assessment and Ritchie articular index) improved in all patients within 7 d of treatment. Improvement lasted from 3 weeks to greater than or equal to 5 months (mean approximately 11 weeks). Rheumatoid factors, immune complexes and other laboratory parameters did not change during or after treatment. Skin reactivity to recall antigens was suppressed in four out of six patients during treatment but returned to pretreatment levels within 6 weeks. Immunofluorescent analysis revealed a short-lasting drop of T cells, mainly of the CD4+ CDw29+ subset, but monocytes were also affected. The injected antibody was detectable on circulating cells for about 10 h. Within 20-24 h, the cell distribution returned to pretreatment levels. In six out of eight patients an anti-mouse-Ig response was seen. We conclude that mouse anti-CD4 monoclonal antibody (MoAb) treatment is well tolerated and that the cellular immunological changes observed are short-lasting. The low incidence of side effects may justify further clinical studies to evaluate the clinical efficacy of such treatment.

Recurrence of Insulitis in the Nod Mouse After Early Prolonged Anti-CD4 Monoclonal Antibody Treatment

The treatment of young NOD mice with continuous anti-CD4 mAb administration has previously been shown to prevent insulitis from developing. It has also been shown that insulitis, once present, can be abolished by anti-CD4 mAb treatment, but that the insulitis recurs after treatment stops. We have studied the effect of treating young NOD/Wehi mice with a limited course of anti-CD4 mAb to determine if any long term prevention of insulitis ensued. Anti-CD4 mAb (H129.19) was given i.p. weekly to 10 female NOD/Wehi mice from 25-100 days of age. Control mice received no treatment. The pancreas of the mice were examined histologically at 100 and 180 days of age and severity of insulitis graded from 0-100%. The insulitis grade in treated mice at 100 days was 5 +/- 2% compared with 29 +/- 26% in age matched control mice (p less than 0.01). At 180 days of age the insulitis grade in treated mice was 29 +/- 23% compared with 47 +/- 22% in aged matched controls (NS). Thus insulitis was prevented from developing by early treatment with anti-CD4 mAb but developed later when treatment was stopped. Early anti-CD4 mAb treatment does not therefore appear to produce a tolerant state so averting insulitis development and, insulitis can still develop in older NOD mice.

Intrathymic deletion of self-reactive cells prevented by neonatal anti-CD4 antibody treatment.

T-cell differentiation in the thymus involves the coordinate expression of genes encoding the alpha and beta chains of the major histocompatibility complex-restricted heterodimeric antigen receptor (TCR) complex, as well as other functionally important molecules such as CD4 and CD8. The repertoire of TCR expressed by T cells is generally thought to be influenced by positive and/or negative selection events occurring when TCRs on developing T cells interact with self-antigens and major histocompatibility complex components. Using a model system in which specific antigen-reactive cells can be monitored by virtue of their preferential expression of certain TCR beta-chain variable (V beta) domains, it has been shown that self-reactive T cells are clonally deleted during development. We report here that clonal deletion of V+ beta 6 cells in Mlsa mice can be prevented by in vivo neonatal administration of monoclonal antibodies directed against CD4. Furthermore, as anti-CD4 monoclonal antibody treatment resulted in the reappearance of V+ beta 6 cells in the mature CD8+ T-cell subset, it is likely that clonal deletion acts on the CD4+CD8+ thymocyte subset and that this subset is an intermediate stage in the differentiation pathway of both CD4+ and CD8+ T-cell lineages.

The mechanism of inhibition of experimental allergic encephalomyelitis in the rat by monoclonal antibody against CD4

Lewis rats with actively induced or passively transferred experimental allergic encephalomyelitis (EAE) were treated with a monoclonal antibody (MAb) which binds to the CD4 antigen of rat helper/inducer T cells. Actively immunized animals treated at the first onset of clinical signs experienced only a mild form of the disease and rapidly recovered while the majority of those treated prophylactically never showed clinical signs of EAE. Passively transferred EAE was also completely inhibited with anti-CD4 MAb. In treated animals which exhibited only mild clinical signs of EAE, spinal cord and cerebellar leukocyte infiltrates were quite similar to those in untreated rats but where anti-CD4 MAb treatment completely prevented clinical EAE, histological signs were minimal or absent. Like Lewis rats which have recovered naturally from EAE, those treated with anti-CD4 MAb were both resistant to a secondary challenge with myelin basic protein and harboured potential encephalitogenic cells which were capable of transferring disease to recipient rats. Disease in these recipients was, however, of much greater severity than that experienced by animals receiving cells from naturally recovered (untreated) donors. These data demonstrate that administration of anti-CD4 MAb to rats can prevent EAE by a mechanism which does not ablate the encephalitogenic CD4 + cells or prevent the development of resistance to EAE but which may inhibit the disease by preventing the function of already activated effector cells.