The treatment of moderate to severe psoriasis with a new anti-CD11a monoclonal antibody.

BACKGROUND: Anti-CD11a (hu1124) is a humanized monoclonal antibody directed against the CD11a subunit of LFA-1. This study investigated whether treatment with anti-CD11a antibody provides clinical benefit to patients with moderate to severe plaque psoriasis. METHODS: This was a double-blind, placebo-controlled, phase II, multicenter study. In total, 145 patients with minimum Psoriasis Area and Severity Index scores of 12 and affected body surface area of 10% or more were sequentially enrolled into low-dose (0.1 mg/kg, n = 22) or high-dose (0.3 mg/kg, n = 75) groups. Within groups, patients were randomized to treatment or placebo (n = 48) in a 2:1 ratio. Drug was administered intravenously at weekly intervals for 8 weeks. RESULTS: The percentage of subjects achieving more than 50% improvement in physician's global assessment at day 56 (1 week after final dose) was 15% and 48% for placebo and 0.3 mg/kg of drug, respectively (P =.002). A physician's global assessment of excellent (>75% improvement) was greater in the 0.3 mg/kg group versus placebo (25% vs 2%, P =.0003). Average Psoriasis Area and Severity Index scores at day 56 were 13.9 +/- 7.5 (placebo) and 10.9 +/- 8.4 (0.3 mg/kg) (P <.0001). Epidermal thickness was reduced in the 0.3 mg/kg group compared with the placebo group (37% vs 19%, P =.004). Treatment was well tolerated; mild to moderate flu-like complaints were the most common adverse events. White blood cell counts and lymphocyte counts transiently increased. Depletion of circulating lymphocytes did not occur. CONCLUSIONS: Anti-CD11a antibody administered intravenously in 8 weekly doses of 0.3 mg/kg was well tolerated and induced clinical and histologic improvements in psoriasis.

Flow cytometry crossmatching as a predictor of acute rejection in sensitized recipients of cadaveric renal transplants.

Flow cytometry crossmatching (FCXM) was developed as a more sensitive assay than the standard complement-dependent cytotoxicity crossmatch (CDCXM) for the detection of anti-donor antibodies, that mediate hyperacute rejection and graft loss in the early post-transplant period in renal transplant recipients. The role of FCXM in predicting long-term clinical outcome in renal allograft recipients is unclear. This study examines the role of FCXM in predicting long-term clinical outcome in highly sensitized recipients of cadaveric renal transplants. All patients (n = 100) with peak panel reactive antibody (PRA) levels > 30%, who received cadaveric renal transplants between 1/1/'90 and 12/31/'95 at our institution, were divided into FCXM + and FCXM - groups. The incidence of acute rejection was determined for each group during the first yr after transplant. Graft survival rates at 1, 2, and 3 yr, and creatinine levels were also compared between groups. FCXM + patients experienced a higher incidence of acute rejection during the first yr after transplant (69 vs. 45%), and a higher percentage of FCXM + patients had more than one episode of acute rejection during the first yr after transplant (34 vs. 8%) when compared to FCXM - patients. There was no statistically significant difference in 1-, 2-, or 3-yr graft survival between FCXM + and FCXM - patients (76 vs. 83, 62 vs. 80, 62 vs. 72%, respectively). These results suggest that sensitized FCXM + cadaveric renal transplant recipients have a higher incidence of acute rejection episodes in the first yr after transplant. Given the association of multiple rejection episodes with poor long-term allograft survival, FCXM may be a useful predictor of long-term clinical outcome in this sub-group of renal transplant recipients.

Population pharmacokinetics and pharmacodynamics of the anti-CD11a antibody hu1124 in human subjects with psoriasis.

