Journal articles: 'Gp100'

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Bakker, A. B., M. W. Schreurs, A. J. de Boer, Y. Kawakami, S. A. Rosenberg, G. J. Adema, and C. G. Figdor. "Melanocyte lineage-specific antigen gp100 is recognized by melanoma-derived tumor-infiltrating lymphocytes." Journal of Experimental Medicine 179, no. 3 (March 1, 1994): 1005–9. http://dx.doi.org/10.1084/jem.179.3.1005.

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We recently isolated a cDNA clone that encodes the melanocyte lineage-specific antigen glycoprotein (gp)100. Antibodies directed against gp100 are an important tool in the diagnosis of human melanoma. Since the gp100 antigen is highly expressed in melanocytic cells, we investigated whether this antigen might serve as a target for antimelanoma cytotoxic T lymphocytes (CTL). Here, we demonstrate that cytotoxic tumor-infiltrating lymphocytes (TIL) derived from a melanoma patient (TIL 1200) are directed against gp100. HLA-A2.1+ melanoma cells are lysed by TIL from this patient. In addition, murine double transfectants, expressing both HLA-A2.1 and gp100, are lysed by TIL 1200, whereas transfectants expressing only HLA-A2.1 are not susceptible to lysis. Furthermore, the HLA-A2.1+ melanoma cell line BLM, which lacks gp100 expression and is resistant to lysis, becomes susceptible after transfection of gp100 cDNA. Finally, HLA-A2.1+ normal melanocytes are lysed by TIL 1200. These data demonstrate that the melanocyte differentiation antigen gp100 can be recognized in the context of HLA-A2.1 by CTL from a melanoma patient. Gp100 may therefore constitute a useful target for specific immunotherapy against melanoma, provided that no unacceptable cytotoxicity towards normal tissue is observed.

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O'Day, S., F. S. Hodi, D. F. McDermott, R. W. Weber, J. A. Sosman, J. B. Haanen, X. Zhu, M. J. Yellin, A. Hoos, and W. J. Urba. "A phase III, randomized, double-blind, multicenter study comparing monotherapy with ipilimumab or gp100 peptide vaccine and the combination in patients with previously treated, unresectable stage III or IV melanoma." Journal of Clinical Oncology 28, no. 18_suppl (June 20, 2010): 4. http://dx.doi.org/10.1200/jco.2010.28.18_suppl.4.

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4 Background: Ipilimumab, a fully human monoclonal antibody against cytotoxic T-lymphocyte antigen-4, demonstrated activity in advanced melanoma. Gp100 vaccine showed immunological and clinical responses, and enhanced clinical activity when combined with other immunotherapy. This phase III study compared efficacy and safety of ipilimumab or gp100 monotherapy and combination. Methods: Eligible patients (HLA-A*0201+ previously treated adults with unresectable stage III/IV melanoma) were randomized 1:3:1 to ipilimumab (3 mg/kg q3w x 4 doses) + placebo (n=137), ipilimumab + gp100 (peptides 209-217[210M] and 280-288 [288V]; 1mg q3w x 4 doses; n=403), or gp100 + placebo (n=136). There was no maintenance phase. Primary endpoint was comparison of overall survival (OS) between patients who received combination versus gp100 alone; secondary endpoints were all other OS comparisons, best overall response rate (BORR), disease control rate (DCR) to W24, progression-free survival (PFS), and safety. Results: The study demonstrated statistically significant results for all efficacy endpoints (below). Ipilimumab alone or combined with gp100 resulted in a significant improvement in OS with risk reduction of 32-34% compared to gp100. Significant differences in DCR, BORR, and PFS were observed. Adverse events with ipilimumab were consistent with prior studies: generally mild, immune-related, and medically manageable. Conclusions: Ipilimumab is the first agent to improve median and long-term OS in a phase III study of previously treated patients with advanced melanoma. Addition of gp100 vaccine to ipilimumab did not improve outcome. [Table: see text] [Table: see text]

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Harvey, Becca, Dawn Lee, Anne-Francoise Gaudin, Beatrice Gueron, Bruno Bregman, Céleste Lebbé, and Isabelle Borget. "Changes in the quality of life of advanced melanoma patients after 12 weeks of treatment with ipilimumab compared to gp100 in a phase III clinical trial." Journal of Clinical Oncology 31, no. 15_suppl (May 20, 2013): 9084. http://dx.doi.org/10.1200/jco.2013.31.15_suppl.9084.

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9084 Background: This study analyses health-related quality of life (HRQL) outcomes for ipilimumab (ipi) with and without gp100 and gp100 alone during the 12 week treatment (T) induction period compared to baseline. The aim of this study was to express the HRQL as proportions of patients experiencing changes during the induction period, when most of the immune-related adverse events (ir-AE) on ipi occur. Methods: Data from the MDX010-20 trial, including 676 previously treated patients (pts) with unresectable stage III or IV melanoma, was analysed. Differences between ipi with/without gp100 and gp100 alone in terms of HRQL 12 weeks after randomisation according to the EORTC QLQ-C30 were searched for. The sample consisted of 388 pts (ipi: 83; ipi + gp100: 227; gp100: 78) completing both baseline and week (W)12 questionnaires. Ordinal regression was performed for these pts to determine a T effect on W12 scores (in ordered categories) whilst adjusting for key covariates. Fisher’s Exact Test was used to detect differences between Ts for each EORTC domain when considering mean change in scores, categorised as “clinically worse” (≤-10 on functional domains, ≥10 on symptoms), “stable” (>-10 to <10) and “clinically improved” (≥10 on functional domains, ≤-10 on symptoms). Results: There were no significant differences for any domain score between the 3 T arms at W12. The odds of attaining high HRQL scores at W12 were heavily dependent on the baseline scores; baseline scores explained most of the variability as per regression analysis. There was no significant difference between the 3 T arms in terms of pts showing a clinically significant reduction or improvement in HRQL. HRQL remained globally stable from baseline to W12, in > 50% of pts for most domains. Conclusions: Ipi with/without gp100 does not have a significant negative HRQL impact in stage III-IV melanoma during the T induction phase relative to gp100 alone after adjustment for differences in baseline scores. The lower rate of pts showing HQRL reduction and the absence of difference with gp100 suggest that ir-AE have little impact on HRQL. Further HRQL studies are needed, as efficacy benefits of ipi continue after W12.

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Lundqvist, Andreas, Sheila Rao, Maria Berg, Aleah Smith, Su Su, Hisayuki Yokoyama, Shivani Srivastava, and Richard Childs. "The Proteasome Inhibitor Bortezomib Simultaneously Enhances NK Cell Tumor Cytotoxicity While Paradoxically Reducing Antigen Specific T-Cell Tumor Cytotoxicity." Blood 110, no. 11 (November 16, 2007): 1789. http://dx.doi.org/10.1182/blood.v110.11.1789.1789.

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Abstract The proteasome inhibitor bortezomib was recently found to render tumor cells susceptible to natural killer (NK) cell-mediated apoptosis in vitro and in vivo. This sensitization appears to occur as a consequence of this agent up-regulating surface expression of tumor necrosis factor-related apoptosis-inducing ligand receptor 2 (TRAIL-R2) on human malignant cells rendering them susceptible to TRAIL-mediated NK cell cytotoxicity. We hypothesized that bortezomib would likewise sensitize tumors to the cytotoxic effects of antigen specific T-cells through similar apoptotic pathways, thereby providing an incentive to use bortezomib as a universal immune-sensitizing agent. The HLA-A2+, gp100+, MART-1+ melanoma cell lines 526 and 624 were treated with 10nM bortezomib for 18 hrs then were analyzed by FACS for expression the cell surface markers (HLA-ABC, MIC-A/B, TRAIL-R1/2 and Fas) and Cr51 cytotoxicity assay for susceptibility to CD8+/HLA-A2+ restricted gp100 and MART-1 specific CTL-mediated lysis. As observed previously, NK cell-mediated apoptosis was significantly higher in tumor cells treated with bortezomib compared to untreated tumor cells. In contrast, an unanticipated and significant reduction in CTL-mediated cytotoxicity was observed in tumors treated with bortezomib compared to untreated tumors; at an effector:target ratio of 3:1, NK cell cytotoxicity increased from 43±2% to 70±2% (p<0.01) while gp100 CTL cytotoxicity decreased from 34±4% to 18±2% (p<0.01) in 624 melanoma cells after exposure to bortezomib (figure). This inhibition in T-cell killing was not due to changes in tumor surface expression of MHC class I, MIC-A/B, TRAIL receptors or Fas. Remarkably, CTL-mediated cytotoxicity was restored to baseline in tumor cells that were pulsed with gp100 antigen following bortezomib treatment, suggesting proteasome inhibition by bortezomib altered or impaired the processing and presentation of the gp100 tumor antigen. Conclusions: Exposure of malignant cells to bortezomib results in simultaneous divergent effects on innate NK cell and adaptive T-cell anti-tumor immunity. While tumors exposed to bortezomib have enhanced susceptibility to NK-cell cytotoxicity, proteasome inhibition appears to disrupt antigen presentation potentially reducing tumor specific CTL effector responses. These findings suggest antigen specific T-cell responses such as graft-vs-host disease, and T-cell mediated graft-vs-tumor effects might be altered when bortezomib is administered following allogeneic hematopoietic cell transplantation. Figure. Melanoma cell line (624) was treated with bortezomib [10 nM] and analyzed for susceptibility to NK cell (left) and gp100-specific CD8+ CTL (middle) - mediated cytotoxicity in a 5h Cr51 cytotoxicity assay. Right - bortezomib-treated and untreated gp100:209 peptide pulsed 624 melanoma cells analyzed for susceptibility to gp100-specific CD8+ CTL-mediated cytotoxicity at a E:T ratio of 4:1 Figure. Melanoma cell line (624) was treated with bortezomib [10 nM] and analyzed for susceptibility to NK cell (left) and gp100-specific CD8+ CTL (middle) - mediated cytotoxicity in a 5h Cr51 cytotoxicity assay. Right - bortezomib-treated and untreated gp100:209 peptide pulsed 624 melanoma cells analyzed for susceptibility to gp100-specific CD8+ CTL-mediated cytotoxicity at a E:T ratio of 4:1

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Dowlath, Sasheen, Katrin Campbell, Farah Al-Barwani, Vivienne L. Young, Sarah L. Young, Greg F. Walker, and Vernon K. Ward. "Dry Formulation of Virus-Like Particles in Electrospun Nanofibers." Vaccines 9, no. 3 (March 3, 2021): 213. http://dx.doi.org/10.3390/vaccines9030213.

