bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3717: Sudocetaxel Zendusortide (TH1902) triggers the cGAS/STING pathway and potentiates anti-PD-L1 immune-mediated tumor cell killing
Authors: Demeule Michel , Currie Jean-Christophe , Charfi Cyndia , Zgheib Alain , Cousineau Isabelle , Lullier Veronique , Beliveau Richard , Marsolais Christian , Annabi Borhane
Published: March 2024

The potential impact of the immune tumor microenvironment (iTME) was assessed with respect to the anticancer efficacy of sudocetaxel zendusortide (TH1902), a peptide-drug conjugate internalized through a sortilin (SORT1)-mediated endocytic pathway. In the triple-negative breast cancer (TNBC)-derived MDA-MB-231 immunocompromised xenograft tumor model, TH1902 induced complete tumor regression for more than 40 days after the last treatment. Surprisingly, immunohistochemistry analysis revealed higher staining of CD45 leukocytes and of STING, a master regulator in the cancer-immunity cycle, in comparison to docetaxel. Additionally, in a murine B16-F10 melanoma syngeneic tumor model, weekly administration of TH1902 as a single agent also demonstrated superior tumor growth inhibition than did docetaxel. Although B16-F10 is considered a non-immunogenic “cold” tumor model, we observed a net increase in CD45 leukocyte infiltration within TH1902-treated tumors, especially for tumor-infiltrating lymphocytes and tumor-associated macrophages. Moreover, in TH1902-treated tumors, increased staining of perforin, granzyme B, and caspase-3 was suggestive of elevated cytotoxic T and natural killer cell activities. Combined TH1902/anti-PD-L1 treatment led to increased tumor growth inhibition and median animal survival. Accordingly, TH1902 inhibited cell proliferation and triggered apoptosis and senescence in B16-F10 cells in vitro, while inducing several downstream effectors of the cGAS/STING pathway and the cell surface expression of MHC-I and PD-L1. This is the first evidence that TH1902 exerts its antitumor activity, in part, through modulation of the iTME and that the combination of TH1902 with checkpoint inhibitors (anti-PD-L1) could lead to improved clinical outcomes.

Citation Format: Michel Demeule, Jean-Christophe Currie, Cyndia Charfi, Alain Zgheib, Isabelle Cousineau, Veronique Lullier, Richard Beliveau, Christian Marsolais, Borhane Annabi. Sudocetaxel Zendusortide (TH1902) triggers the cGAS/STING pathway and potentiates anti-PD-L1 immune-mediated tumor cell killing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3717.
Abstract 372: Cell polarity proteins as novel regulators of macropinocytosis in pancreatic cancer
Authors: Barjau Guillem Lambies , Lee Szu-Wei , Commisso Cosimo
Published: March 2024

Pancreatic ductal adenocarcinoma (PDAC) is considered one of the deadliest cancers. This is likely because it is detected at a very late stage and the therapeutic approaches that have been used for its treatment have proven to be quite ineffective. Therefore, new therapeutic strategies are needed for a better prognosis in patients suffering of pancreatic cancer. PDAC cells often rewire their metabolism to survive and progress. This metabolic rewiring permits the acquisition of nutrients by the cells when these are exposed to a poor nutrient microenvironment. One of the most used processes in nutrient stress conditions is the “cell-drinking” pathway macropinocytosis, an endocytic pathway that cancer cells exploit to support cancer metabolism in such unfavorable conditions. Macropinocytosis inhibition has shown to suppress PDAC growth in mice and thus, it has become a promising therapeutic target for PDAC treatment. Here, we describe that atypical Protein Kinase C (aPKC) isoforms Protein Kinase C zeta (PKCζ) and Protein Kinase C iota (PKCι) regulate PDAC macropinocytosis in the context of glutamine stress. Genetic depletion and pharmacological inhibition of aPKC impair macropinocytosis promotion in PDAC cells exposed to a glutamine-deprived environment and in cells treated with DON, a glutamine analog that blocks glutamine metabolism. In epithelial cells, aPKCs are known to regulate the establishment of cell polarity in association with the scaffold proteins Par3 and Par6, constituting the cell polarity complex, which controls the function of different targets such as Par1 kinases. Loss of Par3, Par6 and Par-1a affects glutamine stress associated macropinocytosis in a similar extent than aPKC depletion, presenting a novel role of the cell polarity protein network in the modulation of macropinocytic uptake.

Citation Format: Guillem Lambies Barjau, Szu-Wei Lee, Cosimo Commisso. Cell polarity proteins as novel regulators of macropinocytosis in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 372.
Abstract 3791: A prognostic signature based on inherited genetic polymorphisms for patients with uveal melanoma
Authors: Verrier Thibault , Ven Anaïs Le , Houy Alexandre , Garcia Agathe , Gaelle Pierron , Cassoux Nathalie , Rodrigues Manuel , Noirel Josselin , Stern Marc-Henri
Published: March 2024

Introduction: Uveal melanoma (UM) is the most frequent eye malignancy in adults. Two UM subtypes exist, defined by Gene Expression Profiling or by genomic status: GEP class 1/disomy 3 [D3] and GEP class 2/monosomy 3 [M3] defining low-risk or high-risk of developing metastasis, respectively. However, these prognostic signatures require tumor samples, which may be difficult to obtain. We previously identified by a genome-wide association study (GWAS) frequent single nucleotide polymorphisms (SNP) in IRF4 and HERC2 associated with risk for D3 or M3 UM, respectively (1). Here, we aim to design a prognostic signature for the D3 and M3 UM subtypes based on these constitutive polymorphisms, available from blood samples.

