bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 6556: Targeting mesenchymal tumor cell-intrinsic factors sensitizes refractory tumors to immune checkpoint blockade therapy
Authors: Bakhle Kimaya M. , Sams Caitie H. , Dongre Anushka
Published: March 2024

Immune checkpoint blockade (ICB) therapy has emerged as a promising approach for treating breast cancer, however, only a fraction of patients derives clinical benefit and the underlying reasons for heterogeneous responses remain elusive. The epithelial-to-mesenchymal transition (EMT) enables these carcinomas to metastasize and acquire resistance to chemotherapy. We have recently demonstrated that the EMT program also confers resistance to immunotherapies. Specifically, epithelial tumors recruit CD8+ T-cells to the tumor microenvironment and are sensitive to anti-CTLA4 ICB. In contrast, mesenchymal tumors assemble an immunosuppressive tumor microenvironment and are resistant to the same treatment. Furthermore, mesenchymal tumors express multiple immunosuppressive paracrine factors relative to their epithelial counterparts. Of these, abrogation of CSF1 or SPP1 from mesenchymal cancer cells generates partial responses to ICB therapy. Strikingly, abrogation of CD73 from mesenchymal carcinoma cells completely sensitizes otherwise refractory tumors to immunotherapy. While targeting cancer cell-intrinsic factors can sensitize mesenchymal tumors to ICB, the mechanisms that dictate partial versus complete responses are unclear. Furthermore, whether abrogation of CD73 can also sensitize mesenchymal tumors to other forms of ICB remains unknown. By performing immunofluorescent analysis of tumor sections, we observed that knockout of CD73 promoted the greatest infiltration of both CD4+ and CD8+ T-cells in response to anti-CTLA4 ICB compared to tumors knocked out for CSF1 and SPP1, which showed only intermediate levels of infiltration. Moreover, treatment of mesenchymal tumor-bearing mice with anti-CD73 generated synergistic responses with anti-CTLA4 ICB, but not with anti-PD1, or combinations of anti-CTLA4 and anti-PD1. In conclusion, our results demonstrate that the EMT program in cancer cells is predictive of responses to ICB. Targeting CD73 in mesenchymal breast tumors promotes an influx of both CD4+ and CD8+ T-cells and potentiates the efficacy of anti-CTLA4 ICB. Through mining of canine mammary tumor RNA sequencing data, we have found that canine tumors associated with EMT have increased expression of CD73, which suggests that these mechanisms of immune evasion may translate across species. These findings could promote therapies for both animals and humans that sensitize highly refractory populations of more-mesenchymal cancer cells.

Citation Format: Kimaya M. Bakhle, Caitie H. Sams, Anushka Dongre. Targeting mesenchymal tumor cell-intrinsic factors sensitizes refractory tumors to immune checkpoint blockade therapy [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 6556.
Abstract 6624: MYC promotes lobuloalveologenesis and mammary tumorigenesis in male mice
Authors: Pulver Emilia M. , van der Burg Eline , Drenth Anne Paulien , Yemelyanenko Julia , Ravensbergen Nils , de Bruijn Roebi , Klarenbeek Sjoerd , Song Ji-Ying , Boeije Manon , Siteur Bjørn , Proost Natalie , Scheele Colinda , Bouwman Peter , Jonkers Jos
Published: March 2024

Male breast cancer is a rare disease that is largely overlooked and clinically regarded to recapitulate female breast cancer biology. The worse overall outcome for male patients compared to females, however, underscores inadequate disease management and a sex-based divergence in disease biology. To gain more insight into male breast biology and pathology, we used mouse models of triple-negative breast cancer and 3D whole-mount confocal imaging techniques to evaluate both in vivo and ex vivo mammary gland development, tumor formation, and hormone signaling dependence. Our results show that Myc overexpression in collaboration with Trp53 deficiency drives mammary tumorigenesis in male mice. We show that MYC overexpression promotes estrogen-independent formation of lobuloalveolar structures, which progress into tumors with a similar histology and immune landscape as female mammary tumors but with significantly more genomic instability. In female mice, MYC overexpression bypasses the dependency on paracrine estrogen signaling in mammary tumorigenesis. These data provide robust evidence that MYC overrides mammary tumor estrogen dependence and abolishes the tumor-preventive effect of a low-estrogen environment. Supporting this notion, in vitro experiments in human female breast cancer cell lines indicate that MYC overexpression may functionally contribute to endocrine therapy resistance and therefore could possibly serve as a prognostic biomarker or clinically targetable pathway for treating both male and female breast cancers. Collectively, this preclinical work presents new insights into male mammary gland biology and mechanisms of cancer progression that might translate to a broader range of breast cancers in the clinic.