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Biologics can be combined with liquid polymer materials and electrospun to produce a dry nanofibrous scaffold. Unlike spray-drying and freeze-drying, electrospinning minimizes the physiological stress on sensitive materials, and nanofiber mat properties such as hydrophobicity, solubility, and melting temperature can be tuned based on the polymer composition. In this study, we explored the dry formulation of a virus-like particle (VLP) vaccine by electrospinning VLP derived from rabbit hemorrhagic disease virus modified to carry the MHC-I gp100 tumor-associated antigen epitope. VLP were added to a polyvinylpyrrolidone (PVP) solution (15% w/v) followed by electrospinning at 24 kV. Formation of a nanofibrous mat was confirmed by scanning electron microscopy, and the presence of VLP was confirmed by transmission electron microscopy and Western blot. VLP from the nanofibers induced T-cell activation and interferon- (IFN-) γ production in vitro. To confirm in vivo cytotoxicity, Pmel mice treated by injection with gp100 VLP from nanofibers induced a gp100 specific immune response, lysing approximately 65% of gp100-pulsed target cells, comparable to mice vaccinated with gp100 VLP in PBS. VLP from nanofibers also induced an antibody response. This work shows that electrospinning can be used to dry-formulate VLP, preserving both humoral and cell-mediated immunity.

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Cassaday, R., P. Sondel, D. King, T. Warner, A. Bridges, J. Gan, H. Schalch, J. Hank, D. Mahvi, and M. Albertini. "Clinical and immunological analysis of melanoma patients receiving immunization using particle-mediated gene transfer of genes for gp100 and GM-CSF into uninvolved skin." Journal of Clinical Oncology 24, no. 18_suppl (June 20, 2006): 13033. http://dx.doi.org/10.1200/jco.2006.24.18_suppl.13033.

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13033 Background: To investigate a new method of activating melanoma-specific immune responses, we examined in vivo particle-mediated gene transfer (PMGT) of cDNAs for gp100 and GM-CSF into uninvolved skin of melanoma patients (pts). We now report the analysis of a completed Phase I clinical study. Methods: Two treatment groups of 6 pts each were evaluated. Group I received PMGT with cDNA for gp100 during each 3 week cycle; Group II received PMGT with cDNA for GM-CSF followed 3 days later by PMGT for gp100 at the same site. PMGT used 0.25 ug DNA and 250 ug gold/treatment. Endpoints included vaccine toxicity, transgene expression, immunological activation, and antitumor effects. Results: No systemic toxicity could be attributed to the vaccines, while local toxicity in both groups included mild erythema and induration which resolved within 2 weeks. Monitoring for autoimmunity showed no induction of pathologic autoantibodies. Biopsies of vaccine sites obtained 2 days after the gp100 PMGT showed 16% of gold beads to be in the dermis in Group I vs 3% in Group II, suggesting the prior GM-CSF PMGT inhibited bead penetration (p < 0.001 by chi-square; each bead penetration was analyzed as an independent event). Biopsies in Group I obtained 2 days after vaccination showed 16% of beads in the dermis vs 22% after 4 days (p < 0.001 by chi-square; each bead penetration was analyzed as an independent event). Transgene expression in vaccinated skin sites was detected by ELISA (GM-CSF) and IHC (gp100). One of 4 HLA-A2+ subjects showed a 5 × 5-mm DTH response to gp100 peptide 210M after Cycle 1. Preliminary in vitro studies suggest minimal immunological activation. Of 4 pts who enrolled with no evidence of disease, 2 remain disease-free after 61–73 months of follow-up. Conclusions: PMGT with cDNA for gp100 and GM-CSF yields transgene expression in normal human skin with minimal local or systemic toxicity. Pathologic autoimmunity was not demonstrated. Bead concentration in the dermis increases over time, suggesting persistence of beads in this skin level. Conclusions related to melanoma-specific immune induction await T-cell and antibody studies. Supported in part by the UW General Clinical Research Center (M01 RR03186). No significant financial relationships to disclose.

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Voelkl, Simon, Tamson Moore, Michael Rehli, Michael Nishimura, Karin Fischer, and Andreas Mackensen. "Characterization of MHC Class-I Restricted TCRαβ+CD4−CD8− Double-Negative T Cells Recognizing the gp100 Antigen from a Melanoma Patient after gp100 Vaccination." Blood 112, no. 11 (November 16, 2008): 4905. http://dx.doi.org/10.1182/blood.v112.11.4905.4905.

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Abstract The immune attack against malignant tumors requires the concerted action of CD8+ cytotoxic T lymphocytes (CTL) as well as CD4+ T helper cells. The contribution of T cell receptor (TCR)αβ+ CD4− CD8− double-negative (DN) T cells to anti-tumor immune responses is widely unknown. In previous studies, we have demonstrated that DN T cells with a broad TCR repertoire are present in humans in the peripheral blood and the lymph nodes of healthy individuals. Here we characterize a human DN T cell clone (T4H2) recognizing an HLA-A2-restricted melanoma-associated antigenic gp100-peptide isolated from the peripheral blood of a melanoma patient. Antigen recognition by the T4H2 DN clone resulted in specific secretion of IFN-γ and TNF. Although lacking the CD8 molecule the gp100-specifc DN T cell clone was able to confer antigen-specific cytotoxicity against gp100-loaded target cells as well as HLA-A2+ gp100 expressing melanoma cells. The cytotoxic capacity was found to be perforin/granzymeB-dependent. Together, these data indicate that functionally active antigen-specific DN T cells recognizing MHC class I-restricted tumor-associated antigen (TAA) may contribute to anti-tumor immunity in vivo.

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McKenna, Philip M., Roger J. Pomerantz, Bernhard Dietzschold, James P. McGettigan, and Matthias J. Schnell. "Covalently Linked Human Immunodeficiency Virus Type 1 gp120/gp41 Is Stably Anchored in Rhabdovirus Particles and Exposes Critical Neutralizing Epitopes." Journal of Virology 77, no. 23 (December 1, 2003): 12782–94. http://dx.doi.org/10.1128/jvi.77.23.12782-12794.2003.

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ABSTRACT Rabies virus (RV) vaccine strain-based vectors show significant promise as potential live-attenuated vaccines against human immunodeficiency virus type 1 (HIV-1). Here we describe a new RV construct that will also likely have applications as a live-attenuated or killed-particle immunogen. We have created a RV containing a chimeric HIV-1 Env protein, which contains introduced cysteine residues that give rise to an intermolecular disulfide bridge between gp120 and the ectodomain of gp41. This covalently linked gp140 (gp140 SOS) is fused in frame to the cytoplasmic domain of RV G glycoprotein and is efficiently incorporated into the RV virion. On the HIV-1 virion, the gp120 and gp41 moieties are noncovalently associated, which leads to extensive shedding of gp120 from virions and virus-infected cells. The ability to use HIV-1 particles as purified, inactivated immunogens has been confounded by the loss of gp120 during preparation. Additionally, monomeric gp120 and uncleaved gp160 molecules have been shown to be poor antigenic representations of virion-associated gp160. Because the gp120 and gp41 portions are covalently attached in the gp140 SOS molecule, the protein is maintained on the surface of the RV virion throughout purification. Surface immunostaining and fluorescence-activated cell sorting analysis with anti-envelope antibodies show that the gp140 SOS protein is stably expressed on the surface of infected cells and maintains CD4 binding capabilities. Furthermore, Western blot and immunoprecipitation experiments with infected-cell lysates and purified virions show that a panel of neutralizing anti-envelope antibodies efficiently recognize the gp140 SOS protein. The antigenic properties of this recombinant RV particle containing covalently attached Env, as well as the ability to present Env in a membrane-bound form, suggest that this approach could be a useful component of a HIV-1 vaccine strategy.

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Overwijk, Willem W., Allan Tsung, Kari R. Irvine, Maria R. Parkhurst, Theresa J. Goletz, Kangla Tsung, Miles W. Carroll, et al. "gp100/pmel 17 Is a Murine Tumor Rejection Antigen: Induction of “Self”-reactive, Tumoricidal T Cells Using High-affinity, Altered Peptide Ligand." Journal of Experimental Medicine 188, no. 2 (July 20, 1998): 277–86. http://dx.doi.org/10.1084/jem.188.2.277.