Experimental Procedures: We used the GWAS cohort of 1476 UM patients genotyped on GSA or Omni5. Risk SNPs coupled with clinical variables were used on a cohort of 497 UM patients (191 D3, 306 M3) in a statistical machine learning process to define a predictive signature of UM subtype. A cohort of 852 UM patients was used for validation.

Results: We retained two SNPs, rs12203592 (in IRF4) and rs12913832 (in HERC2), tumor diameter and thickness, age at diagnosis and gender. We validated the predictive model in the validation cohort. The predicted high-risk group showed a decrease in time to relapse (TR) (HR: 3.72 [2.57 - 5.41], logrank p-value = 1.35E-15), overall survival (OS) (HR: 3.85 [2.58 - 5.73], logrank p-value = 1.24E-14) and progression-free-survival (PFS) (HR: 3.61 [2.32 - 5.61], logrank p-value = 6.55E-11), compared with the predicted low-risk group.

Conclusion: Common germline polymorphisms contribute to the prediction of metastatic risk in patients with localized uveal melanoma. This information could be especially useful when access to the tumor is problematic.

1. Mobuchon et al, J Natl Cancer Inst, 114, 2, (2022)

Citation Format: Thibault Verrier, Anaïs Le Ven, Alexandre Houy, Agathe Garcia, Pierron Gaelle, Nathalie Cassoux, Manuel Rodrigues, Josselin Noirel, Marc-Henri Stern. A prognostic signature based on inherited genetic polymorphisms for patients with uveal melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3791.
Abstract 3798: Advancements in multiplexed spatial phenotyping
Authors: Montoya Leticia , Clarke Scott , Harvey Austin , Serafin Jen , Voeun Mae
Published: March 2024

Spatial omics is an expanding research area focused on integrating spatial knowledge of tissue with transcriptomics (RNA) and proteomics (protein). Understanding the complexity of the biological structure(s) is an important biological process in cancer immunotherapy research which requires accurate target classification. Translational profiling of 4+ targets on a single sample at one time can be difficult in many ways, such as, panel design, staining protocols, and data analysis. Furthermore, to design a reliable multi-target biomarker panel, there are many parts to overcome, such as protein abundance and localization, fluorophore compatibility, varying tissue types, and data characterization, to name a few. To address the needs of high target multiplex ability, labeling methods have advanced in cyclic detection enabling iterative labeling and detection using automated fluidics, however these approaches suffer from low simultaneous target throughput. Thermo Fisher Scientific can now provide a streamline process for tissue labelling; a process that enables sufficient labelling of high multiplexed panels completed within a couple of hours. We show successful detection of a wide range of various protein markers across numerous tissue organs used in various biological research applications with our uniform technique of labelling.

Citation Format: Leticia Montoya, Scott Clarke, Austin Harvey, Jen Serafin, Mae Voeun. Advancements in multiplexed spatial phenotyping [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3798.
Abstract 3842: Neuroendocrine differentiation and treatment of human SCLC: NRF2 has a role
Authors: Hamad Samera , Joshi Hansa , Li Yahui , Toma Helen , Bendjilali Nasrine , Jefferys Stuart , Corcoran David , Elbezanti Weam , Major Ben , Bernard Weissman , Spitz Francis
Published: March 2024

SCLC is a lethal form of lung cancer and highly metastatic disease with limited therapeutic options. Four main neuro and non-neuroendocrine SCLC molecular subtypes have been identified which further complicate the efforts of finding targeted therapies. NRF2 is a transcription factor that is known to protect cells from oxidative damage by regulating more than 500 cytoprotective genes. NRF2 is frequently mutated in non-small cell lung cancer (NSCLC), however, whether alterations occur in small cell lung cancer (SCLC) is not clear due to limited sequencing data. Recently, our group has discovered a previously unrecognized role of NRF2 signaling pathway in the development of SCLC using a novel genetically engineered mouse model (GEMM). In this study, we have investigated the role of NRF2 in the progression and regression of human SCLC. Human SCLC cell lines were generated with NRF2E79Q/+ mutation (one of the most common mutations found in human tumors) to allow the constitutive activation of NRF2 in these cells. Results from hSCLC cell lines showed that NRF2 activation in NRF2-negative SCLCs can significantly inhibit their growth. Further, human SCLC gene expression data showed reduced levels of a NRF2 signature as hSCLC neuroendocrine SCLC transforms to non-neuroendocrine subtypes. As an anti-inflammatory pathway, NRF2 is known to inhibit immune cell gene signature. Our in vivo and invitro studies showed that NRF2 inhibits immune cell infiltration in SCLC; and increases PDL1 protein expression. Currently, we are investigating the effect of NRF2 activation in hSCLC growth, neuroendocrine differentiation and immune gene activation in vitro and in vivo. We are also using immune checkpoint inhibitors to determine the best therapeutic options for each SCLC subtype.