The pharmacokinetics of hu1124, a human anti-CD11a antibody, were investigated in human subjects with psoriasis. CD11a is a subunit of LFA-1, a cell surface molecule involved in T cell mediated immune responses. Subjects received a single dose of 0.03, 0.1, 0.3, 0.6, 1, 2, 3, or 10 mg/kg of hu1124 intravenously over 1-3 hr. Blood samples were collected at selected times from 60 min to 72 days after administration. Plasma samples were assayed for hu1124 by ELISA, and pharmacokinetic analyses were performed on the drug plasma concentrations. As the dose of hu1124 was increased, the clearance decreased from 322 ml/day per kg at 0.1 mg/kg to 6.6 ml/day per kg at 10 mg/kg of hu1124. The plasma hu1124 concentration-time profile suggested that the clearance of hu1124 was saturable above 10 micrograms/ml. In addition, treatment with hu1124 caused a rapid reduction in the level of CD11a expression on CD3-positive lymphocytes (T cells) to about 25% of pretreatment levels. Regardless of the hu1124 dose administered, cell surface CD11a remained at this reduced level as long as hu1124 was detectable (> 0.025 microgram/ml) in the plasma. When hu1124 levels fell below 3 micrograms/ml, the drug was rapidly cleared from the circulation and expression of CD11a returned to normal within 7-10 days thereafter. In vitro, half-maximal binding of hu1124 to lymphocytes was achieved at about 0.1 microgram/ml and saturation required more than 10 micrograms/ml. One of the receptor-mediated pharmacokinetic/pharmacodynamic models which was developed describes the dynamic interaction of hu1124 binding to CD11a, resulting in the removal of hu1124 from the circulation and reduction of cell surface CD11a. The model accounts for the continually changing number of CD11a molecules available for removing hu1124 from the circulation based on prior exposure of cells expressing CD11a to hu1124. In addition, the model also accounts for saturation of CD11a molecules by hu1124 at drug concentrations of approximately 10 micrograms/ml, thereby reducing the clearance rate of hu1124 with increasing dose.

HLA compatibility requirements for CD8(+)-T-cell-mediated suppression of human immunodeficiency virus replication.

CD8(+) T cells from human immunodeficiency virus (HIV)-infected individuals can suppress HIV replication in cultured CD4(+) cells by a noncytotoxic mechanism. Efficient suppression of HIV replication (>90% reduction) does not require HLA class I or class II histocompatibility between the effector CD8(+) T cells and the infected target CD4(+) T cells. However, maximal control of HIV production occurs when the CD8(+) effector cells and CD4(+) target cells are syngeneic. In some cases, more than 20-fold fewer syngeneic CD8(+) T cells were required to achieve the same degree of HIV inhibition as HLA-mismatched CD8(+) T cells. The increased antiviral activity seen in the syngeneic setting did not map exclusively to either the HLA class I or class II locus. These findings suggest that genetic compatibility (potentially, but not necessarily, at the HLA class I and class II loci) regulates CD8(+) T-cell noncytotoxic antiviral activity against infected CD4(+) T cells.

Differential activation of T cells by natural antigen peptide analogues: influence on autoimmune and alloimmune in vivo T cell responses.

Recent studies using synthetic altered peptide ligands (Analogues) have led to the fine dissection of TCR-mediated T cell functions elicited by Ag recognition. Certain Analogues behave as full agonists of the antigenic peptide while others are partial agonists in that they only trigger selected T cell functions. Additionally, peptide Analogues can behave as antagonists by inhibiting functions of T cell clones when coincubated with the wild-type peptide. In fetal thymic organ cultures, synthetic altered peptide ligands can impact T cell repertoire selection. However, the influence of naturally occurring peptide Analogues on T cell immunity in vivo remains hypothetical. We previously reported that, in B10.A mice, immunogenicity and tolerogenicity of the self-MHC class I peptide, Ld 61-80, were influenced by the presentation of a cross-reactive self-peptide, Kk 61-80. Here, we show that Kk 61-80 self-peptide represents a partial agonist of Ld 61-80 in that it induced the proliferation but not the lymphokine production of Ld 61-80-primed T cells. Next, we showed that presentation of Kk 61-80 Analogue peptide mediated T cell tolerance toward Ld 61-80 self-peptide. Alternatively, when Ld protein represented an alloantigen displayed on transplanted cells, immunization with Kk 61-80 Analogue sensitized recipient mice to Ld 61-80 peptide, thus inducing potent immune responses to donor cells. These results show that the presentation of natural Analogue peptides may represent an essential component of T cell responses involved in autoimmunity and transplant rejection.

