Dong, Haidong

Abstract Spontaneous antitumor immunity remains challenging to detect and monitor largely due to the absence of established biomarkers. We selected the integrin CD11a as a biomarker to define antitumor T cell responses. CD11a is up-regulated on effector and memory T cells and is involved in the killing of tumor cells by cytotoxic T lymphocytes. In primary (subcutaneously inoculated) and metastatic (intravenously injected) tumors, CD11a high CD8 T cells were detected in the primary (skin) or metastatic (lung) tumor sites, but not in secondary lymphoid organs. Once tumors were established, CD11a high CD8 T cells were identified in tumor draining lymph nodes and peripheral blood. Phenotype analysis revealed that CD11a high CD8 T cells were proliferating (Ki67+) activated T cells (CD62L low, CD69 high, PD-1 high). CD11a high CD8 T cells expressed the transcription factor T-bet but not Foxp3. Live, but not dead, tumor cells, induced expansion of CD11a high CD8 T cells in the lung, suggesting that the access of tumor cells to target tissues is required to induce CD11a high CD8 T cell responses. Our results suggest that CD11a may provide a new T cell biomarker to measure ongoing natural tumor-reactive T cell responses and to predicate the efficacy of tumor immunotherapy.

This webinar is the first of a new series that brings together international experts to discuss ongoing developments in cancer research. This webinar focusses on emerging therapeutics, with speakers covering areas including immunotherapy, metabolic targets, and the treatment of lung and colorectal cancers. Our panellists discuss these developments, their ongoing challenges, and their global perspectives.

Abstract Introduction: Pediatric rhabdomyosarcoma (RMS) is the most common soft tissue tumor in children, with nearly 20% of children presenting with locally aggressive and/or metastatic disease. Despite aggressive chemotherapy and radiotherapy, the 5-year overall survival rate is still less than 40%. In addition, resistance to chemotherapy remains the major cause of cancer-related mortality. Therefore, there is an unmet need to develop more effective and tolerable therapeutic strategies for the treatment of RMS. Our overall objective is to leverage transcriptomic and proteomic profiling methods to identify RMS-specific surface proteins targetable with immune-based therapies. Methods: To identify therapeutic targets, we performed cell-surface capture and mass spectrometry on fusion-positive (FP), fusion-negative (FN) RMS and normal skeletal muscle. Transcriptomic datasets of RMS tumor specimens and normal tissue were employed to identify high-confidence RMS-specific cell-surface proteins. Expression of immune checkpoint molecules was confirmed by flow cytometry, western blot and PCR. Paraffin-embedded tissue sections were used to determine protein expression and prognostic value in RMS patients. We developed an in vitro T-cell killing assay to assess the functional role of immune checkpoint molecules on the antitumor immune response. Results: Surfaceome profiling of RMS cells revealed unique protein signatures specific for each tumor type and normal tissue. We have identified 5061 cell-surface proteins in RMS and 31.6% of proteins were correctly located at the plasma membrane. By integrating proteomic and transcriptomic data, we have identified 30 and 38 proteins significantly overexpressed in fusion-negative and fusion-positive RMS respectively compared to normal muscle. We have identified the immune checkpoint molecule B7-H3 as cell-surface protein overexpressed in both RMS subtypes. Strong expression of B7-H3 was also observed in patient’s tumor specimens. Genetic deletion of B7-H3 gene in RMS cells was associated with higher antitumor immune response in tumor cells co-cultured with CD8+-T cells. No difference was observed in tumor cell proliferation and radioresistance. Conclusions: Through integrated transcriptomic and proteomic profiling, we have discovered B7-H3 as a key regulator of tumor immune-evasion in RMS. B7-H3 is a suitable immunotherapy target for the treatment of RMS. Further studies are warranted to elucidate underlying mechanisms of B7-H3 function in RMS. Citation Format: Roxane R. Lavoie, Fabrice Lucien, Patricio C. Gargollo, Mohamed E. Ahmed, Yohan Kim, Emily Baer, Doris Phelps, Cristine M. Charlesworth, Benjamin J. Madden, Liguo Wang, Peter J. Hougthon, John Cheville, Haidong Dong, Candace F. Granberg. Surfaceome profiling of rhabdomyosarcoma reveals B7-H3 as a mediator of immune evasion [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr A036.