Citation Format: Samera Hamad, Hansa Joshi, Yahui Li, Helen Toma, Nasrine Bendjilali, Stuart Jefferys, David Corcoran, Weam Elbezanti, Ben Major, Weissman Bernard, Francis Spitz. Neuroendocrine differentiation and treatment of human SCLC: NRF2 has a role [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3842.
Abstract 3851: Beta-lapachone promotes tumor associated neutrophils (TANs) polarized towards anti-tumor (N1) phenotype
Authors: Huang Xiumei , Tumbath Soumya , Jiang Liangxiang , Fu Yang-Xin
Published: March 2024

NAD(P)H:quinone oxidoreductase1 (NQO1) is overexpressed in most solid cancers, emerging as a promising target for tumor selective killing. β-lapachone (β-lap), an NQO1bioactivatable drug, exhibits significant antitumor effects on NQO1+ human solid cancer cell lines. It also induces immunogenic cell death in the tumor microenvironment (TME)and synergizes with anti-PD-L1 immunotherapy. However, the interaction between β-lap-mediated anti-tumor immune response and neutrophils, a novel antigen presenting cells(APCs), remains unknown. This study found β-lap treatment increased intracellular ROS formation and DNA damage to kill NQO1+ murine tumor cell lines and dicoumarol spared the lethality. Furthermore, flow cytometric analysis of immune cells in TME of subcutaneous tumor models indicated that there is a significant infiltration of neutrophils(CD11b+ Gr1high cells) or Tumor Associated Neutrophils (TANs) following β-lap treatment. Depletion of neutrophils by co-addition of anti-Ly6G antibody abolished β-lap-induced anti-tumor efficacy in immune competent mice. Neutrophil mediated tumor cell killing determined ex vivo using bone marrow derived neutrophils and it was observed that in the presence of β-lap induced antigen, neutrophils were killing murine tumor cells. Moreover, exposure of these neutrophils to β-lap induced antigen medium induce antitumor N1-like properties, include increase in certain surface markers such as CD95(Fas) and CD54 (ICAM-1) as well as increase in intracellular cytokine IFN-β and decrease in TGF-β revealed by flow cytometric analysis. Corroborating to this, there is a significant increase of CD95 and IFN-β and a decrease in TGF-β were observed with TANs infiltrated into β-lap treated TME compared to vehicle treated tumors. Additionally, TLR/MyD88signaling deficiency decreased β-lap induced neutrophil tumor infiltration. BlockingHMGB1, depleting neutrophils, or TLR4/MyD88 deficiency lessened the antigen-specific T cell response. This study highlights that β-lap kills NQO1+ murine tumor cells through ROS production and DNA damage, and neutrophils play a pivotal role in its antitumor effects.

Citation Format: Xiumei Huang, Soumya Tumbath, Liangxiang Jiang, Yang-Xin Fu. Beta-lapachone promotes tumor associated neutrophils (TANs) polarized towards anti-tumor (N1) phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3851.
Abstract 3863: Flow cytometry using stabilized whole blood to assess innate and adaptive immune cells (Monocytes, NK, T, and B cells) in immunotherapies
Authors: Benko Jackie , Splitt Samantha , Bisconte Angelina , Lozza Laura , Owen Rachel , Phippard Deborah
Published: March 2024

Flow cytometry assays using fresh whole blood which are interrogating either an extremely rare cell subset or a depleting immunotherapy, may require samples to be processed from global clinical sites over a wide geographical area, which can pose a major challenge for clinical operations. Precision for Medicine (PfM) has addressed these operational challenges using commercial stabilizers which are added to the blood allowing samples to be frozen at -80°C and subsequently batch processed for sample testing. We have performed a fit for purpose assay qualification of a multiparameter flow cytometry panel allowing the immunophenotypic analysis of monocyte, T cell, B cell and NK immune populations using Cytodelics™ fixed/stabilized healthy donor whole blood. Additionally, the assay was verified in whole blood samples from multiple solid tumor disease states including melanoma, colorectal cancer, head & neck, non-small cell lung cancer to demonstrate clinical application. In these oncology samples, we were able to assess both immune cell frequencies and their activation status in both fresh whole blood as well as fixed whole blood over a short-term sample stability study, where the samples had been stored at -80°C for several months. This data provides a representative example of the use of commercial stabilizing fixatives for clinical studies to extend the stability window of whole blood for subsequent assessment by flow cytometry. Fixed whole blood can be stored at -80°C, shipped on dry ice allowing multiple timepoints in a cohort to be assessed together, thus minimizing assay variability as compared to fresh whole blood analysis, and mitigating the impact of shipping delays and temperature excursions on the samples. With sufficient time for assay development there are several commercial stabilizers that can be tested to determine if the markers in the flow assay are compatible with these reagents, providing a valuable option to assess the impact of immunotherapies on the immune cells of interest and reducing operational challenges in global studies.

Citation Format: Jackie Benko, Samantha Splitt, Angelina Bisconte, Laura Lozza, Rachel Owen, Deborah Phippard. Flow cytometry using stabilized whole blood to assess innate and adaptive immune cells (Monocytes, NK, T, and B cells) in immunotherapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3863.
Abstract 395: An alternating magnetic field suppresses tumor growth
Authors: Umemura Masanari , Akimoto Taisuke , Islam Rafikul , Kishi Kazuhito , Nagasako Akane , Nakakaji Rina , Yamaguchi Takashi , Mizuno Yuto , Ishikawa Soichiro , Yamamoto Tetsuya , Ishikawa Yoshihiro
Published: March 2024

Application of physical forces, ranging from ultrasound to electric field, is recommended in various clinical practice guidelines, including those for bone fracture or cancer treatment. However, mechanistic information for such treatments has been poorly provided, due to the lack of comprehensive study model. Here, we show an alternating magnetic field (AMF) per se exhibits direct anti-cancer effect through enhancing oxidative phosphorylation (OXPOHS) and thus metabolic reprograming. Proliferation of human glioblastoma cells (U87 and LN229), but not normal human astrocytes (NHA), was inhibited upon exposure to AMF within the narrow frequency range. Extracellular acidification rate was decreased while oxygen consumption was increased. These data indicated that AMF induced metabolic reprograming. This was accompanied by increased reactive oxygen species (ROS) production. Mouse models grafted with human glioblastoma cells showed that AMF exposure, 30 minutes per day for two weeks, suppressed the tumor growth and prolonged the overall survival. Our results suggest that the AMF exposure may be a simple strategy to inhibit cancer cell growth by utilizing oxidative stress via metabolic reprograming.

