Biomedical subjects

A M Krensky

Publications and source records attributed to A M Krensky.

At least 19 recordsLinked to original sources

PubMed1996

Identification of a novel regulatory region critical for expression of the RANTES chemokine in activated T lymphocytes.

The RANTES chemokine is a T cell-expressed, proinflammatory cytokine recently implicated as a suppressive agent of HIV replication. We have identified tandem kappaB-like sequences within the promoter for RANTES that are critical for RANTES promoter-reporter gene activity in both the T cell tumor line Hut78 and in PHA-activated PBL. This region binds not only Rel family members (including p50-p65 heterodimers and p50-p50 homodimers) but also non-Rel factors up-regulated in PBL 3 to 5 days following activation. The expression of these "late" expressed nuclear factors correlates with an up-regulation of RANTES message found at this point in T cell activation. These factors are also constitutively expressed in functionally mature CD8+ T cells. We hypothesize that these apparently novel proteins are responsible in part for the temporal regulation of RANTES seen in peripheral blood T cells and represent a component of transcriptional regulatory machinery newly expressed at this "late" stage of peripheral T cell development.

Base Sequence↗

PubMed1996

Tumor-specific, cytotoxic T-lymphocyte response after idiotype vaccination for B-cell, non-Hodgkin's lymphoma.

Patients with non-Hodgkin's B-cell lymphoma who received an antitumor vaccine of idiotypic ig protein showed humoral and proliferative immune responses. Because immunity to some antigens, including tumor antigens and human pathogenic viruses, may be better correlated with the cytolytic cellular immune response, we evaluated 16 non-Hodgkin's lymphoma patients immunized with autologous idiotypic ig molecules for changes in tumor-specific cytotoxic T-lymphocyte precursor (CTLp) frequency using limiting dilution analysis. Eleven patients had a significant increase in tumor-specific CTLp. Eight of these 11 patients remain without evidence of disease or with stable minimal disease. In contrast, all five patients who did not have a significant change in tumor-specific CTLp have developed progressive disease. Patient vaccination with tumor associated protein antigens can increase tumor-specific CTLp frequencies. The correlation of increased tumor specific CTLp with freedom from progression is significant at P = .002. This study indicates that measurement of CTLp frequencies are relevant to the clinical evaluation of human tumor vaccines and suggests that cell-mediated cytolytic immune responses may be an important determinant of vaccine efficacy.

Adult↗

PubMed1996

HLA-derived peptides which inhibit T cell function bind to members of the heat-shock protein 70 family.

Synthetic peptides corresponding to sequences of HLA class I molecules have inhibitory effects on T cell function. The peptides investigated in this study have sequences corresponding to the relatively conserved region of the alpha 1 helix of HLA class I molecules that overlaps the "public epitope" Bw4/Bw6. These HLA-derived peptides exhibit inhibitory effects on T lymphocytes and have beneficial effects on the survival of allogenic organ transplants in mice and rats. Peptides corresponding to the Bw4a epitope appear most potent as they inhibit the differentiation of T cell precursors into mature cytotoxic T lymphocytes (CTL) and target cell lysis by established CTL lines and clones. To elucidate the mechanism through which these peptides mediate their inhibitory effect on T lymphocytes, peptide binding proteins were isolated from T cell lysates. We show that the inhibitory Bw4a peptide binds two members of the heat-shock protein (HSP) 70 family, constitutively expressed HSC70 and heat-inducible HSP70. Peptide binding to HSC/HSP70 is sequence specific and follows the rules defined by the HSC70 binding motif. Most intriguing, however, is the strict correlation of peptide binding to HSC/HSP70 and the functional effects such that only inhibitory peptides bind to HSC70 and HSP70 whereas noninhibitory peptides do not bind. This correlation suggests that small molecular weight HLA-derived peptides may modulate T cell responses by directly interacting with HSPs. In contrast to numerous reports of HSP70 expression at the surface of antigen-presenting cells and some tumor cells, we find no evidence that HSC/HSP70 are expressed at the surface of the affected T cells. Therefore, we believe that the peptides' immunodulatory effects are not mediated through a signaling event initiated by interaction of peptide with surface HSP, but favor a model similar to the action of other immunomodulatory compounds, FK506 and cyclosporin A, with a role for HSC/HSP70 similar to that for immunophilins, FKBPs and CyP40.