Citation Format: Emilia M. Pulver, Eline van der Burg, Anne Paulien Drenth, Julia Yemelyanenko, Nils Ravensbergen, Roebi de Bruijn, Sjoerd Klarenbeek, Ji-Ying Song, Manon Boeije, Bjørn Siteur, Natalie Proost, Colinda Scheele, Peter Bouwman, Jos Jonkers. MYC promotes lobuloalveologenesis and mammary tumorigenesis in male mice [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 6624.
Abstract 6633: Tracking cell-cell interactions using intercellular barcode transfer
Authors: Tay Neil Q. , Juan Tiffany , Muldoon Joseph J. , Eyquem Justin , McManus Michael T.
Published: March 2024

Cell-cell interaction is one of the fundamental biological mechanisms by which cells communicate and is a key modality by which cancer cells interact with other cells in their microenvironment. Current methods, such as enzymatic labeling and proximity-based tagging, allow the experimental tracking of these interactions but are not compatible with sequencing-based readouts, which are essential for high-throughput analyses. We have developed a highly scalable intercellular barcode transfer technology that bridges this gap. Our novel “Relay” technology offers a scalable solution to monitor cell-cell interactions with unprecedented single-cell resolution, seamlessly integrating with existing high-throughput and sequencing platforms. Relay employs RNA barcodes to mark and track cell interaction events, providing definitive quantitative data on cell interaction partners. We used Relay to track interactions between cancer cells and T cells, capturing both potent antigen-specific engagements and subtler contacts involved in antigen sampling. Additionally, by integrating CRISPR libraries with Relay, we pioneered a novel approach for pooled CRISPR screens, focusing on intercellular interactions across different cell types. In this context, the cell harboring genetic perturbation may differ from the cell under analysis. To demonstrate this, we are conducting genome-wide perturbations in cancer cells and interrogating cancer cell factors that affect antigen sampling and trogocytosis by T cells and macrophages. Beyond its application in CRISPR screening, Relay can also be used with single-cell RNA sequencing to directly track and measure immune-cancer and immune-immune cell interactions at the single cell level. We applied single-cell Relay to a dendritic cell-T cell system to link dendritic cell signatures to T cell signatures based on direct cell contact and regulatory signaling. The design of Relay as a highly modular system extends its utility beyond cancer research, making it broadly applicable across various biological models and systems where cell-cell communication is a fundamental property in complex multicellular settings. This tool promises to accelerate the transition from theoretical understanding to practical exploration of cell-cell interactions, with profound implications for immunology, oncology, and beyond.

Citation Format: Neil Q. Tay, Tiffany Juan, Joseph J. Muldoon, Justin Eyquem, Michael T. McManus. Tracking cell-cell interactions using intercellular barcode transfer [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 6633.
Abstract 6664: In situ vaccination with hyper-interferon sensitive influenza induces adaptive immune response and sensitizes murine non-small cell lung cancer to PD-1 blockade
Authors: Salehi-Rad Ramin , Du Yushen , Zhang Tianhao , Shi Yuan , Liu Bin , Ren Sun
Published: March 2024

Despite recent advances in immunotherapy with immune checkpoint inhibitors (ICI), many patients with non-small cell lung cancer (NSCLC) fail to respond or develop resistance after an initial response. The exclusion of T cells from the tumor or the presence of a dysfunctional T cell compartment within the tumor microenvironment (TME) constitute two central hallmarks of resistance to ICI in patients with NSCLC. Seminal studies have identified that loss of LKB1 in KRAS-mutant NSCLC drives resistance to ICI, potentially by suppressing STING, thereby leading to dysregulation of the interferon (IFN) signaling. There is a pressing demand for the development of innovative immune therapies designed to bolster anti-tumor T cell responses to overcome resistance to ICI. In situ vaccination (ISV) with oncolytic viruses has emerged as a promising, antigen-agnostic strategy capable of conditioning the TME and amplifying anti-tumor T cell responses. We engineered a live attenuated viral vaccine, Hyper-Interferon Sensitive virus (HIS), by conducting a genome-wide functional screening and introducing eight interferon (IFN)-sensitive mutations in the influenza A genome. We utilized multiple syngeneic murine models of NSCLC with distinct driver mutations [KrasG12D/P53+/−/Lkb1−/− (KPL); KrasG12D/P53+/− (KP); and KrasG12D (K)] and varying tumor mutational burden to assess the safety and efficacy of intratumoral HIS virotherapy. Compared to wild-type (WT) influenza, HIS replication was attenuated, enhancing its potential as a safe option for cancer therapy. HIS ISV elicited robust yet transient type I IFN responses in murine NSCLCs, leading an influx of polyfunctional effector Th1 CD4 and cytotoxic CD8 T cells into the tumor. HIS ISV demonstrated enhanced anti-tumor efficacy compared to WT in multiple syngeneic murine models of NSCLC. This efficacy was dependent on host type 1 IFN responses and T lymphocytes. HIS ISV synergized with anti-PD-1 to eradicate immune resistant LKB-1 deficient NSCLC in a subset of mice, resulting in improved overall survival and enduring systemic tumor-specific immunity. These studies provide compelling evidence to support further clinical evaluation of HIS as a novel ‘off-the-shelf’ ISV strategy for patients with NSCLC refractory to ICI.