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Many tumor-associated antigens are nonmutated, poorly immunogenic tissue differentiation antigens. Their weak immunogenicity may be due to “self”-tolerance. To induce autoreactive T cells, we studied immune responses to gp100/pmel 17, an antigen naturally expressed by both normal melanocytes and melanoma cells. Although a recombinant vaccinia virus (rVV) encoding the mouse homologue of gp100 was nonimmunogenic, immunization of normal C57BL/6 mice with the rVV encoding the human gp100 elicited a specific CD8+ T cell response. These lymphocytes were cross-reactive with mgp100 in vitro and treated established B16 melanoma upon adoptive transfer. To understand the mechanism of the greater immunogenicity of the human version of gp100, we characterized a 9-amino acid (AA) epitope, restricted by H-2Db, that was recognized by the T cells. The ability to induce specific T cells with human but not mouse gp100 resulted from differences within the major histocompatibility complex (MHC) class I–restricted epitope and not from differences elsewhere in the molecule, as was evidenced by experiments in which mice were immunized with rVV containing minigenes encoding these epitopes. Although the human (hgp10025–33) and mouse (mgp10025–33) epitopes were homologous, differences in the three NH2-terminal AAs resulted in a 2-log increase in the ability of the human peptide to stabilize “empty” Db on RMA-S cells and a 3-log increase in its ability to trigger interferon γ release by T cells. Thus, the fortuitous existence of a peptide homologue with significantly greater avidity for MHC class I resulted in the generation of self-reactive T cells. High-affinity, altered peptide ligands might be useful in the rational design of recombinant and synthetic vaccines that target tissue differentiation antigens expressed by tumors.

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Kaufman, Howard, Jose Lutzky, Joseph Clark, Kim Allyson Margolin, David H. Lawson, Asim Amin, Frances A. Collichio, et al. "Safety and efficacy of ipilimumab in melanoma patients who received prior immunotherapy on phase III study MDX010-020." Journal of Clinical Oncology 31, no. 15_suppl (May 20, 2013): 9050. http://dx.doi.org/10.1200/jco.2013.31.15_suppl.9050.

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9050 Background: MDX010-020 was a phase III comparison of ipilimumab (Ipi), gp100 vaccine or the combination for advanced melanoma. A subset of patients (pts) received other immunotherapy (IM) for advanced disease prior to receiving Ipi, providing the opportunity to evaluate safety and efficacy of Ipi following IM. A prior analysis has shown that pts receiving prior IL-2 had a similar overall survival (OS) to pts who had not received prior IL-2 [Hodi et al NEJM2010]; we now report expanded results for pts receiving any prior IM (interferons and/or interleukin). Methods: Eligible pts (n=676) had unresectable stage III/IV melanoma and were randomized 3:1:1 to q3 wks x 4 doses of Ipi + gp100 or Ipi + placebo or gp100 + placebo. All Ipi doses were 3 mg/kg i.v. OS was retrospectively analyzed for pts receiving any prior IM; immune-related adverse events (irAEs) during induction were evaluated for pts who received any prior IM (322 pts, 48%) and for pts who received prior IL-2 (154 pts, 23%). Results: Demography and OS are summarized below. irAEs of any grade were reported for 60% (Ipi) and 54% (Ipi + gp100) of pts receiving any prior IM. Those receiving prior IL-2 specifically had 73% (Ipi) and 58% (Ipi + gp100) incidence of any grade irAEs. Incidence was similar for those not receiving prior IM or prior IL-2. Diarrhea, rash, and pruritus were the most common events in all groups. Conclusions: Results for OS in this subgroup analysis were similar for both those receiving any prior IM and those who did not receive prior IM and to the overall 020 population. In addition, safety profiles were similar irrespective of prior immunotherapy. Clinical trial information: NCT00094653. [Table: see text]

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Khattar, Sunil K., Anthony L. DeVico, Celia C. LaBranche, Aruna Panda, David C. Montefiori, and Siba K. Samal. "Enhanced Immune Responses to HIV-1 Envelope Elicited by a Vaccine Regimen Consisting of Priming with Newcastle Disease Virus Expressing HIV gp160 and Boosting with gp120 and SOSIP gp140 Proteins." Journal of Virology 90, no. 3 (November 18, 2015): 1682–86. http://dx.doi.org/10.1128/jvi.02847-15.

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Newcastle disease virus (NDV) expressing HIV-1 BaL gp160 was evaluated either alone or with monomeric BaL gp120 and BaL SOSIP gp140 protein in a prime-boost combination in guinea pigs to enhance envelope (Env)-specific humoral and mucosal immune responses. We showed that a regimen consisting of an NDV prime followed by a protein boost elicited stronger serum and mucosal Th-1-biased IgG responses and neutralizing antibody responses than NDV-only immunizations. Additionally, these responses were higher after the gp120 than after the SOSIP gp140 protein boost.

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Middleton, Mark R., Neil Matthew Steven, TR Jeffry Evans, Jeffrey R. Infante, Omid Hamid, Alexander Noor Shoushtari, Philippa Gail Corrie, et al. "Pharmacodynamic effect of IMCgp100 (TCR–CD3 bispecific) on peripheral cytokines and association with overall survival in patients with advanced melanoma." Journal of Clinical Oncology 37, no. 15_suppl (May 20, 2019): 9523. http://dx.doi.org/10.1200/jco.2019.37.15_suppl.9523.

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9523 Background: ImmTAC molecules are unique TCR–anti-CD3 bispecifics that redirect T cells against intracellular antigens. IMCgp100, an ImmTAC targeting melanocyte-expressed gp100 antigen, has demonstrated monotherapy activity in advanced melanoma and can cause rash and cytokine-mediated AEs, hypothesized to be on-target (gp100) or effector (CD3) mediated. A preclinical MoA for T cell bispecifics suggests chemokine CXCL10 redirection of CXCR3+ T cells from blood into antigen-positive tissues; this has not been clinically validated. Methods: 84 HLA-A2+ pts with advanced melanoma (n = 61 cutaneous [CM], n = 19 uveal [UM], n = 4 other) received IMCgp100 (NCT01211262). Serum (n = 40) and PBMC (n = 22) samples were taken pre- and post-infusion to analyze changes in cytokines and circulating T cells. Pre- (n = 16) and post-treatment (n = 11) tumor biopsies were analyzed by IHC for CD3, PD-L1 and gp100 expression; tumor RNA (n = 12) was analyzed for gene expression. Results: IMCgp100 induced a transient increase in IFNg-inducible cytokines, most prominently CXCL10. A greater increase in serum CXCL10 was associated with longer OS (p = 0.0002), tumor shrinkage (p = 0.003), and greater transient reduction in peripheral CXCR3+CD8+ T cells (p = 0.001). Reduction in CXCR3+ CD8+ T cells also trended with longer OS (p = 0.02), and tumor shrinkage (p = 0.03). 3/16 pre-treatment biopsies had < 1% gp100 expression (all progressive disease). 8/11 biopsies post-IMCgp100 had increased CD3+ T cells compared with matched pre-treatment samples (associated with baseline gp100 but not PD-L1 expression). Based on tumor biopsy gene expression analysis, IMCgp100 increased T cell markers, IFNg-inducible and cytotoxicity-related genes. Conclusions: The association of clinical benefit with increased serum CXCL10 and decreased peripheral CXCR3+ T cells supports the MoA of IMCgp100-induced T cell redirection and activation. Tumor biopsy results support IMCgp100 redirection of T cells to antigen-positive tumor. A Phase II trial in CM (NCT02535078), a Phase I/II trial in UM (NCT02570308), and a Pivotal RCT in UM (NCT03070392) are ongoing. Clinical trial information: NCT01211262.

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Moss, D. J., and C. A. White. "A Ca2(+)-sensitive glycoprotein, GP90, associated with the cytoskeleton from brain and gizzard." Journal of Cell Science 93, no. 1 (May 1, 1989): 85–94. http://dx.doi.org/10.1242/jcs.93.1.85.

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We describe here the expression during development, tissue distribution and molecular properties of GP90: a major concanavalin A (ConA)-binding glycoprotein present in the neuronal membrane skeleton from chicken brain. GP90 is co-isolated with, and has a similar developmental profile to contactin (previously called GP130). In whole brain, GP90 undergoes rapid synthesis between embryonic days 10 and 12. Unlike contactin, it is not restricted to nervous tissue and is quite abundant in gizzard, where there are two antigenically related proteins of 100K and 90K (K = 10(3) Mr). In both brain and gizzard GP90 and (GP100) are enriched in the membrane skeleton fraction. Trypsinization of live cells suggest that GP90 from gizzard is related to GP100 by the removal of a polypeptide chain. GP90 from both neurones and gizzard cells is protected from proteolysis by the presence of extracellular Ca2+. In the absence of Ca2+ a soluble fragment of approximately 70K can be released from the surface of cells indicating that a large fraction of GP90 is extracellular. Deglycosylation of GP90 from brain using endoglycosidase F demonstrates the presence of at least five carbohydrate chains and a polypeptide chain of approximately 80K. Immunofluorescence studies show that GP90 is exposed on the surface of cultured neurones, gizzard cells and most glial cells with the exception of Schwann cells. It is observed in clusters or patches even when cells are prefixed, suggesting this may be the normal distribution of GP90.

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de Vries, I. Jolanda M., Monique R. Bernsen, W. Joost Lesterhuis, Nicole M. Scharenborg, Simon P. Strijk, Marie-Jeanne P. Gerritsen, Dirk J. Ruiter, Carl G. Figdor, Cornelis J. A. Punt, and Gosse J. Adema. "Immunomonitoring Tumor-Specific T Cells in Delayed-Type Hypersensitivity Skin Biopsies After Dendritic Cell Vaccination Correlates With Clinical Outcome." Journal of Clinical Oncology 23, no. 24 (August 20, 2005): 5779–87. http://dx.doi.org/10.1200/jco.2005.06.478.