Amino Acid Sequence↗

PubMed1996

Stimulus specificity of matrix metalloproteinase dependence of human T cell migration through a model basement membrane.

Chemotaxis of human T lymphoblastoma cells of the Tsup-1 line, which migrate similarly to blood T cells, through a layer of basement membrane-like Matrigel on a polycarbonate micropore filter was evoked by vasoactive intestinal peptide (VIP; concentration for a maximal response, 10(-7)M), IL-2 (10(-9)M), and the chemokines RANTES (10(-10)M) and macrophage inflammatory protein-1 alpha (10(-10)M). Chemotactic concentrations of each factor increased Tsup-1 cell secretion of matrix metalloproteinase-9 (MMP-9), with significant responses by 4 h for VIP, IL-2, and IL-4, but only after 24 h for macrophage inflammatory protein-1 alpha and RANTES, as quantified by Western blots and zymography. 3H-Labeled type IV human collagen incorporated in the Matrigel layer was degraded by migrating Tsup-1 cells, as assessed by release of radioactive fragments of the collagen. The in situ degradation of type IV collagen in Matrigel by migrating Tsup-1 cells was enhanced most significantly by VIP, IL-2, and IL-4 after 4 h at concentrations that increased the secretion of MMP-9 optimally, but only after 24 h by macrophage inflammatory protein-1 alpha and RANTES. The specific MMP inhibitor GM6001 suppressed Tsup-1 cell MMP activity evoked by all stimuli, as determined by zymography and in situ degradation of 3H-Labeled type IV human collagen. The chemotactic migration of Tsup-1 cells through Matrigel, but not through a filter alone, in response to optimal concentrations of VIP, IL-2, and IL-4, but not the chemokines, was inhibited by GM6001, with a concentration dependence similar to that for suppression of MMP activity. Thus elicitation of T cell chemotactic migration through a model basement membrane by stimuli that increase MMP activity early in the response depends on degradation of matrix proteins by MMP, whereas stimuli that recruit MMP late may rely on early activation of other proteases.

Basement Membrane↗

PubMed1996

RANTES chemokine expression in diseased and normal human tissues.

RANTES is a member of a large family of cytokines, called chemokines, which are thought to play a regulatory role in inflammatory processes. We have made recombinant human RANTES protein which was used to generate a panel of anti-RANTES monoclonal antibodies. Following characterization, select anti-RANTES monoclonal antibodies were used for immunohistologic staining of a large panel of normal, diseased and fetal tissue sections. Diseased tissues included eleven lymphomas and eight renal tumors. Most tissues were also tested in parallel for RANTES mRNA by in situ hybridization using RANTES mRNA specific oligomeric probes. As expected, most normal adult tissues contain few, if any, RANTES positive cells. In contrast, RANTES expression dramatically increases in inflammatory sites. In addition, megakaryocytes, some tumours, and select fetal tissues express high levels of RANTES message and protein. These results indicate a wider expression of RANTES than previously appreciated and suggest multiple physiologic roles for this soluble factor.

Animals↗

PubMed1996

Cytotoxic T lymphocyte recognition of non-Hodgkin's B cell lymphomas.

Human cytolytic T lymphocytes specific for autologous Burkitt's lymphoma express the gamma, delta T cell receptor and recognize immunoglobulin idiotype in an MHC-unrestricted manner. Antibodies against a member of the heat shock protein 70 family inhibit this specific cytotoxicity, implicating these molecules in tumor recognition and antigen presentation. Such data is relevant to the design of novel immunotherapies for cancer and provides new insights into target recognition by gamma, delta T cells.

Humans↗

PubMed1996

Kinetics of transcription factors regulating the RANTES chemokine gene reveal a developmental switch in nuclear events during T-lymphocyte maturation.