Citation Format: Ramin Salehi-Rad, Yushen Du, Tianhao Zhang, Yuan Shi, Bin Liu, Sun Ren. In situ vaccination with hyper-interferon sensitive influenza induces adaptive immune response and sensitizes murine non-small cell lung cancer to PD-1 blockade [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 6664.
Abstract 6670: Could immunotherapy offer potential in managing bacterial dysbiosis in prostate cancer
Authors: Oliver Tim , Nedjai Belinda , Lane Emily , Chinegwundoh Frank , Rajan Prabhakar , Cuzick Jack
Published: March 2024

Background: Since 2006 three studies, involving 289,089 adolescents, demonstrated a significant association (1.67(0.79-3.55),1.7(1.03-2.70),2.37(1.19-4.73)) between pubertal acne and poor outcome prostate cancer(PC) 30+ years later. This suggested Cutibacterium acnes, amicroaerophylic bacterium, might be contributing to PC bacterial dysbiosis analogous to the role of H. pylori in stomach cancer and explain why circumcision reduces PC risk. Six years ago, using MALDI-TOF, we demonstrated that the presence of seven obligate anaerobe species in addition to C. acnes were increased in PC and linked to increased PSA. This presentation studies stored samples from the PROVENT chemoprevention randomised PC active surveillance phase 2 trial analysed using 16S rDNA sequencing, and comparing obligate anaerobe frequency with those detected by culture and MALDI-TOF identification and their influence on PSA expression.

Methods: 89 urines of PROVENT trial patients were studied using 16S rDNA sequencing and followed for 24 months. In addition, 39 previously reported MALDI-TOF screened cohort patients (21 PCs and 18 BPHs) followed for 43 months. The frequency of 7 obligate bacterial genera (Actinobaculum, Finegoldia, Prevotella, Peptoniphilus, Peptostreptococcus & Viellonella) was studied.

Results: Twenty six of 89 (29%) PROVENT urine samples had a majority of bacteria detected as obligate anaerobes (mean=68%), while it was 12 of the 39 (31%) in PC tested in MALDI-TOF. In the combined series, those patients (n=38) with obligate anaerobes had mean PSA of 9.59 ug/L, while those without obligate anaerobes had a mean PSA of 6.29 ug/L (n=90), (t=-2.925, p = 0.0058). In the MALDI-TOF series, statistically increased urological intervention rate was associated with the presence of anaerobes (p=0.045, Fisher’s exact test).

Discussion: These findings suggest that obligate anaerobe bacterial dysbiosis could be contributing to raised PSA through prostate cell damage, and possibly participating in the chronic inflammation and hypervascular changes that are seen in precancerous lesions. Checkpoint inhibitors have shown little benefit in prostate cancer. Recent reports from animal model studies of BCG resistance have suggested a short course combination of anti PDLI and anti NKG2a could break immune tolerance associated with expression of HLA-E and this approach could be of use to break tolerance to bacterial dysbiosis in the future.