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Purpose Tumor-specific immunomonitoring is essential to evaluate the efficacy of vaccination against cancer. In this study, we investigated the predictive value of the presence or absence of antigen-specific T cells in biopsies from delayed-type hypersensitivity (DTH) sites. Patients and Methods In our ongoing clinical trials, HLA-A2.1+ melanoma patients were vaccinated with mature dendritic cells (DC) pulsed with melanoma-associated peptides (gp100 and tyrosinase) and keyhole limpet hemocyanin. Results After intradermal administration of a DTH challenge with gp100- and tyrosinase peptide-loaded DC, essentially all patients showed a positive induration. In clinically responding patients, T cells specific for the antigen preferentially accumulated in the DTH site, as visualized by in situ tetramer staining. Furthermore, significant numbers of functional gp100 and tyrosinase tetramer-positive T cells could be isolated from these DTH biopsies, in accordance with the applied antigen in the DTH challenge. We observed a direct correlation between the presence of DC vaccine-related T cells in the DTH biopsies of stage IV melanoma patients and a positive clinical outcome (P = .0012). Conclusion These findings demonstrate the potency of this novel approach in the monitoring of vaccination studies in cancer patients.

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Vardeu, Mariantonella, David Depoil, Camille Britton-Rivet, Jane Houghton, Jane Harper, Gabrielle Le Provost, Laura Collins, Koustubh Ranade, and Adel Benlahrech. "624 IFNγ secreted by tebentafusp (IMCgp100)-redirected T cells inhibits expression of melanin synthesis pathway genes in healthy melanocytes." Journal for ImmunoTherapy of Cancer 8, Suppl 3 (November 2020): A660. http://dx.doi.org/10.1136/jitc-2020-sitc2020.0624.

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BackgroundTebentafusp (IMCgp100) is a bispecific T cell redirector comprised of an affinity-enhanced TCR recognising melanocyte lineage antigen gp100 and a T cell engaging anti-CD3 scFv domain. Tebentafusp has shown activity as monotherapy in advanced cutaneous and uveal melanoma (Middleton et al., ASCO 2019), and we have previously reported that over half of uveal melanoma patients treated with tebentafusp display melanocyte-related adverse events (MRAE). These include vitiligo/skin hypopigmentation, leukotrichia, and hyperpigmentation and, collectively, are associated with better overall survival in uveal patients receiving tebentafusp (Orloff et al, AACR 2020). In this study, we dissected the mechanisms by which tebentafusp may induce MRAE and highlight the potential clinical significance.MethodsIn vitro studies were conducted to assess the direct and indirect effects of tebentafusp on epidermal melanocytes from healthy donors. Expression of gp100 and the gp100:HLA*02:01 target complex by melanocytes were quantified at the mRNA level and on the cell surface by confocal microscopy, respectively. Melanocytes co-cultured with PBMC and increasing concentrations of tebentafusp were assessed for their susceptibility to lysis and/or ability to stimulate cytokine production. These readouts were compared to gp100-positive and negative melanoma cancer cell lines. Melanin production by melanocytes was quantified and the melanin synthesis pathway interrogated at the mRNA and protein level following exposure to secretomes from tebentafusp-redirected PBMC against melanoma cancer cells.ResultsHealthy melanocytes expressed 2 to 3-fold lower levels of gp100 peptide-HLA complexes on their surface compared to gp100-positive melanoma cell lines. In the presence of tebentafusp, this lower target expression translated into 3–6 fold lower levels of IFNγ and more than 100 fold lower granzyme B production by redirected T cells and these melanocytes were resistant to direct tebentafusp-induced killing (EC50 for melanocytes greater than 1nM vs EC50 melanoma cell lines of 23–50 pM). Supernatants from T cells activated in response to melanoma cancer cells by tebentafusp downregulated the melanin content of healthy melanocytes (20–30% reduction). Western blotting revealed 30–40% inhibition of two key components of the melanin synthesis pathway; the tyrosinase-related protein (TRP)-1 and TRP-2. This inhibition was reversed by blocking IFNγ in supernatants from activated T cells.ConclusionsMRAEs, especially vitiligo, associated with response to tebentafusp, may be explained, at least in part, by the downregulation of melanin biosynthesis pathway genes by IFNγ secreted by tebentafusp-activated T cells.Ethics ApprovalThe study was approved by the South Central - Oxford A Research Ethics Committee (UK), REC reference 13/SC/0226ReferencesMiddleton, et al., Relationship between clinical efficacy and AEs of IMCgp100, a novel bispecific TCR–anti-CD3, in patients with advanced melanoma. Journal of Clinical Oncology. 2019.Orloff, et al., Vitiligo and other clinical melanocyte-related adverse events following tebentafusp (IMCgp100) exposure in patients with uveal melanoma. AACR (American Association for Cancer Research), 2020.

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van Dinther, Dieke, Miguel Lopez Venegas, Henrike Veninga, Katarzyna Olesek, Leoni Hoogterp, Mirjam Revet, Martino Ambrosini, et al. "Activation of CD8+ T Cell Responses after Melanoma Antigen Targeting to CD169+ Antigen Presenting Cells in Mice and Humans." Cancers 11, no. 2 (February 5, 2019): 183. http://dx.doi.org/10.3390/cancers11020183.

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The lack of tumor-reactive T cells is one reason why immune checkpoint inhibitor therapies still fail in a significant proportion of melanoma patients. A vaccination that induces melanoma-specific T cells could potentially enhance the efficacy of immune checkpoint inhibitors. Here, we describe a vaccination strategy in which melanoma antigens are targeted to mouse and human CD169 and thereby induce strong melanoma antigen-specific T cell responses. CD169 is a sialic acid receptor expressed on a subset of mouse splenic macrophages that captures antigen from the blood and transfers it to dendritic cells (DCs). In human and mouse spleen, we detected CD169+ cells at an equivalent location using immunofluorescence microscopy. Immunization with melanoma antigens conjugated to antibodies (Abs) specific for mouse CD169 efficiently induced gp100 and Trp2-specific T cell responses in mice. In HLA-A2.1 transgenic mice targeting of the human MART-1 peptide to CD169 induced strong MART-1-specific HLA-A2.1-restricted T cell responses. Human gp100 peptide conjugated to Abs specific for human CD169 bound to CD169-expressing monocyte-derived DCs (MoDCs) and resulted in activation of gp100-specific T cells. Together, these data indicate that Ab-mediated antigen targeting to CD169 is a potential strategy for the induction of melanoma-specific T cell responses in mice and in humans.

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Voss, Ralf-Holger, Simone Thomas, Christina Pfirschke, Beate Hauptrock, Sebastian Klobuch, Jürgen Kuball, Margarete Grabowski, et al. "Coexpression of the T-cell receptor constant α domain triggers tumor reactivity of single-chain TCR-transduced human T cells." Blood 115, no. 25 (June 24, 2010): 5154–63. http://dx.doi.org/10.1182/blood-2009-11-254078.

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Abstract Transfer of tumor antigen–specific T-cell receptors (TCRs) into human T cells aims at redirecting their cytotoxicity toward tumors. Efficacy and safety may be affected by pairing of natural and introduced TCRα/β chains potentially leading to autoimmunity. We hypothesized that a novel single-chain (sc)TCR framework relying on the coexpression of the TCRα constant α (Cα) domain would prevent undesired pairing while preserving structural and functional similarity to a fully assembled double-chain (dc)TCR/CD3 complex. We confirmed this hypothesis for a murine p53-specific scTCR. Substantial effector function was observed only in the presence of a murine Cα domain preceded by a TCRα signal peptide for shuttling to the cell membrane. The generalization to a human gp100-specific TCR required the murinization of both C domains. Structural and functional T-cell avidities of an accessory disulfide-linked scTCR gp100/Cα were higher than those of a dcTCR. Antigen-dependent phosphorylation of the proximal effector ζ-chain–associated protein kinase 70 at tyrosine 319 was not impaired, reflecting its molecular integrity in signaling. In melanoma-engrafted nonobese diabetic/severe combined immunodeficient mice, adoptive transfer of scTCR gp100/Cα transduced T cells conferred superior delay in tumor growth among primary and long-term secondary tumor challenges. We conclude that the novel scTCR constitutes a reliable means to immunotherapeutically target hematologic malignancies.

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Boutin, P., S. Shankara, C. Nicolette, and B. Roberts. "Maximizing antigen presentation of GP100 using adenoviral vectors." Journla of Immunotherapy 22, no. 5 (September 1999): 455. http://dx.doi.org/10.1097/00002371-199909000-00013.

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Luiten, Rosalie M., Esther W. M. Kueter, Wolter Mooi, Maarten P. W. Gallee, Elaine M. Rankin, Winald R. Gerritsen, Shirley M. Clift, et al. "Immunogenicity, Including Vitiligo, and Feasibility of Vaccination With AutologousGM-CSF–Transduced Tumor Cells in Metastatic Melanoma Patients." Journal of Clinical Oncology 23, no. 35 (December 10, 2005): 8978–91. http://dx.doi.org/10.1200/jco.2005.01.6816.