RANTES is a chemoattractant cytokine (chemokine) whose gene is expressed immediately after stimulation of several cell types but upregulated late (3 to 5 days) after activation in normal T lymphocytes. Here we describe two cis-acting elements in the human RANTES promoter that act in T lymphocytes. One site interacts with NFIL6, which is activated within the first 24 h after T-cell activation. The second site binds an apparently novel complex that is upregulated later, between days 3 and 5. These data provide an explanation for the immediate-early expression of RANTES in some cell types and identify apparently novel factors contributing to late RANTES transcription in T cells. The results reveal a developmental switch occurring during normal T-cell maturation coincident with the onset of terminal differentiation and the binding of late-acting factors to sequences of the RANTES promoter.

Base Sequence↗

PubMed1996

Structure of HLA molecules and immunosuppressive effects of HLA derived peptides.

Elucidation of the structure of MHC molecules has provided profound new insights into their function in antigen presentation. In addition, structural studies have implicated certain regions of MHC molecules in specific functions. Although much of MHC biology has concentrated on the extensive polymorphism among these molecules, there is also evolutionary pressure to maintain the relatively monomorphic portions of these molecules. Drs. Krensky and Clayberger have found that synthetic peptides corresponding to linear sequences of HLA molecules have immunomodulatory effects both in vitro and in vivo. In this paper, they review the structure of HLA molecules and their studies of HLA derived peptides as novel immunotherapeutics. Members of the heat shock protein 70 family are implicated in the HLA derived peptide immunosuppressive pathway.

Alleles↗

PubMed1996

Immunologic tolerance: tailored antigen.

In summary, synthetic peptides corresponding to linear sequences of HLA class I molecules can inhibit T-cell responses in vitro and in vivo. These peptides induce immunologic tolerance by binding to hsp-70 family members, causing an increase in intracellular calcium, and down-regulating the nuclear factor of activated T cells, NF-AT. We suggest that heat shock proteins may function as novel immunophilins (Fig 2). Like cyclophilins and FK 506 binding proteins, heat shock proteins are ubiquitous, are involved in protein folding and trafficking, and bind exogenous drugs. Cyclosporine and FK 506 exert immunosuppressive effects by binding immunophilins, which as a result interrupt the phosphatase activity of calcineurin. Although the precise pathways involved in the synthetic HLA peptide effects are not as well worked out, it seems likely that peptide binding to heat shock protein is disrupting normal events in T-cell activation, giving rise to an apparently permanent state of anergy.

Amino Acid Sequence↗

PubMed1996

Novel immunotherapeutic strategies using MHC derived peptides.

Organ transplantation is now the treatment of choice for end stage organ failure. The ultimate goal in transplantation remains the development of strategies to induce specific tolerance to the allograft. The major histocompatibility complex (MHC) antigens are the principal targets of the immune response to allografts and T cell recognition of allo-MHC is the initial event which initiates allograft rejection. The availability of sequences of MHC genes in mice, rats and humans has made it possible to prepare synthetic peptides for the study of the role of MHC peptides in allorecognition and tolerance induction. New evidence confirms that there are at least two distinct, but not necessarily mutually exclusive, pathways of allorecognition. In the so-called "direct" pathway T cells recognize intact allo-MHC molecules on the surface of donor cells. These MHC molecules contain an array of endogenous peptides bound in their antigen presentation groove. In the "indirect" pathway, T cells recognize specific processed alloantigen presented as peptides in the context of self MHC by antigen-presenting cells (APCs). In addition, there is ample evidence that synthetic MHC peptides can immunomodulate the alloimmune response both in vitro and vivo, and that potent allo-tolerance can be induced with synthetic MHC peptides. Two types of effects mediated by synthetic MHC peptides have been demonstrated: (1) suppression of the alloimmune response by relatively non-polymorphic peptides and (2) antigen-specific unresponsiveness induced by polymorphic peptides. The mechanisms mediating the immunomodulatory effects of synthetic effects of synthetic class I and class II MHC peptides and the potential for clinical applications are reviewed.

Animals↗

PubMed1995

Gamma delta T cell recognition of tumor Ig peptide.