Citation Format: Masanari Umemura, Taisuke Akimoto, Rafikul Islam, Kazuhito Kishi, Akane Nagasako, Rina Nakakaji, Takashi Yamaguchi, Yuto Mizuno, Soichiro Ishikawa, Tetsuya Yamamoto, Yoshihiro Ishikawa. An alternating magnetic field suppresses tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 395.
Abstract 3951: Timing the development of chemoresistance in relapsed pediatric cancer
Authors: Blay Sasha , Layeghifard Mehdi , Davidson Scott , Chen David , Schwertschkow Astra , Ramaswamy Vijay , Taylor Michael , Papaemmanuil Elli , Villani Anita , Malkin David , Alexandrov Ludmil , Cowley Mark , Shlien Adam
Published: March 2024

Survivors of pediatric cancer face lifelong battles with severe morbidities, which includes a significant risk of recurrence. Pre-existing genetic variation within primary tumors may offer certain cell populations an evolutionary advantage, increasing the likelihood that some will be resistant to treatment. But the inherent complexity of the tumor means finding these markers of resistance has yet to be fully explored across childhood cancer. To address this, we developed a pipeline that combines clonal evolution reconstruction with mutational signature extraction at the subclonal level to elucidate changes in mutational processes. Using this pipeline to analyze 1,743 pediatric tumor genomes we found a common pattern of parallel evolution in most pediatric cancers, irrespective of their tissue of origin. Further, we detected therapy-associated mutational signatures linked to specific treatments such as temozolomide, platinum, fluorouracil, and thiopurine in post-treatment tumor subpopulations. Of 235 samples with known treatment exposure, 37.9% displayed one or more treatment-associated signatures. In the context of the phylogenetic tree, we identified specific subpopulations with therapy signatures. In multi-sample cases, resistant subpopulations had ancestors present in primary tumors at the time of diagnosis, suggesting certain subpopulations possessed the ability to withstand chemotherapy-induced pressures from an early stage and then expanded following treatment. Finally, we observed differences in the timing of mutational signatures between antimetabolite and alkylating agent treatments, reflecting the mechanisms of these drugs. Antimetabolite agents showed subclonal mutational signatures occurring in later branches, likely due to dependence on cell cycling. In contrast, alkylating agents displayed signatures in both early and late subpopulations, as they are unaffected by cell cycle state. Thus, investigating mutational signatures at the subclonal level unveils new insights into the clonal dynamics of pediatric cancers and the development of chemoresistance. This research offers valuable insights for identifying pediatric cancer patients at greater risk of recurrence, enabling the development of more effective treatment plans to enhance their quality of life and overall survival rates.

Citation Format: Sasha Blay, Mehdi Layeghifard, Scott Davidson, David Chen, Astra Schwertschkow, Vijay Ramaswamy, Michael Taylor, Elli Papaemmanuil, Anita Villani, David Malkin, Ludmil Alexandrov, Mark Cowley, Adam Shlien. Timing the development of chemoresistance in relapsed pediatric cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3951.
Abstract 3967: ISM9182A, a novel HPK1 inhibitor, exhibits immune modulatory activity and robust monotherapy anti-tumor effects in preclinical studies
Authors: Chen Celia X.-J. , Pu Congying , Peng Jingjing , Ding Xiaoyu , Lu Hongfu , Ding Xiao , Zhang Man , Bavadekar Supriya , Rao Sujata , Ren Feng , Zhavoronkov Alex
Published: March 2024

The hematopoietic progenitor kinase 1 (HPK1 or MAP4K1) is a member of the Ste20 serine/threonine kinase super family that negatively regulates immune cells, including T cells, B cells and dendritic cells (DCs). Thus HPK1 is an attractive immuno-oncology target in many tumor types. Here, we report ISM9182A as a novel, potent, and selective inhibitor of HPK1.ISM9182A demonstrated nanomolar enzymatic potency against HPK1 with >100-fold selectivity against other MAP4K family kinases. ISM9182A inhibited the phosphorylation of SLP-76 at Ser376 dose-dependently with an IC50 of 30nM in human peripheral blood mononuclear cells. Moreover, ISM9182A enhanced IL-2 and IFNγ secretion upon CD3/CD28 co-stimulation in a dose-dependent manner without affecting T cell proliferation. Treatment of human pan T cells with ISM9182A also significantly reversed PGE2-mediated immunosuppression of human pan T cell activation and further boosted IL-2 and IFNγ secretion compared to CD3/CD28 co-stimulation. In monocyte-derived DCs stimulated with TNF-α/IL-1β/PGE-2, ISM9182A significantly upregulated CD80, CD86, and CD83 expression, demonstrating its potency to promote DC maturation. ISM9182A showed no obvious toxicity towards human T cell, B cells, and monocytes without CD3/CD28 co-stimulation. In vivo PK data revealed favorable ADME properties and good oral bioavailability of ISM9182A across different species. In a CT-26 murine syngeneic mouse model, oral administration of ISM9182A exhibited potent antitumor effects with p-SLP76 inhibition in both murine spleen and tumor, without body weight loss. In summary, ISM9182A is a potent and selective HPK1 inhibitor and a promising immuno-oncology agent.