In utero hematopoietic stem cell transplants prolong survival of postnatal kidney transplantation in monkeys.

The authors hypothesized that in utero transplantation of T-cell-depleted paternal marrow into rhesus monkey fetuses would induce tolerance to postnatal kidney grafts from the marrow donor. T-cell-depleted paternal bone marrow was transplanted intraperitoneally into two female fetal rhesus monkeys at 61 +/- 1 days' gestation. Chimeric monkeys (n = 2) received kidney transplants from paternal donors. Control monkeys (n = 2) underwent kidney transplants without prior in utero stem cell transplants. Both chimeric monkeys demonstrated low level (<0.1% donor cells) engraftment in the bone marrow and peripheral blood using the polymerase chain reaction assay for the Y chromosome. The mixed lymphocyte reaction demonstrated hyporeactivity to the donor. Control animals demonstrated severe acute rejection and graft failure 1 week posttransplant. The first chimeric monkey had no significant clinical or sonographic evidence of renal failure until 7 weeks after the transplant. Biopsy findings showed mild rejection 1 week postoperatively, but rejection did not significantly progress until 5 weeks later. The second chimeric monkey had no significant clinical or sonographic changes for 4 weeks, but evidence of moderate rejection was seen on biopsy results. This monkey was given a 10-week course of immunosuppression, and had no clinical or sonographic renal deterioration, although biopsy results showed chronic rejection that was confirmed when electively euthanized 8 months later. Our data suggest that in utero transplantation of hematopoietic stem cells can increase the survival of a kidney allograft in the rhesus monkey.

Successful immunomodulation with intravenous gamma globulin and cyclophosphamide in an alloimmunized heart transplant recipient.

A highly human leukocyte antigen-sensitized heart transplant candidate underwent immunomodulation with intravenous gamma globulin and cyclophosphamide. His panel reactive antibody screen fell from 64% to 14%. He underwent successful orthotopic heart transplantation with a histoincompatible, T-cell cross-match-negative heart without the development of hyperacute rejection. The combination of intravenous gamma globulin and cyclophosphamide may be a simple and effective means to overcome the human leukocyte antigen-barrier in sensitized heart transplant candidates.

Sequence-independent inhibition of RNA transcription by DNA dumbbells and other decoys.

DNA dumbbells are stable, short segments of double-stranded DNA with closed nucleotide loops on each end, conferring resistance to exonucleases. Dumbbells may be designed to interact with transcription factors in a sequence-specific manner. The internal based paired sequence of DNA dumbbells in this study contains the X-box, a positive regulatory motif found in all MHC class II DRA promoters. In electrophoretic mobility shift assays (EMSAs), dumbbells and other oligonucleotides ('decoys') with the core X-box sequence were found to compete with the native strand for binding to X-box binding proteins (including RFX1). However, only the X-box dumbbell was capable of forming detectable complexes with such proteins using EMSA. In a model cell system, dumbbells were tested for their ability to block RFX1VP16 activation of a plasmid containing multiple repeats of the X-box linked to the CAT gene. While it appeared that dumbbells could block this activation, the effect was non-specific. This and further evidence suggests an inhibition of transcription, most likely via an interaction with the general transcriptional machinery.

An oligonucleotide blocks interferon-gamma signal transduction.

Interferon (IFN)-gamma is an important mediator of transplant graft rejection. It induces endothelial cell expression of HLA-DR and intercellular adhesion molecule-1, which render transplant grafts more susceptible to rejection by the host. Oligonucleotide 5'-GGG GTT GGT TGT GTT GGG TGT TGT GT-RNH2 (oligo I) blocks multiple IFN-gamma effects in human K562 cell cultures. A systematic approach revealed that oligo I has a novel, and potentially important, mode of action--it blocks the binding of IFN-gamma to its receptor, thus preventing activation of the IFN-gamma signal transduction pathway. The results are consistent with an aptamer mechanism of action, because oligo I exerts its inhibitory effects by interacting with protein, not intracellular nucleic acid targets, such as mRNA or genomic DNA.

