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D M Pardoll

Publications and source records attributed to D M Pardoll.

At least 55 records · Page 3Linked to original sources
PubMed1996

The immunodominant major histocompatibility complex class I-restricted antigen of a murine colon tumor derives from an endogenous retroviral gene product.

Tumors express peptide antigens capable of being recognized by tumor-specific cytotoxic T lymphocytes (CTL). Immunization of mice with a carcinogen-induced colorectal tumor, CT26, engineered to secrete granulocyte/macrophage colony-stimulating factor, routinely generated both short-term and long-term CTL lines that not only lysed the parental tumor in vitro, but also cured mice of established tumor following adoptive transfer in vivo. When either short-term or long-term CTL lines were used to screen peptides isolated from CT26, one reverse-phase high performance liquid chromatography peptide fraction consistently sensitized a surrogate target for specific lysis. The bioactivity remained localized within one fraction following multiple purification procedures, indicating that virtually all of the CT26-specific CTL recognized a single peptide. This result contrasts with other tumor systems, where multiple bioactive peptide fractions have been detected. The bioactive peptide was identified as a nonmutated nonamer derived from the envelope protein (gp70) of an endogenous ecotropic murine leukemia provirus. Adoptive transfer with CTL lines specific for this antigen demonstrated that this epitope represents a potent tumor rejection antigen. The selective expression of this antigen in multiple non-viral-induced tumors provides evidence for a unique class of shared immunodominant tumor associated antigens as targets for antitumor immunity.

Animals↗
PubMed1996

Can bone marrow-derived thymic stromal cells mediate the positive selection of class I-restricted T cells?

Positive selection of T lymphocytes expressing self. MHC-restricted T cell receptors is mediated by MHC molecules expressed on thymic stroma. Among the more controversial aspects of this process is the relative role of MHC molecules on epithelial versus bone marrow-derived stromal elements (BM-APC). For CD4+ T cells, the weight of evidence suggests that positive selection is driven solely by MHC class II molecules expressed on thymic epithelial cells. In contrast, recent experiments have been interpreted to show that CD8+ T cell development can be driven by MHC class I molecules expressed on BM-APCs as well as epithelial cells. To directly address this issue, we have examined the development of T cells expressing a transgenic, MHC class I-restricted TCR in mice deficient in the rearrangement of endogenous TCR genes. Since the transgenic TCR is the only TCR expressed, this system is extremely sensitive and specific for detecting even inefficient events in T cell development. Our experiments demonstrate that MHC class I expression exclusively on BM-APC is incapable of driving positive selection of CD8+ T cells. This finding, together with earlier experiments for MHC class II, suggests a qualitatively unique function of thymic epithelial cells in mediating positive selection.

Animals↗
PubMed1996

Does B7-1 expression confer antigen-presenting cell capacity to tumors in vivo?

Tumors engineered to express the costimulatory molecule B7-1 can elicit CD8+ cytotoxic T lymphocyte (CTL)-dependent antitumor responses in immunocompetent mice. It has been postulated that this result reflects direct priming of CTL by the modified tumor in vivo. Previous studies of the immune response to a B7-1- murine colon carcinoma expressing influenza nucleoprotein (NP) as a model tumor antigen have demonstrated the crucial role of bone marrow-derived antigen-presenting cells (APCs) in the priming of NP-specific CTL in vivo. In this system, no evidence of direct CTL priming by tumor was detected. We have performed a similar analysis to determine if B7-1 transfectant of this tumor results in the direct priming of CTL, and to compare this response to that primed by host APCs. When H-2b-->H-2bxd bone marrow chimeras were immunized with a single injection of CT26/NP/B7-1 (H-2d), NP-specific CTL were detected that were restricted to the bone marrow haplotype (H-2b), but not to the tumor haplotype. In contrast, CTL recognizing the NP antigenic epitope in the context of the tumor's major histocompatibility complex were detectable only after multiple immunizations. These results suggest that whereas B7-1+ tumor vaccines result in some degree of direct presentation to CD8+ T cells, the dominant mechanism of CTL priming is through the uptake and presentation of tumor antigens by bone marrow-deprived APCs. However, repeated immunization with B7-1+ tumor cells can efficiently expand the directly primed CD8+ CTL population.

Amino Acid Sequence↗
PubMed1996

Treatment of established tumors with a novel vaccine that enhances major histocompatibility class II presentation of tumor antigen.