Citation Format: Celia X.-J. Chen, Congying Pu, Jingjing Peng, Xiaoyu Ding, Hongfu Lu, Xiao Ding, Man Zhang, Supriya Bavadekar, Sujata Rao, Feng Ren, Alex Zhavoronkov. ISM9182A, a novel HPK1 inhibitor, exhibits immune modulatory activity and robust monotherapy anti-tumor effects in preclinical studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3967.
Abstract 3978: B-cell activating factor (BAFF) enhances antitumor immunity
Authors: Aziz Khaled , Munjal Kabeer , Howe Kathryn , Yarchoan Mark
Published: March 2024

Immunotherapies that modulate T cell function have been firmly established in cancer immunotherapy, whereas the potential for B cells in the antitumor immune response is not as well understood. B cell-activating factor (BAFF) is a cytokine belonging to the TNF ligand family that activates B cells and is linked to autoimmunity. Our preliminary research has identified BAFF as a target for enhancing antitumor immunity through enhancing presentation of tumor neoantigens and inducing tertiary lymphoid structures which can orchestrate tumor-specific T cell immune responses and correlate with improved clinical response. Since broad activation of B cell response could be detrimental and possibly induce autoimmune side effects, identification of specific B cell receptors that coordinate anti-tumor immunity is prudent. BAFF has three known receptors (BAFF-R, TACI and BCMA), and the effects of these specific BAFF ligand-receptor interactions on antitumor immunity are not known. Here we first generated BAFF-overexpressing cell lines using genomic editing [KPC-BAFF (pancreas), B16F10-BAFF (melanoma), and PancO2-BAFF (pancreas)] and examined their tumor growth in syngeneic immunocompetent host. Secondly, we investigated the effects of BAFF-overexpression on tumor response when individual receptors are knocked out using BAFF-R-KO, TACI-KO and BCMA-KO mice. Our results show significantly hindered tumor growth when the BAFF expressing stable cell lines are introduced to immunocompetent host, compared to parental cell lines. Tumor response experiments in receptor knockout mice is currently in progress. Successful identification of the BAFF receptor candidate that drives anti-tumor response is an exciting prospect for clinical translation. Development of agonists for the specific receptor may allow selective modulation of this particular B cell function in context of cancer therapy without perturbation of broad systemic humoral immunity.

Citation Format: Khaled Aziz, Kabeer Munjal, Kathryn Howe, Mark Yarchoan. B-cell activating factor (BAFF) enhances antitumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3978.
Abstract 4004: Fine-tuning affinity/avidity of anti-CAIX CAR T cells to mitigate on-target off-tumor toxicity in clear cell renal cell carcinoma
Authors: Yuan Stephen Hsien-Chi , Wang Yufei , Abbas Rabia , Mathenge Peterson , Chien Charles , Coherd Christian D. , Bajoua Branson , Chang Matthew R. , Murugan Nithyassree , Kastrunes Gabriella M. , Wang Guilin , Hoang Song-My , Marasco Wayne A.
Published: March 2024

On-target off-tumor toxicity (OTOT) is one of the major hurdles preventing tumor-associated antigen (TAA) targeted therapy from effectively treating patients with solid tumors due to the shared presence of the same TAA on normal tissues. We have previously demonstrated high expression of carbonic anhydrase IX (CAIX) in ccRCC patient samples and low-density expression on healthy bile duct epithelium. In 2006, a clinical trial was stopped due to OTOT bile duct toxicity observed in patients infused with anti-CAIX CAR T cell therapy. To address the issue of OTOT in the bile duct, we performed affinity/avidity fine-tuning of the anti-CAIX CAR G36 to specifically target ccRCC tumor cells with high CAIX expression while sparing cholangiocytes with low CAIX expression. We generated a complementarity-determining region (CDR) single variant mutagenesis library of G36 and displayed the library on yeast for screening. Using flow cytometry, we were able to identify six populations based on antigen-binding affinity: high, med-high, med, med-low, low, and loss of binding. These libraries were sequenced via PacBio, a next generation sequencing (NGS) platform that excels in long-read sequencing, providing a powerful tool to detect single mutations in the scFv. The results were analyzed to identify enriched mutations compared to the wild-type G36, and inspected using in silico modeling and docking. Furthermore, enriched mutants from med-low and low populations were tested in CAR-T format for their cytotoxic capacity on tumor cells and bile duct cholangiocytes, as well as their avidity. We demonstrate that mutations in CDR2 and CDR3 of the heavy chain successfully mitigate OTOT toxicity on cholangiocytes with physiological levels of CAIX expression while maintaining tumor-killing capacity, shedding light on CDR mutagenesis as a useful tool to mitigate OTOT toxicity.