The use of intravenous immunoglobulin in recurrent pregnancy loss associated with combined alloimmune and autoimmune abnormalities.

PROBLEM: Several studies have evaluated the effect of intravenous gammaglobulin (IVIG) in women with unexplained recurrent spontaneous abortions (RSA). Data regarding the underlying immunologic abnormalities in these patients is scant. This study reports the pregnancy outcome and immunologic changes observed in a large group of women with RSA associated with well-defined alloimmune and autoimmune abnormalities treated with IVIG. METHODS: Thirty-five patients with three or more recurrent miscarriages were studied. None of the patients had identifiable alloimmune response to paternal lymphocytes. Twenty-four patients had anti-thyroid antibodies, ten patients had high levels of circulating immune complexes, and six patients had anti-cardiolipin antibodies. Five patients had Hashimoto's disease, one had immune thrombocytopenic purpura, and one had Crohn's disease. Twenty-three patients had more than one autoimmune abnormality. All patients received IVIG infusions (200-250 mg/kg) every 3 weeks during the first 8 months of pregnancy. RESULTS: Twenty-eight patients (80%) had a successful pregnancy. Decrease of the level of autoantibodies and circulating immune complexes was observed in all patients who had a successful pregnancy. Only three of these patients developed measurable alloimmune response to paternal antigens. CONCLUSIONS: This preliminary study suggests that IVIG may be of benefit to patients with recurrent pregnancy associated with combined alloimmune and autoimmune abnormalities. This benefit was seen in spite of lack of detectable correction of the alloimmune abnormality in the majority of patients.

Identification of high- and low-risk second kidney grafts.

The purpose of this study was to identify recipients who are at low or high risk of early cadaveric regraft failure by segregating results of the flow cytometric crossmatch (FCXM) test with previous graft survival time (PGST). Early immunologic kidney regraft failure was analyzed in 103 multicenter recipients by cross-stratifying FCXM negative/positive status with < or =3- and >3-month PGST. T cell and B cell cytotoxicity crossmatches were negative. All were tested retrospectively in the T cell FCXM and 60 of the 103 were also tested in the B cell FCXM. A positive T and B cell FCXM was defined as a mean channel shift of > or = 9 (256 channel log scale) or > or = 40 (1024 channel log scale) for pretransplant crossmatch serum above negative control serum. Recipients received triple immunosuppression therapy and limited-use antilymphocyte induction therapy. Early cadaveric regraft losses were biopsied. Comparably good rates of second kidney graft survival at 3 years were found among three ow risk subsets: 78% for 18 FCXM-positive patients with PGST >3 months, 78% for 49 FCXM-negative patients with PGST >3 months, and 84% for 19 FCXM-negative patients with PGST < or =3 months. in contrast, 53% 3-month and 44% 3-year regraft survival rates occurred in 17 high-risk FCXM-positive recipients with a PGST < or =3 months. The odds ratio for increased relative risk of early second graft loss was 4.5 (confidence interval: 1.32-1.67) for the high-risk versus low-risk subsets (P = 0.009). Within the high-risk subset, 56% (5 of 9) of those who were FCXM T negative B positive experienced early regraft loss. A positive B cell FCXM has an adverse clinical impact only for high-risk regraft recipients. Pretransplant panel reactive antibody levels, pregnancy, number of blood transfusions between grafts, repeat donor HLA mismatches, and regraft recipient HLA mismatches did not correlate with early regraft loss. We conclude that kidney regraft survival rates in low-risk recipients (PGST >3 months/FCXM negative or positive [T and/or B cell] and PGST < or = 3 months/FCXM negative) approach primary graft survival rates and justify retransplantation, but the rate in high-risk regraft candidates (PGST < or =3 months/FCXM positive T and/or B cell) suggests that retransplantation should be performed only with a negative FCXM.

T cell tolerance is influenced by concomitant T cell recognition of cross-reactive self-peptides.