Citation Format: Tim Oliver, Belinda Nedjai, Emily Lane, Frank Chinegwundoh, Prabhakar Rajan, Jack Cuzick. Could immunotherapy offer potential in managing bacterial dysbiosis in prostate 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 6670.
Abstract 6672: Unraveling the microbiome: antibiotic impact on CAR T therapy and the synergistic potential of vancomycin in enhancing antitumor immunity
Authors: Facciabene Andrea , Uribe-Herranz Mireia , Bhoj Vijay
Published: March 2024

The landscape of treating hematologic malignancies has undergone a revolutionary transformation with the advent of Anti-CD19 chimeric antigen receptor T cell therapy (CART-19). Despite its success, persistent challenges like disease relapse and CAR-mediated toxicities remain. A comprehensive multicenter study involving 228 individuals revealed a significant association between antibiotic administration, particularly piperacillin/tazobactam, meropenem, and imipenem/cilastatin (P-I-M), targeting anaerobes within four weeks preceding CART-19 therapy, and adverse outcomes, including worsened survival and increased neurotoxicity. Intriguingly, another antibiotic, Cefepime, also used for neutropenic fever, displayed a positive trend when compared with P-I-M. This highlights the intricate relationship between the modulation of the intestinal microbiome and outcomes in CART-19 therapy for B cell malignancies. Building on these critical observations, our study delved into the impact of vancomycin-induced gut microbiota dysbiosis on chimeric antigen receptor (CAR) T immunotherapy. Utilizing preclinical models of CD19+ A20 lymphoma and CD19+ B16 melanoma, we discovered that mice receiving vancomycin alongside CART-19 therapy exhibited enhanced tumor control and increased cross-presentation of tumor-associated antigens (TAAs) compared to those receiving CART-19 alone. This effect was further validated through fecal microbiota transplants from healthy human donors to pre-conditioned mice, replicating the favorable findings. Importantly, our study extends to clinical correlates, as B cell acute lymphoblastic leukemia patients treated with CART-19 and oral vancomycin displayed elevated CART-19 peak expansion compared to unexposed patients. These compelling results underscore the profound influence of the gut microbiota on CAR T cell therapy and suggest that vancomycin-induced gut microbiota modulation may emerge as a potent strategy to enhance therapeutic outcomes across various tumor types.

Citation Format: Andrea Facciabene, Mireia Uribe-Herranz, Vijay Bhoj. Unraveling the microbiome: antibiotic impact on CAR T therapy and the synergistic potential of vancomycin in enhancing 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 6672.
Abstract 6714: AI-guided tailor-made anti-TROP2 T-cell engagers for T-cell mediated cancer immunotherapy with enhanced therapeutic index
Authors: Chen Chuan , Tian Liang , Su Chenpeng , Liu Dandan , Chen Xiaoqian , Zhou Yifei , Jiang Suze , Wang Guifang , Tian Jiyuan , Yuan Yuan , Yang Bing , Peng Jian
Published: March 2024

Bispecific T-cell engagers (TCEs) have shown significant therapeutic advances in the treatment of hematologic cancers. However, the clinical efficacy of TCEs in treating solid tumors remains to be demonstrated. Accumulating evidence suggests novel TCE modalities that simultaneously engage both CD3 (signal 1) and CD28 (signal 2) co-stimulatory signals may offer potential advantage to enhance clinical efficacy of TCEs with improved therapeutic index. To explore the full potential of TCEs as a powerful approach of immunotherapy for solid tumors, we developed an integrated AI-guided TCE platform to streamline the discovery, optimization and high-throughput function screening and characterization of TCE molecules. Here we report the construction and characterization of a novel panel of TROP2-targeting TCEs by leveraging our AI-guided optimization technology, plus a high-throughput in vitro screening platform. These tailor-made TCE molecules exhibit favorable developability and functional properties. Our data show that a novel TROP2 × CD3 x CD28 trispecific TCE that contains a CD3-targeting arm in concomitant with a CD28-targeting arm can induce potent T-cell co-stimulation and promising synergistic T cell-mediated tumor cell killing in vitro, compared to its counterparts with a single CD3-targeting arm. We further generate a trivalent TROP2 x CD3 bispecific TCE constructed as an asymmetric ‘2+1’ format with bivalent binding to TROP2 but a monovalent CD3-targeting arm. This molecule stimulates very low levels of cytokine release, but drives robust antigen-specific killing in in vitro assessment. Anti-tumor activity of the various TROP2 x CD3 TCEs is currently being investigated in multiple tumor xenograft mouse models.