Presentation of antigenic peptides by MHC class II molecules to CD4+ T cells is critical to the generation of antitumor immunity. In an attempt to enhance MHC class II antigen processing, we linked the sorting signals of the lysosome-associated membrane protein (LAMP-1) to the cytoplasmic/nuclear human papilloma virus (HPV-16) E7 antigen, creating a chimera (Sig/E7/LAMP-1). Previously, we found that expression of this chimera in vitro and in vivo with a recombinant vaccinia vector targeted E7 to endosomal and lysosomal compartments and enhanced MHC class II presentation to CD4+ T cells compared to vaccinia expressing wild-type E7. In the current study, we tested these recombinant vaccinia for in vivo protection against an E7+ tumor, TC-1, which was derived from primary epithelial cells of C57BL/6 mice cotransformed with HPV-16 E6 and E7 and c-Ha-ras oncogenes. All mice vaccinated with 1 x 10(7) plaque-forming units of wild-type E7-vaccinia showed progressive tumor growth when challenged with a tumorigenic dose of TC-1 tumor cells; in contrast, 80% of mice vaccinated with the chimeric Sig/E7/LAMP1 vaccinia remained tumor free 3 months after tumor injection. Furthermore, treatment with the Sig/E7/LAMP-1 vaccinia vaccine cured mice with small established TC-1 tumors, whereas the wild-type E7-vaccinia showed no effect on this established tumor burden. These findings point out the therapeutic limitations of recombinant vaccinia expressing unmodified tumor antigens. Further, they demonstrate that modifications that reroute a cytosolic tumor antigen to the endosomal/lysosomal compartment can profoundly improve the in vivo therapeutic potency of recombinant vaccines.

Animals↗
PubMed1996

Murine tumor antigens: is it worth the search?

Standardized techniques that allow the direct identification of tumor antigens are now available. Several murine antigens recognized by T cells have already been identified. So far, the majority of these antigens derive from cellular proteins similar to those that give rise to human tumor antigens. While many of the known human tumor antigens are widely shared, most of the murine tumor antigens appear to be unique to the individual tumor from which they were isolated. Nonetheless, common features between murine and human tumor antigens are emerging, suggesting that these murine antigens will provide essential tools in the evaluation of antigen-based vaccines for the future treatment of cancer.

Animals↗
PubMed1996

In vivo cross-priming of MHC class I-restricted antigens requires the TAP transporter.

Recent in vitro evidence suggests two alternative mechanisms by which bone marrow-derived APCs may process exogenous antigens for presentation to CTL in vivo, a phenomenon termed cross-priming. Although in vitro studies have suggested that both TAP-dependent and TAP-independent pathways exist, we have now demonstrated an absolute requirement for a functional TAP for cross-priming to occur in vivo. Bone marrow chimeras reconstituted with marrow from TAP-defective donors develop functional CD8+ CTL, but have APCs with disrupted TAP function. In such chimeras, in vivo priming of naive CTL was observed when antigen was targeted to the ER in a TAP-independent fashion, but cross-priming could not be demonstrated. These results support the TAP-dependent mechanism of cross-priming.

ATP-Binding Cassette Transporters↗
PubMed1996

Enhanced immune priming with spatial distribution of paracrine cytokine vaccines.

In preclinical models, tumor cells genetically modified to express cytokines or other costimulatory molecules can generate systemic antitumor immunity. In some cases, these tumor vaccines have been shown to eradicate micrometastases. These results have led to the initiation of numerous phase I clinical trials employing either autologous or allogeneic tumor vaccines genetically modified to express cytokines and other genes. In this report, we use our murine model to identify a number of parameters that may be critical for enhancing vaccine efficacy. In addition to antigen dose and cytokine level, the distribution of vaccine inoculation was found to have a significant impact on vaccine potency. These results require consideration in early clinical trials designed to evaluate cellular vaccine therapy.

Animals↗
PubMed1996

Systemic and local paracrine cytokine therapies using transduced tumor cells are synergistic in treating intracranial tumors.

Development of an effective immunotherapeutic approach for treatment of CNS tumors must take into account the unique anatomic and immunologic features of the brain. We explored the antitumor immune response in the brain elicited by nonreplicating melanoma cells genetically engineered to produce either granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-2 (IL-2) in a paracrine fashion. Using a new model of intracranial melanoma in C57BL/6 mice, the cytokine-producing cells were given either as a subcutaneous vaccine to induce systemic antitumor immunity or as a direct injection into the brain as local immunotherapy. We found that GM-CSF-transduced cells, as a subcutaneous vaccine but not as an intracranial injection, afforded some protection from intracranial challenge with the wild-type tumor. In contrast, direct intracranial injection of tumor cells secreting IL-2 was protective whereas flank vaccination with IL-2 transductants was not. Combination therapy with both the subcutaneous GM-CSF-transductants as a vaccine and local administration of IL-2-transductants in the brain achieved a synergistic response. These findings provide a basis for the application of paracrine cytokine delivery to brain cancer therapy both as a systemic vaccine and via local administration. The demonstration of synergy between paracrine cytokine therapies holds promise as a novel therapy for brain tumors.