Citation Format: Stephen Hsien-Chi Yuan, Yufei Wang, Rabia Abbas, Peterson Mathenge, Charles Chien, Christian D. Coherd, Branson Bajoua, Matthew R. Chang, Nithyassree Murugan, Gabriella M. Kastrunes, Guilin Wang, Song-My Hoang, Wayne A. Marasco. Fine-tuning affinity/avidity of anti-CAIX CAR T cells to mitigate on-target off-tumor toxicity in clear cell renal cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4004.
Abstract 4006: IL12-secreting CAR-T cells eliminate the requirement for lymphodepletion and promote epitope spreading via local antigen presenting cells
Authors: Mohan Aditya , Shen Steven , Hotchkiss Kelly , Quackenbush Sarah , Sampson John , Fecci Peter , Patel Anoop
Published: March 2024

Introduction: EGFRvIII is a commonly studied tumor-specific antigen in glioblastoma (GBM) and poses an enticing target for immunotherapy. Its varied expression, however, complicates its effectiveness as a Chimeric Antigen Receptor (CAR) T cell target. Given the increasing prominence of Interleukin-12 (IL12) in treating solid tumors, its synergy with EGFRvIII specific CARs in treating preclinical models of GBM is explored.

Methods: To simulate tumor heterogeneity, mice were implanted with equal ratios of CT2A and CT2A EGFRvIII+ cells. We developed EGFRvIII-specific third-generation CAR T cells, both with and without IL12 using a MSGV retrovirus backbone and transduced them into murine T cells. Their antitumor effectiveness was gauged by evaluating survival rates, and their capability to enhance and modify the tumor environment was determined using single-cell sequencing.

Results: Standalone EGFRvIII CARs were unsuccessful in curing mice with heterogeneous tumors. However, 60% of mice treated with EGFRvIII CARs combined with IL12 were long term survivors. IL12 secreting CARs exhibited prolonged persistence in vivo, and there was no need for prior lymphodepletion before ALT introduction. Our studies reveal that EGFRvIII CARs secreting IL12 can eliminate EGFRvIII negative tumor cells by drawing on the inherent CD8 T cell compartment. Moreover, IL12 prominently impacted microglia among all Antigen Presenting Cells in the tumor environment, amplifying antigen presentation processes and broadening epitope exposure.

Conclusion: By leveraging IL12, EGFRvIII CARs can counteract antigen heterogeneity in tumors by recruiting endogenous CD8 T cells and polarizing microglia. These IL12-secreting CARs outlast their traditional counterparts without requiring lymphodepletion. Harnessing CAR-T cells that secrete IL12 represents a novel strategy for overcoming tumor heterogeneity and treating GBM in a clinical context.

Citation Format: Aditya Mohan, Steven Shen, Kelly Hotchkiss, Sarah Quackenbush, John Sampson, Peter Fecci, Anoop Patel. IL12-secreting CAR-T cells eliminate the requirement for lymphodepletion and promote epitope spreading via local antigen presenting cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4006.
Abstract 4013: Increasing mitochondrial respiration by silencing MCJ/DnaJC15 enhances CD8 CAR-T cell therapy efficacy
Authors: Wu Meng-Han , Giddings Emily L. , Valenca-Pereira Felipe , Pham-Danis Catherine , Cendali Francesca , Yarnell Michael C. , Novak Amanda J. , Henao-Mejia Jorge , Flavell Richard A. , D'Alessandro Angelo , Kohler Mark Eric , Rincon Mercedes
Published: March 2024

Chimeric antigen receptor (CAR)-T cell therapy is a promising treatment against leukemia and lymphoma. However, poor initial treatment response and post-therapy relapse have found to be common and remained to be resolved. Modulating T cell immunometabolism is considered a novel therapeutic approach to enhance CAR-T cell therapy. We have identified MCJ (DnaJC15) as an endogenous negative regulator of Complex I of the electron transport chain in mitochondria. MCJ is abundantly expressed in mouse CD8 cells relative to other immune cells. We propose that MCJ can be a therapeutic target to enhance mitochondrial respiration in CD8 CAR-T cells. We will show that loss of MCJ in mouse CD8 cells leads to superior anti-tumor response against melanoma. In addition, we will show that MCJ-deficient murine CD8 CAR-T cells have enhanced killing activity against leukemia both in vitro and in vivo. We have recently revealed that MCJ is also expressed in human CD8 cells and the levels of MCJ are heterogenous among human population. Similar to murine CD8 cells, MCJ knockdown in human CD8 cells enhances mitochondrial membrane potential and effector functions. We have designed a new CAR construct to silence MCJ in human CD8 CAR-T cells. MCJ-deficient human CD8 cells show superior cytotoxicity against leukemia with increased mitochondrial metabolism. Together, these data demonstrate that MCJ is a promising target to improve CAR-T cell therapy through modulating the T cell metabolism. We are currently investigating the effect of MCJ-deficient CD8 CAR-T cells against solid malignancy.

Citation Format: Meng-Han Wu, Emily L. Giddings, Felipe Valenca-Pereira, Catherine Pham-Danis, Francesca Cendali, Michael C. Yarnell, Amanda J. Novak, Jorge Henao-Mejia, Richard A. Flavell, Angelo D'Alessandro, Mark Eric Kohler, Mercedes Rincon. Increasing mitochondrial respiration by silencing MCJ/DnaJC15 enhances CD8 CAR-T cell therapy efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4013.
Abstract 4040: Hypoxia-induced HIF-1α/USP3 pathway promoting immune escape in lung squamous cell carcinoma
Authors: Zhang Xiuwen , Huang Hua , Ding Chen , Chen Chen , Li Yongwen , Liu Hongyu , Jun Chen
Published: March 2024

Background: Immune escape poses a significant mechanism for tumor progress as well as enormous challenge for the treatment of lung squamous cell carcinoma (LUSC). The close association between hypoxic environments and tumor immune escape, with PD-L1 serving as a key marker, has garnered considerable attention. This study aims to delve into the molecular mechanisms underlying the regulation of PD-L1 deubiquitination by the hypoxia-induced HIF-1α/USP3 pathway, shedding light on the immune escape mechanisms in LUSC.