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PurposeTo determine the feasibility, toxicity, and immunologic effects of vaccination with autologous tumor cells retrovirally transduced with the GM-CSF gene, we performed a phase I/II vaccination study in stage IV metastatic melanoma patients.Patients and MethodsSixty-four patients were randomly assigned to receive three vaccinations of high-dose or low-dose tumor cells at 3-week intervals. Tumor cell vaccine preparation succeeded for 56 patients (88%), but because of progressive disease, the well-tolerated vaccination was completed in only 28 patients. We analyzed the priming of T cells against melanoma antigens, MART-1, tyrosinase, gp100, MAGE-A1, and MAGE-A3 using human leukocyte antigen/peptide tetramers and functional assays.ResultsThe high-dose vaccination induced the infiltration of T cells into the tumor tissue. Three of 14 patients receiving the high-dose vaccine showed an increase in MART-1– or gp100-specific T cells in the peripheral blood during vaccination. Six patients experienced disease-free survival for more than 5 years, and two of these patients developed vitiligo at multiple sites after vaccination. MART-1– and gp100-specific T cells were found infiltrating in vitiligo skin. Upon vaccination, the T cells acquired an effector phenotype and produced interferon-γ on specific antigenic stimulation.ConclusionWe conclude that vaccination with GM-CSF–transduced autologous tumor cells has limited toxicity and can enhance T-cell activation against melanocyte differentiation antigens, which can lead to vitiligo. Whether the induction of autoimmune vitiligo may prolong disease-free survival of metastatic melanoma patients who are surgically rendered as having no evidence of disease before vaccination is worthy of further investigation.

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Strobel, Sophia B., Devayani Machiraju, Ingrid Hülsmeyer, Jürgen C. Becker, Annette Paschen, Dirk Jäger, Winfried S. Wels, Michael Bachmann, and Jessica C. Hassel. "Expression of Potential Targets for Cell-Based Therapies on Melanoma Cells." Life 11, no. 4 (March 24, 2021): 269. http://dx.doi.org/10.3390/life11040269.

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Tumor antigen-specific redirection of cytotoxic T cells (CTLs) or natural killer (NK) cells including chimeric antigen receptor (CAR-) and T cell receptor (TCR-) cell therapy is currently being evaluated in different tumor entities including melanoma. Expression of melanoma-specific antigen recognized by the respective CAR or TCR directly or presented by HLA molecules is an indispensable prerequisite for this innovative therapy. In this study, we investigated in 168 FFPE tumor specimens of patients with stage I-IV melanoma the protein expression of HER2, TRP2, ABCB5, gp100, p53, and GD2 by immunohistochemistry (IHC). These results were correlated with clinical parameters. Membrane expression of HER2 and GD2 was also investigated in ten melanoma cell lines by flow cytometry for which corresponding tumors were analyzed by IHC. Our results demonstrated that gp100 was the most frequently overexpressed protein (61%), followed by TRP2 (50%), GD2 (38%), p53 (37%), ABCB5 (17%), and HER2 (3%). TRP2 expression was higher in primary tumors compared to metastases (p = 0.005). Accordingly, TRP2 and ABCB5 expression was significantly associated with lower tumor thickness of the primary (p = 0.013 and p = 0.025). There was no association between protein expression levels and survival in advanced melanoma patients. Flow cytometric analysis revealed abundant surface expression of GD2 and HER2 in all melanoma cell lines. The discordant HER2 expression in situ and in vitro suggests a tissue culture associated induction. In summary, our data support the use of gp100 and GD2 as a potential target for developing engineered TCR- or CAR-cell therapies, respectively, against melanoma.

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Kawakami, Y., S. Eliyahu, K. Sakaguchi, P. F. Robbins, L. Rivoltini, J. R. Yannelli, E. Appella, and S. A. Rosenberg. "Identification of the immunodominant peptides of the MART-1 human melanoma antigen recognized by the majority of HLA-A2-restricted tumor infiltrating lymphocytes." Journal of Experimental Medicine 180, no. 1 (July 1, 1994): 347–52. http://dx.doi.org/10.1084/jem.180.1.347.

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Four melanoma proteins, MART-1, gp100, tyrosinase, and tyrosinase-related protein-1 (gp75) were evaluated for recognition by HLA-A2-restricted melanoma-specific cytotoxic T lymphocytes (CTLs) derived from the tumor-infiltrating lymphocytes (TIL) of 10 different patients. 9 of 10 TIL recognized MART-1, 4 recognized gp100 (including 3 that also recognized MART-1), but none of the TIL recognized tyrosinase or gp75. Based on the known HLA-A2.1 peptide binding motifs, 23 peptides from MART-1 were synthesized in an attempt to identify the epitopes recognized by TIL. Three peptides were recognized by TIL when pulsed on T2 target cells. One of the 9-mer peptides, AAGIGILTV, was most effective in sensitizing the T2 cells for TIL lysis. This peptide was recognized by 9 of 10 HLA-A2-restricted melanoma-specific CTLs. Therefore, this peptide appears to be a very common immunogenic epitope for HLA-A2-restricted melanoma-specific TIL and may be useful for the development of immunotherapeutic strategies.

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Frankenburg, Shoshana, Igor Grinberg, Ziva Bazak, Lena Fingerut, Jacob Pitcovski, Raphael Gorodetsky, Tamar Peretz, Ram M. Spira, Yehuda Skornik, and Ronald S. Goldstein. "Immunological activation following transcutaneous delivery of HR-gp100 protein." Vaccine 25, no. 23 (June 2007): 4564–70. http://dx.doi.org/10.1016/j.vaccine.2007.04.025.

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Yasumoto, Ken-ichi, Hidenori Watabe, Julio C. Valencia, Tsuneto Kushimoto, Takeshi Kobayashi, Ettore Appella, and Vincent J. Hearing. "Epitope Mapping of the Melanosomal Matrix Protein gp100 (PMEL17)." Journal of Biological Chemistry 279, no. 27 (April 19, 2004): 28330–38. http://dx.doi.org/10.1074/jbc.m401269200.

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Jakobsen, B. K. "ImmTAC-gp100: A novel, targeted therapy for malignant melanoma." Journal of Clinical Oncology 28, no. 15_suppl (May 20, 2010): e19036-e19036. http://dx.doi.org/10.1200/jco.2010.28.15_suppl.e19036.

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Koch, G. L. E., M. J. Smith, and R. A. Mortara. "An abundant ubiquitous glycoprotein (GP100 ) in nucleated mammalian cells." FEBS Letters 179, no. 2 (January 7, 1985): 294–98. http://dx.doi.org/10.1016/0014-5793(85)80537-8.

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Dionne, Sara O., Margaret H. Smith, Francesco M. Marincola, and Douglas F. Lake. "Functional characterization of CTL against gp100 altered peptide ligands." Cancer Immunology, Immunotherapy 52, no. 4 (April 2003): 199–206. http://dx.doi.org/10.1007/s00262-002-0358-3.

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Adema, G. J., A. J. de Boer, A. M. Vogel, W. A. Loenen, and C. G. Figdor. "Molecular characterization of the melanocyte lineage-specific antigen gp100." Journal of Biological Chemistry 269, no. 31 (August 1994): 20126–33. http://dx.doi.org/10.1016/s0021-9258(17)32136-1.

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Slingluff, Craig L., Gina R. Petroni, Galina V. Yamshchikov, Sarah Hibbitts, William W. Grosh, Kimberly A. Chianese-Bullock, Eric A. Bissonette, et al. "Immunologic and Clinical Outcomes of Vaccination With a Multiepitope Melanoma Peptide Vaccine Plus Low-Dose Interleukin-2 Administered Either Concurrently or on a Delayed Schedule." Journal of Clinical Oncology 22, no. 22 (November 15, 2004): 4474–85. http://dx.doi.org/10.1200/jco.2004.10.212.

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Purpose A phase II trial was performed to test whether systemic low-dose interleukin-2 (IL-2) augments T-cell immune responses to a multipeptide melanoma vaccine. Forty patients with resected stage IIB-IV melanoma were randomly assigned to vaccination with four gp100- and tyrosinase-derived peptides restricted by human leukocyte antigen (HLA) -A1, HLA-A2, and HLA-A3, and a tetanus helper peptide plus IL-2 administered daily either beginning day 7 (group 1), or beginning day 28 (group 2). Patients and Methods T-cell responses were assessed by an interferon gamma ELIspot assay in peripheral blood lymphocytes (PBL) and in a lymph node draining a vaccination site (sentinel immunized node [SIN]). Patients were followed for disease-free and overall survival. Results T-cell responses to the melanoma peptides were observed in 37% of PBL and 38% of SINs in group 1, and in 53% of PBL and 83% of SINs in group 2. The magnitude of T-cell response was higher in group 2. The tyrosinase peptides DAEKSDICTDEY and YMDGTMSQV were more immunogenic than the gp100 peptides YLEPGPVTA and ALLAVGATK. T-cell responses were detected in the SINs more frequently, and with higher magnitude, than responses in the PBL. Disease-free survival estimates at 2 years were 39% (95% CI, 18% to 61%) for group 1, and 50% (95% CI, 28% to 72%) for group 2 (P = .32). Conclusion The results of this study support the safety and immunogenicity of a vaccine composed of four peptides derived from gp100 and tyrosinase. The low-dose IL-2 regimen used for group 1 paradoxically diminishes the magnitude and frequency of cytotoxic T lymphocyte responses to these peptides.

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Attia, Peter, Giao Q. Phan, Ajay V. Maker, Michael R. Robinson, Martha M. Quezado, James C. Yang, Richard M. Sherry, et al. "Autoimmunity Correlates With Tumor Regression in Patients With Metastatic Melanoma Treated With Anti–Cytotoxic T-Lymphocyte Antigen-4." Journal of Clinical Oncology 23, no. 25 (September 1, 2005): 6043–53. http://dx.doi.org/10.1200/jco.2005.06.205.