Animals↗
PubMed1995

Engineering an intracellular pathway for major histocompatibility complex class II presentation of antigens.

The presentation of antigenic peptides by major histocompatibility complex (MHC) class II molecules to CD4+ T cells is critical to the function of the immune system. In this study, we have utilized the sorting signal of the lysosomal-associated membrane protein LAMP-1 to target a model antigen, human papillomavirus 16 E7 (HPV-16 E7), into the endosomal and lysosomal compartments. The LAMP-1 sorting signal reroutes the antigen into the MHC class II processing pathway, resulting in enhanced presentation to CD4+ cells in vitro. In vivo immunization experiments in mice demonstrated that vaccinia containing the chimeric E7/LAMP-1 gene generated greater E7-specific lymphoproliferative activity, antibody titers, and cytotoxic T-lymphocyte activities than vaccinia containing the wild-type HPV-16 E7 gene. These results suggest that specific targeting of an antigen to the endosomal and lysosomal compartments enhances MHC class II presentation and vaccine potency.

Amino Acid Sequence↗
PubMed1995

A reassessment of the role of B7-1 expression in tumor rejection.

Introduction of the B7-1 gene into murine tumor cells can result in rejection of the B7-1 transductants and, in some cases, systemic immunity to subsequent challenge with the nontransduced tumor cells. These effects have been largely attributed to the function of B7-1 as a costimulator in directly activating tumor specific, major histocompatibility class I-restricted CD8+ T cells. We examined the role of B7-1 expression in the direct rejection as well as in the induction of systemic immunity to a nonimmunogenic murine tumor. B-16 melanoma cells with high levels of B7-1 expression did not grow in C57BL/6 recipient mice, while wild-type B-16 cells and cells with low B7-1 expression grew progressively within 21 d. In mixing experiments with B7-1hi and wild-type B-16 cells, tumors grew out in vivo even when a minority of cells were B7-1-. Furthermore, the occasional tumors that grew out after injection of 100% B-16 B7-1hi cells showed markedly decreased B7-1 expression. In vivo antibody depletions showed that NK1.1 and CD8+ T cells, but not CD4+ T cells, were essential for the in vivo rejection of tumors. Animals that rejected B-16 B7-1hi tumors did not develop enhanced systemic immunity against challenge with wild-type B-16 cells. These results suggest that a major role of B7-1 expression by tumors is to mediate direct recognition and killing by natural killer cells. With an intrinsically nonimmunogenic tumor, this direct killing does not lead to enhanced systemic immunity.

Animals↗
PubMed1995

Simplified high-sensitivity sequencing of a major histocompatibility complex class I-associated immunoreactive peptide using matrix-assisted laser desorption/ionization mass spectrometry.

Cytotoxic T cells (CTL) are known to recognize small peptide fragments of cytoplasmic proteins bound to major histocompatibility complex (MHC) class I molecules on cell surfaces. Recent work indicates that tumor antigens are processed and presented in a manner similar to viral antigens. Identification of the peptides recognized by tumor-specific CTL would provide valuable information about their parent proteins, as well as allowing for the development of recombinant antigen-specific tumor vaccines. While highly represented MHC-bound peptides have been routinely purified by reversed-phase HPLC for Edman degradation sequencing, identification and sequencing of infrequent peptides that represent the biologically relevant targets of tumor-specific CTL have proved elusive. We have combined matrix-assisted laser desorption/ionization mass spectrometry with on-slide exopeptidase digestion to successfully identify and directly sequence a model tumor-specific peptide antigen derived from an integrated viral gene. The enhanced sensitivity of this technique (femtomolar range) allows for the sequencing of specific MHC-bound peptides derived from as few as 1 x 10(9) cells.

Amino Acid Sequence↗
PubMed1995

Molecular characterization of defective antigen processing in human prostate cancer.