Although gamma delta T cells have been postulated to act as a surveillance mechanism that eliminates transformed or otherwise damaged cells, little is known about tumor recognition by gamma delta T cells, including the Ags that are recognized and the molecules that present them. Previously, we described human gamma delta CTL that recognize the autologous B cell lymphoma. Here we report that these gamma delta CTL lyse heterologous cells transfected with the tumor Ig lambda chain gene. Furthermore, the lambda chain is recognized as processed peptide in an Id-specific manner. T cell recognition does not involve classical MHC molecules, but it could be blocked by Abs directed against the heat shock protein grp75. These findings show a specific gamma delta T cell response to a highly polymorphic Ag such as tumor Id and implicate heat shock protein as a molecule required for recognition.

Amino Acid Sequence↗

PubMed1995

Novel tumor-associated accessory molecules involved in the gamma/delta cytotoxic T-lymphocyte-Burkitt's lymphoma interaction.

BACKGROUND: Tumor specific cytotoxic T lymphocytes (CTL) recognize antigen via the T-cell receptor (TCR). In addition, recognition requires accessory molecules involved in adhesion and signal transduction. The authors previously have characterized an autologous, Burkitt's lymphoma specific CTL line that uses the gamma-delta TCR to recognize antigen in a nonclassical context. The current study was undertaken to identify novel accessory molecules involved in this unusual TCR-tumor cell interaction. METHODS: A panel of monoclonal antibodies was generated against a Burkitt's lymphoma cell line and was screened for inhibition of autologous, tumor specific, cytolysis by a gamma-delta CTL line. Proteins identified by these monoclonal antibodies were further characterized by fluorescent-activated cell sorter analysis, Western blot and immunoprecipitation. RESULTS: Three known (CD5, CD43, and CD11a/CD18) and three novel (BAM-1, BAM-2, and BAM-3) cell surface molecules involved in the gamma-delta CTL-Burkitt's lymphoma interaction were identified and characterized. CONCLUSIONS: This study identifies and provides a preliminary characterization of three novel Burkitt's lymphoma-associated molecules involved in the gamma-delta CTL-tumor cell interaction and demonstrates that CD5, CD43, and CD11a/CD18 are involved in this interaction. It is likely that other unidentified accessory molecules are also involved in this and other effector cell-tumor interactions. Identification of such molecules may be useful in the design of new immunotherapeutic approaches.

Animals↗

PubMed1995

HLA-derived peptides as novel immunotherapeutics.

Soluble forms of HLA may be natural immunoregulatory molecules and may explain the "transfusion effect" seen in transplant patients in the precyclosporine era. Synthetic peptides corresponding to short linear sequences of HLA molecules have inhibitory effects on human T lymphocytes in vitro and cause tolerance induction in a rat heterotopic heart transplant model. Recent studies indicate that a subset of these peptides causes an increase in intracellular calcium which appears to account for T cell unresponsiveness and tolerance induction. The peptides bind to members of the heat-shock protein 70 family in a specific manner, depending upon peptide sequence. These studies provide a mechanism to explain the biologic activities of these new immunotherapeutic reagents.

Animals↗

PubMed1995

Immunosuppressive peptides corresponding to MHC class I sequences.

The induction of tolerance is a long-standing goal in transplantation. Intact MHC molecules, or fragments of them, are being used to render T cells unresponsive both in vitro and in vivo. Elucidation of the mechanisms underlying the effects of these treatments should aid the design of novel therapies for transplantation.

Crystallization↗

PubMed1995

HLA-derived peptides as novel immunosuppressives.