Summary Exploratory analysis of a phase III trial in esophageal cancer found that the patients who most contributed to an overall survival benefit from PD-1 blockade were not responders, but non-responders. The analysis has limitations but may have implications for investigating the optimal timing of immunotherapy relative to other treatments. See related article by Okada et al., p. 3277

Abstract Contemporary approaches for vaccination and immunotherapy are often capable of eliciting strong T-cell responses against tumor antigens. However, such responses are not parallel to clinical tumor regression. The development of evasion mechanisms within tumor microenvironment may be responsible for poor therapeutic responses. We report here that constitutive or inducible expression of B7-H1, a B7 family molecule widely expressed by cancers, confers resistance to therapeutic anti-CD137 antibody in mice with established tumors. The resistance is accompanied with failure of antigen-specific CD8+ CTLs to destroy tumor cells without impairment of CTL function. Blockade of B7-H1 or PD-1 by specific monoclonal antibodies could reverse this resistance and profoundly enhance therapeutic efficacy. Our findings support that B7-H1/PD-1 forms a molecular shield to prevent destruction by CTLs and implicate new approaches for immunotherapy of human cancers.

Abstract Background and Aims: Tumor necrosis factor related apoptosis inducing ligand (TRAIL) is predominantly expressed on immune cells. Although TRAIL biology has garnered considerable attention as a potential anti-cancer strategy, TRAIL agonists have had very limited anti-cancer activity in humans. TRAIL signaling can attenuate the T lymphocyte response. However, this potential immunosuppressive function of TRAIL has not been examined in cancer biology. Cholangiocarcinoma (CCA), a highly lethal biliary tract cancer, has a dense immunosuppressive microenvironment. Accordingly, CCA provides a model to examine the potential immune regulatory function of TRAIL in cancer biology. Methods: Using a syngeneic, orthotopic murine model of CCA (PMID: 29464042), murine CCA cells (SB cells) that express both Trail and Trail receptor (Tr) were implanted into livers of WT and Tr-/- mice. Hence, in this model the host immune cells express Trail but not the receptor; therefore, they would be capable of inducing TRAIL-mediated apoptosis in CCA cells but would be resistant to TRAIL-mediated immunosuppression. After 4 weeks of tumor growth, mice were sacrificed, and tumors were characterized using flow cytometry. Results: We observed that Tr-/- mice had a significant reduction in tumor burden compared to WT mice. Moreover, tumor bearing Tr-/- mice had a significant increase in cytotoxic T lymphocytes (CTLs) and enhanced CTL effector function. Myeloid-derived suppressor cells (MDSCs) were significantly decreased in Tr-/- tumors compared to WT tumors. Furthermore, implantation of SB cells devoid of Trail (SB-Trail-/-) into WT mice resulted in a significant reduction in tumor burden and MDSC infiltration. In vitro functional studies employing SB cells deficient in Tr (SB-Tr-/-) and MDSCs were carried out. There was a significant reduction in proliferation and immunosuppressive function of MDSCs when cocultured with SB-Tr-/- compared to SB cells. These results indicate that TRAIL-TR fosters MDSC growth and immunosuppressive function. In conclusion, we have demonstrated that Tr-/- mice have a significant reduction in CCA tumor burden and MDSC infiltration. Consequently, Tr-/- tumors have enhanced CTL infiltration and function. These data suggest that the TRAIL-TR system mediates tumors immune evasion via MDSCs, and direct targeting of TRAIL on CCA cells is a potential anti-tumor strategy. Citation Format: Emilien Loeuillard, Jingchun Yang, Haidong Dong, Gregory J. Gores, Sumera I. Ilyas. Tumor necrosis factor related apoptosis inducing ligand fosters myeloid derived suppressor cell mediated tumor immune evasion in cholangiocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1315.