Although the current dogma of T cell recognition stresses its exquisite specificity, T cell clones selected for a given peptide can recognize other sequentially or structurally related peptides. Here, we have examined the immunogenicity and tolerogenicity of various self-peptides derived from region 61-80 of different MHC class I proteins co-expressed in the same mouse. Following immunization of B10.A mice (K(k), A(k), E(k), L(d), D(d)) with self-L(d) 61-80 peptide, vigorous MHC class II-restricted T cell proliferation was elicited after restimulation with either the immunogen or with self-K(k) 61-80 but not with self-D(d) 61-80. Furthermore, adult B10.A mice, tolerized with L(d) 61-80 prior to immunization with L(d) 61-80 did not respond to challenge with L(d) 61-80 and the cross-reactive K(k) 61-80. However, following K(k) 61-80 immunization, L(d) 61-80-tolerized mice responded to K(k) 61-80 but not to L(d) 61-80. Thus, tolerance induction to L(d) 61-80 resulted in the elimination/inactivation of L(d) 61-80-reactive T cells including the subpopulation that cross-reacted with K(k) 61-80. However, T cells that recognized K(k) 61-80 exclusively were preserved. Moreover, we showed that immunization with K(k) 61-80 resulted in tolerance breakdown to the cross-reactive, dominant self-peptide D(b) 61-80 in B10.A(4R) mice (K(k), A(k), L(d),D(b)). Together, these results show that the autoimmune T cell repertoire is influenced by the concomitant recognition of different cross-reactive self-peptides within the same individual.

Induction of T cell responses to a self-antigen following allotransplantation.

T cell tolerance to self-antigens is established through the recognition by immature T cells of dominant self-peptides presented in association with self-MHC molecules in the developing thymus (negative selection). The self-peptide Dd 61-80 is dominant in syngeneic BALB/c mice (H2d). T cell tolerance to Dd 61-80 in this mouse strain resulted in the absence of T cell proliferation following in vivo priming with Dd 61-80 peptide. Here, we show that transplantation of BALB/c mice with allogeneic B10.A (H2a) splenocytes led to an autoimmune T cell response toward the dominant self-peptide Dd 61-80. No T cell responses to Dd 61-80 peptide were observed after transplantation of C57BL/6 (H2b) splenocytes into BALB/c recipients. In addition, we provide evidence indicating that the breakdown of tolerance to Dd 61-80 self-peptide resulted from the presentation of the donor crossreactive peptide Kk 61-80 at the surface of recipient antigen-presenting cells. Taken together, our results suggest that following allotransplantation, T cell responses to donor antigens could spread to crossreactive determinants on self-proteins, thus perpetuating and amplifying the rejection process and presumably initiating tissue-specific autoimmune disorders.

Self determinant selection and acquisition of the autoimmune T cell repertoire.

Autologous proteins are continuously processed and presented in the form of peptides associated with self major histocompatibility (MHC) molecules at the surface of antigen-presenting cells for interaction with autoreactive T cells. During thymic selection, the presentation of self peptides is an essential element in the establishment of the T cell repertoire. Developing T cells which recognize self peptide/self MHC complexes with sufficient affinity are clonally deleted. However, we and others have recently demonstrated that a variety of self peptides, despite their high binding affinity to MHC molecules, never reach the threshold of presentation to ensure negative selection (cryptic self peptides). This mechanism may have been selected to avoid excessive purging of T cell repertoire during ontogeny. However, T cells directed to cryptic self determinants represent a continuous threat for the initiation of autoimmunity in adults. Supporting this view, recent studies have documented the involvement of cryptic self peptide presentation in different autoimmune diseases. In this article, we examine the factors that govern the selection of self peptides for presentation to autoreactive T cells in vivo and discuss their contribution to both the induction and the maintenance of self tolerance. In addition, we analyze the mechanisms by which the hierarchy of determinants on a self protein can be disrupted, thereby leading to the presentation of previously cryptic self peptides and the induction of an autoimmune T-cell-mediated process.

Direct evidence for in vivo induction of CD8+ cytotoxic T cells directed to donor MHC class I peptides following mouse allotransplantation.