Methods: NSCLC cells were cultured under hypoxia condition and the expression of USP3, HIF-1α and PD-L1 were measured by Western blot. Chromatin Immunoprecipitation (ChIP) and Co-immunoprecipitation (CO-IP) were employed to elucidate the direct regulatory mechanisms of the HIF-1α/USP3 pathway in PD-L1 deubiquitination. In vitro T cell-mediated killing assays was used to investigate the functional roles of USP3 in antitumor immunity in NSCLC.

Results: We demonstrated that hypoxia score was increased in LUSC and negatively correlated with CD8+ T cell infiltration. Hypoxia as a major factor triggering LUSC cell immunosuppression and against T cell surveillance via stabilization of PD-L1. Mechanistically, USP3, induced by HIF-1α, is required for hypoxia-mediated PD-L1 stabilization in LUSC cells. USP3 inhibits the ubiquitination and degradation of PD-L1. The combination of USP3 inhibitors with PD-1/PD-L1 monoclonal antibodies significantly enhances the anti-LUSC effect, possibly through promoting the infiltration of CD8+ T and NK cells.

Conclusion: This research unveils a novel mechanism by which hypoxia, through the HIF-1α/USP3 pathway, regulates PD-L1 deubiquitination, thereby promoting immune escape in LUSC. Furthermore, our study provides a new strategy involving USP3 inhibitors in combination with PD-1/PD-L1 monoclonal antibodies, offering novel insights for molecular targeted therapy and reversal of drug resistance in LUSC.

Citation Format: Xiuwen Zhang, Hua Huang, Chen Ding, Chen Chen, Yongwen Li, Hongyu Liu, Chen Jun. Hypoxia-induced HIF-1α/USP3 pathway promoting immune escape in lung squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4040.
Abstract 4045: Dependence of human immune cell type, and impact of IL-7R or TLR1/2 agonism, upon trastuzumab-mediated ADCC
Authors: Bryant Mark , Tut Gokhan , Harrison Neale , Welsh Emma , Morano Ines , Kaur Arshpreet , Pegg Ashley , Chan Hujo , Li Zhi , Cowley Jamie , Brady Catherine , Gordon John , Barnes Nicholas , Qureshi Omar
Published: March 2024

Targeting of tumors over-expressing HER2 using trastuzumab provides clinical benefit for patients by inhibiting HER2 signalling and enhancing immune responses against the tumor by antibody-dependent cell-mediated cytotoxicity (ADCC). Cytokines such as IL-7 augment cytotoxic anti-viral responses through up-regulation of FasL, whereas TLR1/2 agonists enhance T cell cytotoxic responses but may also promote tumor cell survival. We therefore investigated the ability of IL-7R or TLR1/2 agonism to enhance trastuzumab-mediated ADCC.

We first determined the contribution of various human immune cell subsets (e.g. monocytes, T cells, NK cells) to deliver ADCC of human tumor cells using a live high-content imaging assay in both 2D and 3D formats. Our results demonstrate that trastuzumab evoked immune cell dependent, concentration-dependent killing of tumor cells that exhibited some differential sensitivity dependent upon the cell culture format. We then tested trastuzumab-mediated killing in the absence or presence of IL-7R or TLR1/2 agonism, and noted evident donor and immune cell dependent modulation of the cytotoxic response. Modulation of this cytotoxic response was associated with differential immune cell marker expression when analysed by flow cytometry.

Our findings provide mechanistic insight into trastuzumab-mediated ADCC and highlight how cytokine receptor and TLR pathways may modulate this response.

Citation Format: Mark Bryant, Gokhan Tut, Neale Harrison, Emma Welsh, Ines Morano, Arshpreet Kaur, Ashley Pegg, Hujo Chan, Zhi Li, Jamie Cowley, Catherine Brady, John Gordon, Nicholas Barnes, Omar Qureshi. Dependence of human immune cell type, and impact of IL-7R or TLR1/2 agonism, upon trastuzumab-mediated ADCC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4045.
Abstract 4101: Immune inhibition of murine colorectal cancer by a p53 peptide mucosal vaccine
Authors: Wang Suhe , Cao Zhengyi , Janczak Katarzyna W. , Feng Ying , Green Maranne , Fearon Eric R. , Baker Jr James R.
Published: March 2024

Colorectal cancer (CRC) is among the most common cancers with high morbidity and mortality for patients despite multi-modality treatment options. Immune checkpoint inhibitors often have impact in the 15% of CRC patients with microsatellite instability-high status CRCs but have limited utility in the remaining 85% of CRC patients. Given the presence of p53 mutations in CRC, we pursued a vaccine approach against tumor cells expressing murine CRC Trp53 codon 270 missense alleles (codon 273 mutation in human cancer) using a combination of R270H p53 peptide, wild type p53 recombinant protein and used a mucosal adjuvant (MA) to drive epithelial immune responses. The vaccine (p53/MA) was studied in a CDX2-CreERT2 (tamoxifen-induced Cre-Lox) transgenic mouse CRC model where tumors arise in the cecum and proximal colon epithelium following combined ApcΔ580, KrasG12D, and Trp53R270H mutations (AKP model). The p53/MA vaccine treatment was administered intranasally to the mice at defined time points after tamoxifen tumor induction. p53/MA vaccinated animals had increased anti-p53 IgG (p<0.01), IgG2a (p<0.01), and IgG2b (p<0.001) in sera compared to control mice. The treatment enhanced antigen-specific Th1 and Th17 cellular immune responses, as shown by increasing of IFN-ɤ (p<0.05), IL-17a (p<0.01) and IL-2 (p<0.01) both in splenocytes and draining lymph nodes measured by Luminex assay. Vaccinated mice also showed significant more cells producing IFN-ɤ (p<0.001) and IL-17a (p<0.001) as analyzed by ELISpot. Importantly, the immunized animals had decreased tumor size/volume (p<0.001) and prolonged survival (p<0.05) compared to control animals that received only saline or MA respectively. Tumors from vaccinated mice also showed more mononuclear cell infiltrates than tumors from the control animals. In conclusion, novel p53/MA vaccinations can induce significant p53-specific immune responses and anti-tumor effects in the mouse AKP CRC model.