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Purpose Previously, we reported our experience treating 14 patients with metastatic melanoma using a fully human antibody to cytotoxic T-lymphocyte antigen-4 (anti–CTLA-4) in conjunction with peptide vaccination. We have now treated 56 patients to evaluate two different dose schedules of anti–CTLA-4 and to explore the relationship between autoimmunity and tumor regression. Patients and Methods A total of 56 patients with progressive stage IV melanoma were enrolled onto the study. All had Karnofsky performance status ≥ 60% with no prior history of autoimmunity. Twenty-nine patients received 3 mg/kg anti–CTLA-4 every 3 weeks, whereas 27 received 3 mg/kg as their initial dose with subsequent doses reduced to 1 mg/kg every 3 weeks. In both cohorts patients received concomitant vaccination with two modified HLA-A*0201-restricted peptides from the gp100 melanoma-associated antigen, gp100:209-217(210M) and gp100:280-288(288V). Results Two patients achieved a complete response (ongoing at 30 and 31 months, respectively) and five patients achieved a partial response (durations of 4, 6, 25+, 26+, and 34+ months, respectively), for an overall objective response rate of 13%. Tumor regression was seen in lung, liver, brain, lymph nodes, and subcutaneous sites. Of 14 patients with grade 3/4 autoimmune toxicity, five (36%) experienced a clinical response compared with only two responses in the 42 patients (5%) with no autoimmune toxicity (P = .008). There were no significant differences in response rate or toxicity between the two dose schedules. Conclusion Administration of anti–CTLA-4 monoclonal antibody plus peptide vaccination can cause durable objective responses, which correlate with the induction of autoimmunity, in patients with metastatic melanoma.

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Neudorfer, Julia, Daniel Sommermeyer, Christian Peschel, Thomas Blankenstein, Wolfgang Uckert, and Helga Bernhard. "Redirecting Human T Lymphocytes toward Tumor-Associated Antigens by T Cell Receptor (TCR) Replacement." Blood 108, no. 11 (November 16, 2006): 3711. http://dx.doi.org/10.1182/blood.v108.11.3711.3711.

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Abstract The gene transfer of alpha and beta chains derived from a defined TCR has been successfully applied to endow T cells with specificities directed against tumor-associated antigens. However, it is still unclear if the transfer of TCR genes into T cells that already express an endogenous TCRalpha and beta chain leads to engineered T cells expressing four different TCR complexes on their cell surface. Mixed TCR heterodimers composed of endogenous and exogenous TCR chains may acquire new specificities, which may cause unwanted reactions in patients following adoptive T cell transfer. We examined the possibility of mixed TCR heterodimer formation using defined conditions of single TCR chain transfer into human cytotoxic T cell (CTL) clones specific for CMV and Melan-A, respectively. After stimulation for three days CTLs were retrovirally transduced with the beta chain derived from a gp100-specific TCR. The expression of the exogenous (transduced) and the endogenous beta chain was distinguished by flow cytometry using antibodies against the different Vbeta motives. Indeed, CTLs that had been transduced with the single beta chain expressed this chain on the cell surface indicating the formation of mixed TCRs, because the expression of the exogenous TCRbeta chain requires the pairing with the endogenous TCRalpha chain. Furthermore, we transduced the CTL clones with both the TCRalpha and beta chain derived from a gp100-specific TCR. The transduced T cells were positively stained with an A2/gp100 multimer documenting the correct formation of the exogenous TCR chains. Functionality of transduced CTL clones was tested by antigen-specific IFN-gamma release and cytolytic activity. The TCR-transduced T cells were sorted with the A2/gp100 multimer and expanded for two weeks. Double staining with HLA multimers for the endogenous and the transduced TCRs showed the downregulation of the endogenous TCRs in three different CTL clones. In one CTL clone, the endogenous TCR was even replaced by the exogenous TCR as documented by flowcytometry and antigen-specific T cell function. In conclusion, transfer of single TCR chains in CTL clones can result in the formation of TCR heterodimers. However, our results also show that complete TCRs are predominantly expressed or can even replace other TCRs following transfer. The development of dominant TCRs will facilitate the therapeutical approaches of adoptive transfer regimens based on TCR-transduced T cells, because dominant TCRs can be selected or TCRs can be modified to be more dominant.

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Schwartzentruber, D. J., D. Lawson, J. Richards, R. M. Conry, D. Miller, J. Triesman, F. Gailani, L. B. Riley, D. Vena, and P. Hwu. "A phase III multi-institutional randomized study of immunization with the gp100: 209–217(210M) peptide followed by high-dose IL-2 compared with high-dose IL-2 alone in patients with metastatic melanoma." Journal of Clinical Oncology 27, no. 18_suppl (June 20, 2009): CRA9011. http://dx.doi.org/10.1200/jco.2009.27.18_suppl.cra9011.

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CRA9011 Background: In a phase II study, 13 (42%) of 31 patients with metastatic melanoma receiving high-dose (HD) IL-2 plus gp100:209–217(210M) peptide experienced objective responses (S.A. Rosenberg, et al, Nature Medicine 4: 321–327, 1998). Other studies showed a lower response rate (RR) but no randomized studies have been done. Methods: A prospective randomized phase III trial was conducted at 21 centers with 185 patients. Eligibility: stage IV or locally advanced stage III cutaneous melanoma, HLA A0201, no brain metastases, eligible for HD IL-2, and no previous HD IL-2 or gp100:209–217(210M). Arm 1 received HD IL-2 alone (720,000 IU/kg/dose) and Arm 2 gp100:209–217(210M) peptide + Montanide ISA followed by HD IL-2. The primary objective was clinical response. Secondary objectives were toxicity, disease free/progression free survival, immunologic response and quality of life. Central HLA typing, pathology review, and blinded response assessment were done at the NIH. Central data monitoring was done by The EMMES Corp. and a Data Safety Monitoring Board. Results: Numbers of patients enrolled, treated, and evaluable for response in Arm 1 were 94, 93, and 93 respectively; in Arm 2 91, 86, and 86. Toxicities were consistent with HD IL-2 ± vaccine. Investigator assessed RR showed significant improvement in overall RR for Arm 2=22.1% vs 9.7% (P=0.0223, Chi-Square) and progression free survival (PFS) in favor of Arm 2=2.9 months (1.7–4.5) vs 1.6 (1.5–1.8) (P=0.0101). Median overall survival favors Arm 2=17.6 months (11.8–26.6) vs 12.8 (8.7–16.3) (P=0.0964). Blinded response review is ongoing. Conclusions: RR and PFS were superior with peptide vaccine and HD IL-2 compared to HD IL-2 alone. This represents the first evidence of clinical benefit of vaccination in patients with melanoma. [Table: see text]

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Derby, Nina R., Zane Kraft, Elaine Kan, Emma T. Crooks, Susan W. Barnett, Indresh K. Srivastava, James M. Binley, and Leonidas Stamatatos. "Antibody Responses Elicited in Macaques Immunized with Human Immunodeficiency Virus Type 1 (HIV-1) SF162-Derived gp140 Envelope Immunogens: Comparison with Those Elicited during Homologous Simian/Human Immunodeficiency Virus SHIVSF162P4 and Heterologous HIV-1 Infection." Journal of Virology 80, no. 17 (September 1, 2006): 8745–62. http://dx.doi.org/10.1128/jvi.00956-06.

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ABSTRACT The antibody responses elicited in rhesus macaques immunized with soluble human immunodeficiency virus (HIV) Env gp140 proteins derived from the R5-tropic HIV-1 SF162 virus were analyzed and compared to the broadly reactive neutralizing antibody responses elicited during chronic infection of a macaque with a simian/human immunodeficiency virus (SHIV) expressing the HIV-1 SF162 Env, SHIVSF162P4, and humans infected with heterologous HIV-1 isolates. Four gp140 immunogens were evaluated: SF162gp140, ΔV2gp140 (lacking the crown of the V2 loop), ΔV3gp140 (lacking the crown of the V3 loop), and ΔV2ΔV3gp140 (lacking both the V2 and V3 loop crowns). SF162gp140 and ΔV2gp140 have been previously evaluated by our group in a pilot study, but here, a more comprehensive analysis of their immunogenic properties was performed. All four gp140 immunogens elicited stronger anti-gp120 than anti-gp41 antibodies and potent homologous neutralizing antibodies (NAbs) that primarily targeted the first hypervariable region (V1 loop) of gp120, although SF162gp140 also elicited anti-V3 NAbs. Heterologous NAbs were elicited by SF162gp140 and ΔV2gp140 but were weak in potency and narrow in specificity. No heterologous NAbs were elicited by ΔV3gp140 or ΔV2ΔV3gp140. In contrast, the SHIVSF162P4-infected macaque and HIV-infected humans generated similar titers of anti-gp120 and anti-gp41 antibodies and NAbs of significant breadth against primary HIV-1 isolates, which did not target the V1 loop. The difference in V1 loop immunogenicity between soluble gp140 and virion-associated gp160 Env proteins derived from SF162 may be the basis for the observed difference in the breadth of neutralization in sera from the immunized and infected animals studied here.