Peptides corresponding to linear sequences of HLA molecules have been synthesized and tested for immunomodulatory activity in in vitro assays using human T lymphocytes. Sequences from different parts of the HLA molecules have different effects. Peptides corresponding to residues 75-84 of an HLA class I supratypic specificity of limited heterogeneity (HLA-Bw4) had profound inhibitory effects in a variety of in vitro assays of human T lymphocyte function. Furthermore, a 2-wk course of human HLA sequences and cyclosporine therapy induced enduring immunologic tolerance in a rat model of heterotopic heart transplantation. These studies prompted clinical trials which are currently in progress. The peptides appear to induce T cell anergy by causing a prolonged intracellular calcium flux and interrupting normal signal transduction pathways. Furthermore, these peptides bind to members of the heat-shock protein 70 family, implicating these ubiquitous proteins in the immunomodulatory pathway. Such peptides may be normal physiologic mediators. In any case, they represent potential new immunotherapeutics for a variety of immune-mediated diseases.

Amino Acid Sequence↗

PubMed1995

Human leukocyte antigen-derived peptides as novel immunosuppressives.

Synthetic peptides corresponding to linear sequences of HLA class I and class II molecules can potently inhibit T lymphocytes responses both in vitro and in vivo. The class I and class II peptides studied to date seem to function by different mechanisms. Nevertheless, several different peptides have been shown to potently induce T cell anergy. Opelz and Terasaki first demonstrated that blood transfusions improved graft survival in transplant patients (19). Data from several sources indicate that blood transfusions are immunosuppressive in: 1) increasing infections in trauma patients who have received transfusions (20), 2) increasing metastases and/or relapses in cancer patients who have been transfused (21), and 3) remissions of autoimmune diseases associated with pregnancy and/or transfusion (22). It is likely that the active constituent of blood transfusions is soluble HLA molecule (23,24). Liver transplants, which are profoundly immunosuppressive in themselves, produce large amounts of soluble HLA (25). Both B and T lymphocytes in culture secrete soluble HLA (26). We hypothesize that soluble HLA is a natural immunoregulatory molecule involved in dampening of the immune response, but, even if this is not the case, synthetic peptides corresponding to HLA sequences have profound effects on T lymphocytes and may prove to be effective for the induction of clinical tolerance in transplant patients.

Animals↗

PubMed1994

Peptides corresponding to the CD8 and CD4 binding domains of HLA molecules block T lymphocyte immune responses in vitro.

CD4 and CD8 are cell surface glycoproteins that serve as co-receptors for Ag with the TCR. Recent studies have shown that both CD4 and CD8 interact with conserved regions of MHC class II and class I, respectively. To investigate further the roles of CD4 and CD8 in the immune response, we prepared synthetic peptides corresponding to the HLA sequences with which CD4 and CD8 are thought to interact. The peptide corresponding to residues 222 to 235 of the HLA class I heavy chain blocked the differentiation of human CTL precursors into active effect cells but affected neither the ability of PBLs to proliferate in response to mitogen nor the cytotoxic activity of established CTLs. In contrast, the peptide corresponding to residues 134 to 152 of the HLA-DR beta-chain inhibited the differentiation of CTL precursors, the proliferative response of freshly isolated PBL, and the proliferation of an established alloreactive CD4+ T cell clone to Ag. The inhibitory effect of the DR.134-152 peptide on CTL differentiation could be overcome by addition of exogenous IL-2 to the limiting dilution cultures, whereas the effect of the HLA-1.222-235 peptide was unaffected by exogenous IL-2. These results directly demonstrate a functional role for these regions of MHC molecules and underscore the central role of both CD4 and CD8 in the effective initiation of a CTL response.

Base Sequence↗

PubMed1994

Induction of allograft tolerance in rats by an HLA class-I-derived peptide and cyclosporine A.

T cell recognition of MHC molecules initiates a cascade of events resulting in allograft rejection. CTLs damage the graft by targeting nonself-MHC class I molecules. We and others have previously shown that small synthetic peptides corresponding to regions of certain MHC class I molecules can inhibit the CTL response against MHC class I alloantigens in vitro. Here we report that rat heart allografts survived survived indefinitely when transplanted into recipients treated with a synthetic peptide corresponding to residues 75-84 of (B7.75-84) in combination with a subtherapeutic dose of cyclosporine A. Furthermore, this treatment induced long-term donor-specific tolerance that was mediated by anergic cells, indicating that such peptides may have potential as therapeutics for human organ transplantation.

Amino Acid Sequence↗