Citation Format: Suhe Wang, Zhengyi Cao, Katarzyna W. Janczak, Ying Feng, Maranne Green, Eric R. Fearon, James R. Baker Jr. Immune inhibition of murine colorectal cancer by a p53 peptide mucosal vaccine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4101.
Abstract 4123: Attenuating breast cancer metastasis with BCL-XL-targeting PROTACs
Authors: Carelock Madison E. , Zhang Peiyi , Jiao Mo , De Umasankar , Jin Zeng , Stump Rachel M. , Williams Sarah G. , Zheng Guangrong , Zhang Weizhou
Published: March 2024

B-cell lymphoma X-large (BCL-XL) emerges as a significant player in cancer progression. In our recent findings, we unveil the efficacy of proteolysis-targeting chimeras (PROTACs) directed against BCL-XL, demonstrating their capacity to not only eliminate cancer cells but also target tumor-induced regulatory T cells (TI-Tregs). Notably, BCL-XL's role in promoting breast cancer metastasis extends beyond its anti-apoptotic function. We posit that BCL-XL-targeting PROTACs (BCL-XL-Ps) hold promise for treating metastatic breast cancer. Their potential lies in directly eliminating circulating tumor cells during vulnerable stages and/or triggering anti-cancer immunity through Treg cell depletion. Our experiments underscore the potency of IAP-based PROTACs in facilitating BCL-XL degradation via the ubiquitin-proteasome system (UPS) while mitigating on-target platelet toxicity. Treatment of breast cancer-bearing mice with these compounds demonstrates a tangible reduction in breast cancer metastasis to the lungs. Additionally, our studies reveal a significant depletion of tumor-infiltrating Tregs in mice treated with BCL-XL-Ps. While affirming BCL-XL as a viable therapeutic target for breast cancer metastasis, further exploration into the mechanisms behind BCL-XL-P targeting is imperative. Our future endeavors will be devoted to unraveling whether our BCL-XL-Ps effectively target both tumor-intrinsic BCL-XL, potentially inducing tumor cell death, and tumor-infiltrating Tregs, thereby fostering an adaptive immune response against the tumor cells.

Citation Format: Madison E. Carelock, Peiyi Zhang, Mo Jiao, Umasankar De, Zeng Jin, Rachel M. Stump, Sarah G. Williams, Guangrong Zheng, Weizhou Zhang. Attenuating breast cancer metastasis with BCL-XL-targeting PROTACs [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4123.
Abstract 4138: Evaluation of breast triple-negative 4T1-derived tumor metastatic development in gender-different mouse
Authors: Shao Chunhong , He Melanie , Tsai Luke Tsung-Huang
Published: March 2024

Background and Objective: Breast cancer in men is a rare and less-studied case, with therapeutic strategies predominantly relying on clinical treatment protocols established for women. To support the clinical treatment concept, Dr. Martin-Martinez's lab observed no differences in the time of orthotopic breast mammary fat pad tumor development, necrosis, or color change of tumor tissue between male and female mice. However, the clinical treatment readout, especially tumor metastasis, is different in men and women, and current models for breast cancer are inadequate due to the absence of metastases. It is necessary to evaluate the development of breast cancer 4T1-derived tumor metastasis in male and female mice.

Method and Result: 4T1-Luc breast cancer cells were implanted in male and female Balb/c mice via various routes, including inguinal fat pad injection, tail vein injection, caudal artery injection, portal vein injection, and Intratibial Injection. Illuminance images showed no significant differences in tumor metastatic development between genders for inguinal fat pad, tail vein, and intratibial injections. However, following caudal artery injection, early-stage metastasis differed; male mice developed femoral head metastasis, while female mice developed lung metastasis. In the late stage, male mice displayed lung and femoral bone metastasis, while female mice primarily showed lung metastasis with barely bone metastasis.

Conclusion: While both male and female mice showed similar primary breast tumor development, mouse gender affected tumor metastasis differently, particularly when tumor cells went through the caudal artery. Male mice presented earlier femoral head metastasis and less lung metastasis than female mice. Based on this work, sex hormones would affect both hormone-related and nonhormone-related tumor metastasis. To elucidate whether this phenomenon is caused by estrogen or androgen or to test if a drug has any beneficial effects, HD Biosciences provides established tumor metastatic cell-delivering platforms and over 1000 cancer cell lines for target validation and drug efficacy study.

Citation Format: Chunhong Shao, Melanie He, Luke Tsung-Huang Tsai. Evaluation of breast triple-negative 4T1-derived tumor metastatic development in gender-different mouse [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4138.