There is accumulating evidence indicating that the T cell response to donor major histocompatibility complex (MHC) peptides plays a crucial role in graft rejection. We and others previously demonstrated the involvement of MHC class-II-restricted recognition of donor MHC class I and II peptides by alloreactive CD4+ T helper cells in graft rejection. Here we studied the in vivo induction of CD8+ cytotoxic T lymphocytes (CTL) directed to donor MHC class I peptides following allotransplantation in the mouse. To address this question, BALB/c irradiated splenocytes (H-2d) (Kd, A(d), E(d), Ld, Dd) were injected into Ld-deficient BALB/c-dm2 (dm2) mutant mice (Kd, A(d), E(d), -, Dd). Nine days after allogeneic cell transplant, recipient lymph node T cells were tested for cytolytic activity using peritoneal macrophages as targets. We observed that in addition to BALB/c targets, dm2 macrophages could also be lysed but only when incubated with a dominant peptide on donor Ld molecule, Ld 61-80. This response was abolished by anti-CD8 but not anti-CD4 monoclonal antibodies. In addition, after immunization of dm2 mice with the peptide Ld 61-80, alloreactive CTL were generated in vivo and shown to destroy allogeneic donor BALB/c target cells in the absence of exogenously added peptide. We conclude that after allotransplantation, concomitant in vivo priming of alloreactive CD8+ CTL by donor MHC class I peptides occurs through both direct and indirect pathways of allorecognition.

HLA class II antigen associations help to define two types of alopecia areata.

BACKGROUND: Multiple HLA class I and class II antigen associations have been described for alopecia areata (AA). As in other immune-mediated diseases, the HLA antigens associated with AA could influence the patient's ability to respond to immune challenge from both self- and non-self-antigens and may offer clues to the cause and prognosis of and potential therapy for the disease. OBJECTIVE: Our purpose was to determine which HLA class II antigens are associated with two forms of long-standing AA, which we define to be long-standing patchy AA and long-standing alopecia totalis (AT) and alopecia universalis (AU). We also examined other factors such as age at onset of disease and familial and patient histories of autoimmune disease for correlation with the two groupings. METHODS: Patients were typed for HLA class I and class II antigens by serologic methods and were typed by molecular methods for the subtypes of the HLA class II antigens. RESULTS: HLA-DR11 (DRB1*1104) and HLA-DQ7 (DQB1*0301) were found to be highly significantly increased in frequency in patients with long-standing AT/AU (group III) but not in patients with long-standing patchy AA (group II); both patient groups showed increased frequencies of HLA-DQ3 (DQB1*03). Group III patients were unique in their early age at onset of disease. Familial incidence of AA was 37% in patients who had their first patch by 30 years of age and 7.1% with the first patch after 30 years of age. CONCLUSION: The data support the differential association of two well-defined clinical forms of AA, namely long-standing AT/AU and long-standing patchy AA, with specific HLA antigens and age at onset; they also suggest that the broad antigen HLA-DQ3, DQB1*03, is a likely candidate for general susceptibility to AA. Our findings also suggest a bimodal pattern of disease with an early-onset form associated with greater severity, long duration, and family history of the disease and a late-onset form characterized by milder severity, shorter duration, and low family incidence.

Presentation of a self-peptide for in vivo tolerance induction of CD4+ T cells is governed by a processing factor that maps to the class II region of the major histocompatibility complex locus.