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Sutmuller, Roger P. M., Luc R. H. M. Schurmans, Leonie M. van Duivenvoorde, John A. Tine, Ellen I. H. van der Voort, René E. M. Toes, Cornelis J. M. Melief, Martine J. Jager, and Rienk Offringa. "Adoptive T Cell Immunotherapy of Human Uveal Melanoma Targeting gp100." Journal of Immunology 165, no. 12 (December 15, 2000): 7308–15. http://dx.doi.org/10.4049/jimmunol.165.12.7308.

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Gelbart, Y., S. Frankenburg, Y. Pinchasov, S. Krispel, D. Eliahu, O. Drize, E. Morag, et al. "Production and purification of melanoma gp100 antigen and polyclonal antibodies." Protein Expression and Purification 34, no. 2 (April 2004): 183–89. http://dx.doi.org/10.1016/j.pep.2003.12.006.

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Zhou, W.-Z., Y. Kaneda, S. K. S. Huang, R. Morishita, and D. S. B. Hoon. "Protective immunization against melanoma by gp100 DNA–HVJ-liposome vaccine." Gene Therapy 6, no. 10 (October 1999): 1768–73. http://dx.doi.org/10.1038/sj.gt.3300998.

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Kawakami, Akinori, Fumio Sakane, Shin-ichi Imai, Satoshi Yasuda, Masahiro Kai, Hideo Kanoh, Hai-Ying Jin, et al. "Rab7 Regulates Maturation of Melanosomal Matrix Protein gp100/Pmel17/Silv." Journal of Investigative Dermatology 128, no. 1 (January 2008): 143–50. http://dx.doi.org/10.1038/sj.jid.5700964.

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Voelkl, Simon, Tamson V. Moore, Michael Rehli, Michael I. Nishimura, Andreas Mackensen, and Karin Fischer. "Characterization of MHC class-I restricted TCRαβ+ CD4− CD8− double negative T cells recognizing the gp100 antigen from a melanoma patient after gp100 vaccination." Cancer Immunology, Immunotherapy 58, no. 5 (October 3, 2008): 709–18. http://dx.doi.org/10.1007/s00262-008-0593-3.

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Andreola, Giovanna, Licia Rivoltini, Chiara Castelli, Veronica Huber, Paola Perego, Paola Deho, Paola Squarcina, et al. "Induction of Lymphocyte Apoptosis by Tumor Cell Secretion of FasL-bearing Microvesicles." Journal of Experimental Medicine 195, no. 10 (May 20, 2002): 1303–16. http://dx.doi.org/10.1084/jem.20011624.

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The hypothesis that FasL expression by tumor cells may impair the in vivo efficacy of antitumor immune responses, through a mechanism known as ‘Fas tumor counterattack,’ has been recently questioned, becoming the object of an intense debate based on conflicting results. Here we definitely show that FasL is indeed detectable in the cytoplasm of melanoma cells and its expression is confined to multivesicular bodies that contain melanosomes. In these structures FasL colocalizes with both melanosomal (i.e., gp100) and lysosomal (i.e., CD63) antigens. Isolated melanosomes express FasL, as detected by Western blot and cytofluorimetry, and they can exert Fas-mediated apoptosis in Jurkat cells. We additionally show that melanosome-containing multivesicular bodies degranulate extracellularly and release FasL-bearing microvesicles, that coexpress both gp100 and CD63 and retain their functional activity in triggering Fas-dependent apoptosis of lymphoid cells. Hence our data provide evidence for a novel mechanism potentially operating in Fas tumor counterattack through the secretion of subcellular particles expressing functional FasL. Such vesicles may form a sort of front line hindering lymphocytes and other immunocompetent cells from entering neoplastic lesions and exert their antitumor activity.

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Hassel, Jessica Cecile, Piotr Rutkowski, Jean-Francois Baurain, Marcus O. Butler, Max Schlaak, Ryan Sullivan, Sebastian Ochsenreither, et al. "Co-primary endpoint of overall survival for tebentafusp (tebe)-induced rash in a phase 3 randomized trial comparing tebe versus investigator’s choice (IC) in first-line metastatic uveal melanoma." Journal of Clinical Oncology 39, no. 15_suppl (May 20, 2021): 9527. http://dx.doi.org/10.1200/jco.2021.39.15_suppl.9527.

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9527 Background: Tebe is a bispecific consisting of an affinity-enhanced T cell receptor fused to an anti-CD3 effector that can redirect T cells to target gp100+ cells. In this Phase (Ph) 3, randomized trial of first line (1L) metastatic uveal melanoma (mUM) [NCT03070392], tebe significantly improved overall survival (OS) vs. investigator’s choice (IC) in the intention-to-treat population (ITT). In previous trials, tebe-related skin adverse events (AEs), hypothesized to be on-target, off-tumor activity against gp100-expressing melanocytes, were associated with improved OS. This association was tested prospectively as a co-primary endpoint in the Ph3 study. Methods: 378 1L HLA-A*02:01+ mUM pts were randomized 2:1 to tebe (n = 252) or IC (n = 126). Co-primary endpoints were 1) OS in all randomized pts (ITT) and 2) OS in tebe-randomized pts who develop any grade rash in week (wk) 1 vs. all receiving IC. Rash was defined as composite of preferred AE terms. Melanocyte-related AEs (MRAEs) were defined as pigment change AEs in the skin or hair. Overall study-wide alpha was controlled at 0.05, with 90% assigned to ITT and 10% to rash. This analysis was conducted on the first interim analysis (data extracted Nov-2020). Results: In the 245 tebe treated pts, the characteristic skin related AEs included most frequently rash (at any time) in 201 pts (82%), pruritis in 167 pts (68%), MRAEs in 109 pts (45%) and erythema in 69 pts (28%). While rash, erythema and pruritis mostly occurred in the first 4 weeks, MRAEs occurred after a median of 2.7 mo. Rash captures most pts, 201/227 (89%), who have any of these skin related AEs. Rash occurred in 146 pts (60%) by wk 1; 179 pts (73%) by wk 2; and 195 pts (80%) by wk 3. Tebe pts with wk 1 rash had significantly longer OS vs. the IC arm, HR 0.35 (95% CI 0.23, 0.53), p < 0.0001. The estimated 1-yr OS rates were 83% vs 58%, respectively. When expanded to include tebe pts with rash through wk 3, the 1-yr OS rate of 75% was still numerically higher than IC. The 50 (20%) tebe pts who did not experience rash by week 3 had 1-yr OS rate of 55%. Conclusions: In 1L mUM pts, tebe significantly improved OS compared to IC in the ITT analysis. Week 1 rash, presumed due to tebe redirection of T cells to gp100+ skin melanocytes, was associated with a very strong OS benefit. Therefore, rash may be a marker that the immune system can be mobilized by tebe to target gp100+ cells. The vast majority of tebe pts will develop a rash at some point, and tebe pts without rash may still derive benefit. Clinical trial information: NCT03070392.

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Przybyla, Anna, Alexander A. Lehmann, Ting Zhang, Jacek Mackiewicz, Łukasz Galus, Greg A. Kirchenbaum, Andrzej Mackiewicz, and Paul V. Lehmann. "Functional T Cell Reactivity to Melanocyte Antigens Is Lost during the Progression of Malignant Melanoma, but Is Restored by Immunization." Cancers 13, no. 2 (January 9, 2021): 223. http://dx.doi.org/10.3390/cancers13020223.

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Healthy human subjects develop spontaneous CD8+ T cell responses to melanoma associated antigens (MA) expressed by normal melanocytes, such as Tyrosinase, MAGE-A3, Melan/Mart-1, gp100, and NY-ESO-1. This natural autoimmunity directed against melanocytes might confer protection against the development of malignant melanoma (MM), where MA are present as overexpressed tumor-associated antigens. Consistent with this notion we report here that functional T cell reactivity to MA was found to be significantly diminished to MAGE-A3, Melan-A/Mart-1, and gp100 in untreated MM patients. Three lines of evidence suggest that the MA-reactive T cells present in healthy subjects undergo exhaustion once MM establishes itself. First, only the MA-specific T cell reactivity was affected in the MM patients; that to third party recall antigens was not. Second, in these patients, the residual MA-specific T cells, unlike third party antigen reactive T cells, were functionally impaired, showing a diminished per cell IFN-γ productivity. Third, we show that immunization with MA restored natural CD8+ T cell autoimmunity to MA in 85% of the MM patients. The role of natural T cell autoimmunity to tumor-associated MA is discussed based on discrete levels of T cell activation thresholds.

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Przybyla, Anna, Alexander A. Lehmann, Ting Zhang, Jacek Mackiewicz, Łukasz Galus, Greg A. Kirchenbaum, Andrzej Mackiewicz, and Paul V. Lehmann. "Functional T Cell Reactivity to Melanocyte Antigens Is Lost during the Progression of Malignant Melanoma, but Is Restored by Immunization." Cancers 13, no. 2 (January 9, 2021): 223. http://dx.doi.org/10.3390/cancers13020223.

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Healthy human subjects develop spontaneous CD8+ T cell responses to melanoma associated antigens (MA) expressed by normal melanocytes, such as Tyrosinase, MAGE-A3, Melan/Mart-1, gp100, and NY-ESO-1. This natural autoimmunity directed against melanocytes might confer protection against the development of malignant melanoma (MM), where MA are present as overexpressed tumor-associated antigens. Consistent with this notion we report here that functional T cell reactivity to MA was found to be significantly diminished to MAGE-A3, Melan-A/Mart-1, and gp100 in untreated MM patients. Three lines of evidence suggest that the MA-reactive T cells present in healthy subjects undergo exhaustion once MM establishes itself. First, only the MA-specific T cell reactivity was affected in the MM patients; that to third party recall antigens was not. Second, in these patients, the residual MA-specific T cells, unlike third party antigen reactive T cells, were functionally impaired, showing a diminished per cell IFN-γ productivity. Third, we show that immunization with MA restored natural CD8+ T cell autoimmunity to MA in 85% of the MM patients. The role of natural T cell autoimmunity to tumor-associated MA is discussed based on discrete levels of T cell activation thresholds.