Abstract Seven different anti-PD-1 and PD-L1 monoclonal antibodies are now widely used in the US to treat a variety of cancer types, but there have been no clinical trials comparing them directly. Furthermore, because many of these antibodies do not cross react between mouse and human proteins, no preclinical models exist in which to consider these types of questions. Thus, we produced humanized PD-1 and PD-L1 mice in which the extracellular domains of both mouse PD-1 and PD-L1 were replaced with the corresponding human sequences. Using this new model, we sought to compare the strength of the immune response generated by FDA-approved antibodies. To do this, we performed an in vivo T cell priming assay in which anti-PD-1/L1 therapies were given at the time of T cell priming against surrogate tumor antigen (OVA), followed by subsequent B16-OVA tumor challenge. We found that anti-PD-1/L1-treated mice exhibited significantly better tumor rejection than controls, although both the control and antibody-treated mice generated comparable numbers of OVA-specific T cells at the time of priming. To determine what could mediate this strong antitumor immune response, we identified the increased production of CX3CR1+PD-1+CD8+ cytotoxic T cells in the anti-PD-1/L1-treated mice, the same subset of effector T cells known to increase in clinical responders to PD-1/L1 therapy. Thus, our model permits the direct comparison of FDA-approved anti-PD-1/L1 monoclonal antibodies and further correlates successful tumor rejection with the level of CX3CR1+PD-1+CD8 + T cells, making this model a critical tool for future studies to optimize and better utilize anti-PD-1/L1 therapeutics.

TPS4170 Background: IMbrave150 is the first study demonstrating the benefit of anti-PDL1 in the frontline treatment of aHCC, and established Atezo/Bev as a new 1st line standard for aHCC. There is currently limited evidence to guide subsequent therapy for aHCC patients progressing on Atezo/Bev. ACCRU-GI-2008 is designed to determine the benefit of continuing Atezo into 2nd line and the safety of Atezo plus a MKI in patients with aHCC who previously received Atezo/Bev. The study is being conducted across 12 centers in the United States (ClincalTrials.gov#: NCT05168163). Methods: This study utilizes a 2:1 randomized phase II design where eligible patients will receive either Atezo/MKI (experimental arm) or MKI alone (control). Patients will be stratified according to the MKI choice (cabozantinib or lenvatinib, per physician’s decision), etiology of HCC (viral vs. non-viral) and alpha-fetoprotein level ( < 400 vs. > = 400 ng/mL). The major eligibility criteria are histological/cytological diagnosis or clinical diagnosis of HCC per the AASLD or WASL 2018 guidelines, has advanced disease not amendable to curative treatment, previously received and progressed on Atezo/Bev, has received only 1 previous line of systemic therapy (2nd line only), ECOG PS 0-1, Child Pugh Class A, adequate organ reserves and RECIST v1.1 measurable disease; previous MKI for advanced disease is excluded. The primary endpoints are overall survival (OS) and progression free survival (PFS). A total sample size of 122, with 89 PFS events, we will have 80% power to detect an improvement in median PFS from 4 to 7 months, assuming a one-sided significance level of 0.05. With approximately 84 deaths, we will have 80% power to detect an improvement in median OS from 10 to 18 months, assuming a one-sided significance level of 0.05. The overall one-sided significance level, for the study, is 0.1. An OS interim analysis will be conducted at 89 PFS events. Secondary endpoints include objective response, duration of response, and adverse events. Archival tumor and serial blood samples will be collected to evaluate for potential prediction biomarkers and mechanisms of sensitivity/resistance. Baseline and on-treatment tumor biopsy specimens will also be collected from the initial 10 patients of each arm. The study is approved by the ethics committee and enrollment to the study will be underway by Q2/3 2022. Clinical trial information: NCT05168163.