BACKGROUND: Gene-modified tumor cell vaccines have shown efficacy in animal models of malignancy, including prostate cancer. Class I major histocompatibility complex (MHC) assembly and function in the cellular targets of such therapies is pivotal in determining the efficacy of specific cytokine-secreting tumor vaccines. PURPOSE: To help guide development of genetically engineered vaccine therapy for human prostate cancer, potential immune resistance pathways were evaluated by analysis of class I MHC assembly in prostate cancer cells. METHOD: Class I MHC assembly in metastasis-derived human prostate cancer cell lines (LNCaP, PPC-1, DU-145, PC-3, and TSU) and a normal prostate-derived cell line (TP-2) were characterized by phenotypic, molecular, and functional assays. Assembled class I MHC and antigen was measured by flow cytometry; mRNA levels of assembly components (class I MHC heavy chain, beta 2-microglobulin, and the antigen transporter gene product TAP-2) were determined; and antigen processing was measured with a chimeric reconstituted system using vaccinia vectors. Restoration of antigen processing was attempted by interferon gamma stimulation and by transfection with mouse class I MHC heavy-chain cDNA. RESULTS: Assembled class I MHC was underexpressed in two (LNCaP and PPC-1) of five prostate cancer cell lines compared with normal prostate-derived controls. PPC-1 cells underexpressed TAP-2 mRNA despite abundant class I MHC and beta 2-microglobulin message. Induction of TAP-2 by interferon gamma indicated that coding sequences for TAP-2 message were present in PPC-1. Resistance to cytotoxic T lymphocytes (CTL) lysis showed a functional defect in antigen transport by PPC-1 cells; reversal of the molecular defect with interferon gamma led to restoration of functional antigen processing. In contrast, LNCaP cells had competent antigen transport but deficient class I MHC heavy-chain function despite abundant class I MHC RNA; though refractory to stimulation by interferon gamma, this defect responded to transfection of class I MHC heavy-chain cDNA. CONCLUSIONS: Metastatic prostate cancer cells can escape T-cell recognition via divergent mechanisms of defective class I MHC assembly. The specific underexpression of TAP-2 gene product in PPC-1 cells contrasts with prior studies of TAP gene underexpression in lung cancer (which concurrently underexpressed class I MHC heavy chain) and provides evidence for a regulatory pathway controlling TAP-2 gene expression in human cancers that may not affect class I MHC heavy-chain expression. IMPLICATIONS: In clinical application of gene therapy for prostate cancer, these findings provide a rationale for focusing on strategies that can circumvent sole reliance on class I MHC-mediated tumor cell recognition by CTL.

Antigen Presentation↗
PubMed1995

Paracrine cytokine adjuvants in cancer immunotherapy.

Advances in our understanding of the molecular events of antigen recognition by T cells and T cell activation are opening up new approaches to cancer immunotherapy. The identification and cloning of cytokines provide one important set of tools for manipulating immunologic responses. For cancer therapy, cytokines such as interleukin-2 have been administered systemically. However, systemic administration of cytokines ignores the paracrine nature of their action. Recently, an alternative approach has been explored that produces high concentrations of cytokines local to the tumor cells. This is achieved either by transduction of the tumor cells with the cytokine gene or by mixture of the tumor cells with cytokine containing biodegradable polymer microspheres. Under these circumstances, the locally released cytokine produces a strong local inflammatory response specific to the particular cytokine. In some cases, a potent tumor-specific T cell response results, capable of mediating regression of systemic tumor deposits. This paracrine delivery of cytokines can therefore be considered as a new type of adjuvant in the design of vaccines for cancer as well as microbial infections.

Adjuvants, Immunologic↗
PubMed1994

In vivo priming of two distinct antitumor effector populations: the role of MHC class I expression.

Downregulation of major histocompatibility complex (MHC) class I expression is an important mechanism by which tumors evade classical T cell-dependent immune responses. Therefore, a system was designed to evaluate parameters for active immunization against MHC class I- tumors. Mice were capable of rejecting a MHC class I- tumor challenge after immunization with an irradiated granulocyte/macrophage colony-stimulating factor (GM-CSF) transduced MHC class I- tumor vaccine. This response was critically dependent on CD4+ T cells and natural killer (NK) cells, but minimally on CD8+ T cells. A strong protective response against MHC class I+ variants of the tumor could be elicited when mice were immunized with irradiated MHC class I+ GM-CSF-secreting tumor cells. This response required CD4+ and CD8+ T cells, and in addition, elimination of NK cells resulted in outgrowth of tumors that had lost expression of at least one MHC class I gene. Finally, class I MHC expression on the vaccinating cells inhibited the response generated against a MHC class I- tumor challenge. These results demonstrate that the host is capable of being immunized against a tumor that has lost MHC class I expression and reveal conditions under which distinct effector cells play a role in the systemic antitumor immune response.

Animals↗