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Stockman, J. A. "gp100 Peptide Vaccine and Interleukin-2 in Patients with Advanced Melanoma." Yearbook of Pediatrics 2013 (January 2013): 31–33. http://dx.doi.org/10.1016/j.yped.2012.03.042.

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Eberlein, T. J. "gp100 Peptide Vaccine and Interleukin-2 in Patients with Advanced Melanoma." Yearbook of Surgery 2012 (January 2012): 350–52. http://dx.doi.org/10.1016/j.ysur.2011.09.022.

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Schwartzentruber, Douglas J., David H. Lawson, Jon M. Richards, Robert M. Conry, Donald M. Miller, Jonathan Treisman, Fawaz Gailani, et al. "gp100 Peptide Vaccine and Interleukin-2 in Patients with Advanced Melanoma." New England Journal of Medicine 364, no. 22 (June 2, 2011): 2119–27. http://dx.doi.org/10.1056/nejmoa1012863.

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Chan, Chi-Chao, Ying Li, Bing Sun, Qian Li, Dawn M. Matteson, De Fen Shen, Robert B. Nussenblatt, and Yifan Zhai. "Recombinant Adenovirus Encoding gp100 Modulates Experimental Melanin-Protein Induced Uveitis (EMIU)." Journal of Autoimmunity 11, no. 2 (April 1998): 111–18. http://dx.doi.org/10.1006/jaut.1997.0187.

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Linette, Gerald P., Michelle Becker-Hapak, Alexander Huang, Amer Alyasiry, Megan Chan, Wen-rong Lie, Lynn Cornelius, et al. "CD40 ligand/interferon-γ matured DC immunization with gp100 antigen HLA class I A *0201 restricted peptides in patients with newly diagnosed metastatic melanoma." Journal of Clinical Oncology 30, no. 15_suppl (May 20, 2012): 2525. http://dx.doi.org/10.1200/jco.2012.30.15_suppl.2525.

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2525 Background: CD40L/IFN-γ matured Dendritic Cells (DCs) produce IL-12 and are potent antigen-presenting cells for naïve resting T cells. We sought to determine the magnitude and kinetics of CD8+ T cell growth in patients receiving autologous CD40L/IFN-γ matured DC and identify biomarkers associated with clinical outcome. Methods: A phase I clinical trial (NCT00683670) incorporating CD40L/IFN-γ for the ex vivo maturation of autologous DCs pulsed with three well characterized gp100 melanoma antigen derived peptides (G154, G209-2M, G280-9V) was initiated with enrollment from 2008-11 at a single center. HLA-A*0201+ individuals with treatment naïve metastatic melanoma were immunized every 3 weeks by intravenous infusion for six doses after a single dose of cyclophosphamide (300 mg/m2 iv). CT imaging was performed at baseline, week 9 and 18 for clinical assessment using RECIST. Responding patients were eligible for maintenance doses every 2-4 months. PBMC were taken weekly for immune monitoring by tetramer analysis and functional assays. DC preparations were characterized to assess for biomarkers of response. Results: 10 patients were screened. Among the 7 treated patients, there were 3 confirmed responses (independently verified), including one durable CR >3 years and 2 PR. Three patients had rapid disease progression and received only 3 doses. Four patients (1 CR, 2 PR, 1 PD) received 6 or more vaccine doses. No SAEs were noted. There was no correlation between tumor volume and response. Using pre-specified immune response criteria, 6 (86%) treated patients developed CD8+ T cell immunity to all three peptides as assessed by tetramer analysis. The vaccine-induced T cells from all 6 individuals were polyfunctional and killed gp100+, HLA-A2+ human melanoma targets in a standard 51Cr release assay. IL-12 production by DCs correlated with TTP (p=0.0198, likelihood ratio test) but not OS (p=0.08). Conclusions: Weekly immune monitoring reveals the rapid onset of CD8+ T cell immunity against gp100 among the responder patients. This is the first DC vaccine clinical trial in melanoma to demonstrate a correlation of IL-12 production and TTP.

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Georgiev, Ivelin S., M. Gordon Joyce, Yongping Yang, Mallika Sastry, Baoshan Zhang, Ulrich Baxa, Rita E. Chen, et al. "Single-Chain Soluble BG505.SOSIP gp140 Trimers as Structural and Antigenic Mimics of Mature Closed HIV-1 Env." Journal of Virology 89, no. 10 (March 4, 2015): 5318–29. http://dx.doi.org/10.1128/jvi.03451-14.

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ABSTRACTSimilar to other type I fusion machines, the HIV-1 envelope glycoprotein (Env) requires proteolytic activation; specifically, cleavage of a gp160 precursor into gp120 and gp41 subunits creates an N-terminal gp41 fusion peptide and permits folding from an immature uncleaved state to a mature closed state. While the atomic-level consequences of cleavage for HIV-1 Env are still being determined, the uncleaved state is antigenically distinct from the mature closed state, and cleavage has been reported to be essential for mimicry of the mature viral spike by soluble versions of Env. Here we report the redesign of a current state-of-the-art soluble Env mimic, BG505.SOSIP, to make it cleavage independent. Specifically, we replaced the furin cleavage site between gp120 and gp41 with Gly-Ser linkers of various lengths. The resultant linked gp120-gp41 constructs, termed single-chain gp140 (sc-gp140), exhibited different levels of structural and antigenic mimicry of the parent cleaved BG505.SOSIP. When constructs were subjected to negative selection to remove subspecies recognized by poorly neutralizing antibodies, trimers of high antigenic mimicry of BG505.SOSIP could be obtained; negative-stain electron microscopy indicated these to resemble the mature closed state. Higher proportions of BG505.SOSIP-trimer mimicry were observed in sc-gp140s with linkers of 6 or more residues, with a linker length of 15 residues exhibiting especially promising traits. Overall, flexible linkages between gp120 and gp41 in BG505.SOSIP can thus substitute for cleavage, and sc-gp140s that closely mimicked the vaccine-preferred mature closed state of Env could be obtained.IMPORTANCEThe trimeric HIV-1 envelope glycoprotein (Env) is the sole target of virus-directed neutralizing antibody responses and a primary focus of vaccine design. Soluble mimics of Env have proven challenging to obtain and have been thought to require proteolytic cleavage into two-component subunits, gp120 and gp41, to achieve structural and antigenic mimicry of mature Env spikes on virions. Here we show that replacement of the cleavage site between gp120 and gp41 in a lead soluble gp140 construct, BG505.SOSIP, with flexible linkers can result in molecules that do not require cleavage to fold efficiently into the mature closed state. Our results provide insights into the impact of cleavage on HIV-1 Env folding. In some contexts such as genetic immunization, optimized cleavage-independent soluble gp140 constructs may have utility over the parental BG505.SOSIP, as they would not require furin cleavage to achieve mimicry of mature Env spikes on virions.

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Yang, Xinzhen, Michael Farzan, Richard Wyatt, and Joseph Sodroski. "Characterization of Stable, Soluble Trimers Containing Complete Ectodomains of Human Immunodeficiency Virus Type 1 Envelope Glycoproteins." Journal of Virology 74, no. 12 (June 15, 2000): 5716–25. http://dx.doi.org/10.1128/jvi.74.12.5716-5725.2000.

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ABSTRACT The human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins function as a membrane-anchored trimer of three gp120 exterior glycoproteins and three gp41 transmembrane glycoproteins. Previously, we reported three approaches to stabilize soluble trimers containing parts of the gp41 ectodomains: addition of GCN4 trimeric helices, disruption of the cleavage site between gp120 and gp41, and introduction of cysteines in the gp41 coiled coil to form intersubunit disulfide bonds. Here, we applied similar approaches to stabilize soluble gp140 trimers including the complete gp120 and gp41 ectodomains. A combination of fusion with the GCN4 trimeric sequences and disruption of the gp120-gp41 cleavage site resulted in relatively homogeneous gp140 trimers with exceptional stability. The gp120 epitopes recognized by neutralizing antibodies are intact and exposed on these gp140 trimers. By contrast, the nonneutralizing antibody epitopes on the gp120 subunits of the soluble trimers are relatively occluded compared with those on monomeric gp120 preparations. This antigenic similarity to the functional HIV-1 envelope glycoproteins and the presence of the complete gp41 ectodomain should make the soluble gp140 trimers useful tools for structural and immunologic studies.

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Stell, A., J. Dobson, and B. Catchpole. "Identification of a Splice Variant of gp100 Expressed in Canine Melanoma Tumours." Veterinary and Comparative Oncology 3, no. 1 (March 22, 2005): 57. http://dx.doi.org/10.1111/j.1476-5810.2005.064al.x.

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Luyten, G. P. M., C. W. van der Spek, K. Sintnicolaas, I. de Waard-Siebinga, M. J. Jager, P. T. V. M. de Jong, P. I. Schrier, and T. M. Luider. "Expression of MAGE, gp100 and tyrosinase genes in uveal melanoma cell lines." Melanoma Research 8, no. 1 (February 1998): 11–12. http://dx.doi.org/10.1097/00008390-199802000-00003.

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Journal articles on the topic 'Gp100'

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