Self-proteins are regularly processed for presentation to autoreactive T cells in association with both class I and class II major histocompatibility complex (MHC) molecules. The presentation of self-peptides plays a crucial role in the acquisition of T cell repertoire during thymic selection. We previously reported that the self-MHC class I peptide Ld 61-80 was immunogenic in syngeneic B10.A mice (H-2a). We showed that despite its high affinity for self-MHC class II molecules, Ld 61-80 peptide failed to induce elimination of autoreactive CD4+ T cells, presumably due to incomplete processing and presentation in the B10.A's developing thymus (cryptic-self peptide). In this report, we showed that the cryptic phenotype was not an intrinsic property of the self-peptide Ld 61-80 since it was found to be naturally presented and subsequently tolerogenic in BALB/c mice (H-2d) (dominant self-peptide). In addition, the self-peptide Ld 61-80 was found to be immunogenic in different H-2a mice while it was invariably tolerogenic in H-2d mice regardless of their background genes. We observed that Ld 61-80 bound equally well to H-2d and H-2k MHC class II molecules. Also, no correlation was found between the quantity of self-Ld protein and the tolerogenicity of Ld 61-80. Surprisingly, Ld 61-80 was not naturally presented in (H-2d x H-2a) F1 mice, indicating that the H-2a MHC locus contained a gene that impaired the presentation of the self-peptide. Analyses of T cell responses to the self-peptide in several H-2 recombinant mice revealed that the presentation of Ld 61-80 was controlled by genes that mapped to a 170-kb portion of the MHC class II region. This study shows that (a) endogenously processed self-peptides presented by MHC class II molecules are involved in shaping the CD4+ T cell repertoire in the thymus; (b) The selection of self-peptides for presentation by MHC class II molecules to nascent autoreactive T cells is influenced by nonstructural MHC genes that map to a 170-kb portion of the MHC class II region; and (c) the MHC locus of H-2a mice encodes factors that prevent or abrogate the presentation by MHC class II molecules of the self-peptide Ld 61-80. These findings may have important implications for understanding the molecular mechanisms involved in T cell repertoire acquisition and self-tolerance induction.

Immunopharmacodynamic studies of cyclosporine in patients awaiting renal transplantation.

The immunopharmacodynamics of cyclosporine were investigated in eight hemodialysis patients awaiting renal transplantation. Cyclosporine was administered orally (10 mg/kg) and intravenously (4 mg/kg), with both administrations separated by at least one week. Plasma samples were processed at 37 degrees C and analyzed for specific cyclosporine and its four major metabolites (AM1, AM1c, AM9, and AM4N) using high-performance liquid chromatography. In addition, the in vitro immunosuppressive activity of these serial plasma samples was estimated as a relative percentage inhibition of third party mitogenic lymphocyte proliferation stimulated with phytohemagglutinin. The relationships between concentration and effect of cyclosporine versus time were noted. These results suggest that unchanged cyclosporine concentrations in plasma correlate with mitogen-induced lymphocyte suppression yielding significant immunosuppressant activity of cyclosporine. Control studies with plasma from healthy volunteers spiked with cyclosporine in the concentration range of 0-10,000 ng/mL were developed. A sigmoidal Emax model was fitted to the effect versus plasma concentration data. The ratio of effect versus predicted effect were calculated for intravenous cyclosporine dosing. There was a good correlation between the observed and predicted inhibitory effect.

Suppression of interferon-gamma induction of MHC class II and ICAM-1 by a 26-base oligonucleotide composed of deoxyguanosine and deoxythymidine.

Interferon-gamma (IFN-gamma) is an important cytokine released by T lymphocytes and natural killer cells which is able to induce expression of class II MHC and ICAM-1, crucial factors in cellular immune response. HeLa S3, HS 27, and NF-71-1 are cell lines which can be induced to express HLA-DR and HLA-DP by exposure to IFN-gamma. When T2 (5'GGGGTTGGTTGTGTTGGGTGTTGTGTRNH(2)3') oligonucleotide was added at 5-20 microM every other day, cell surface induction of HLA-DR and HLA-DP by IFN-gamma was suppressed in a dose-dependent manner in HeLa S3. T2 suppressive effect on HLA class II was also observed in four different nontransformed human cell lines, HS 27 at passage 18, NF-71-1 at passage 5, human corneal endothelial cell at passage 5, and human retinal pigmented epithelial cell at passage 3. Control oligonucleotides had no suppressive effect. Northern hybridization showed that HLA-DR A mRNA induction by IFN-gamma was blocked by T2 in HeLa S3 and fibroblast 143B. The suppressive effect of T2 was also reversible as continued culture of the treated cells without further addition of the oligonucleotide allowed full re-expression of HLA-DR. Further experiments showed that T2 oligonucleotide was also able to inhibit IFN-gamma enhancement of ICAM-1 (CD54) on human corneal endothelial cell and human retinal pigmented epithelial cell. We conclude that T2 oligonucleotide is effective at suppressing HLA-DR, HLA-DP and ICAM-1 induction by IFN-gamma in transformed and nontransformed cells in vitro.