Citation Format: Chuan Chen, Liang Tian, Chenpeng Su, Dandan Liu, Xiaoqian Chen, Yifei Zhou, Suze Jiang, Guifang Wang, Jiyuan Tian, Yuan Yuan, Bing Yang, Jian Peng. AI-guided tailor-made anti-TROP2 T-cell engagers for T-cell mediated cancer immunotherapy with enhanced therapeutic index [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 6714.
Abstract 6752: Targeting of neoantigen cancer vaccines to antigen presenting cells results in faster and broader T cell responses whether encoded in DNA or mRNA
Authors: Weinert Brian T. , Skullerud Lise , Solbakken Eirik , Textor Ana , Dürkoop Mirjam , Winge Inga Patrycja , Norheim Gunnstein , Stubsrud Elisabeth , Granum Stine , Pedersen Mikkel W. , Norell Håkan
Published: March 2024

Individualized cancer vaccines based on patient-specific neoantigens represent a groundbreaking approach in oncology, offering a highly individualized treatment option for cancer patients. Unlike traditional vaccines, which are designed to prevent diseases by stimulating an immune response against a specific pathogen, personalized cancer vaccines are therapeutic, aiming to treat cancer by training the immune system to recognize and attack the patient’s own cancer cells. The key is to induce a fast, strong, broad, and persistent CD8+ T cell response.

Nykode Therapeutics has developed a platform that targets antigens directly to antigen presenting cells (APCs) using a modular dimeric protein format known as a VaccibodyTM. Individualized neoantigen vaccines delivered in a DNA-encoded Vaccibody format have been shown to induce strong and broad T cell responses in pre-clinical and clinical settings.

Here, we wanted to determine if the APC-targeted approach leads to superior T cell responses if Vaccibody vaccines are encoded in mRNA and administered in a lipid nanoparticle (LNP) formulation. For this purpose, we designed and evaluated a model cancer vaccine composed of 20 unstructured 27 amino acid long neoantigens derived from the murine MC38 tumor model.

A DNA-encoded Vaccibody vaccine elicited faster T cell responses compared to an untargeted mRNA-LNP-encoded vaccine. DNA-encoded Vaccibody responses were also broader, targeting a substantially greater number of neoantigens.

Next, we compared the APC-targeting Vaccibody vaccine to an untargeted vaccine when both were formulated in an mRNA-LNP format. The APC-targeting Vaccibody vaccine elicited a greater breath and magnitude of T cell responses compared to the untargeted vaccine. Further studies to evaluate the functional differences associated with the various formats, in terms of T cell phenotype and response to tumor challenge, are ongoing.

In summary, the data demonstrate that an APC-targeted Vaccibody vaccine containing MC38 neoantigens can be encoded in both DNA and mRNA-LNP to induce faster, broader, and stronger T cell responses compared to a non-targeted neoantigen vaccine.

Citation Format: Brian T. Weinert, Lise Skullerud, Eirik Solbakken, Ana Textor, Mirjam Dürkoop, Inga Patrycja Winge, Gunnstein Norheim, Elisabeth Stubsrud, Stine Granum, Mikkel W. Pedersen, Håkan Norell. Targeting of neoantigen cancer vaccines to antigen presenting cells results in faster and broader T cell responses whether encoded in DNA or mRNA [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 6752.
Abstract 6759: Antigen-pHLIP for tumor treatment
Authors: DuPont Michael , Visca Hannah , Moshnikova Anna , Engelman Donald , Reshetnyak Yana , Andreev Oleg
Published: March 2024

Targeted antigen delivery allows activation of the immune system to kill tumor cells. Among the immunogenic epitopes widely investigated is α-Gal, since humans exhibit specific anti-Gal reactivity. However, the repertoire of immunogenic epitopes could be significantly enhanced if therapeutic efficacy did not rely on the presence of natural antibodies, but instead relied on the production of anti-epitope endogenous antibodies induced by immunization against the selected epitope. Therefore, we designed and tested several HA-pHLIP agents, where the HA peptide (YPYDVPDYA) from the influenza virus was selected as an exogenous highly immunogenic epitope, since it is absent in the human genome, and a pH Low Insertion Peptide (pHLIP) was selected as the tumor targeting agent. HA peptides were attached to the N-termini of pHLIPs either directly or via PEG polymers. To enhance the overall binding affinity of HA-pHLIP to anti-HA antibodies, double headed HA-pHLIP agents were introduced, where two HA epitopes were linked to a single pHLIP peptide via PEG12 polymers. The pHLIP sequence was modified to ensure proper pKs of insertion into cell membranes. HA epitopes were targeted to cancer cells in mouse tumors, and remained exposed to the extracelullar space within tumors for about 24 hours. Double-headed HA-pHLIP treatments of imunized mice (a high titer of anti-HA antibodies was developed in mice after immunization with KLH-HA), bearing 4T1 triple negative or B16F10 MHC-I negative melanoma tumors resulted in a 55% reduction of tumor growth. Further reduction of tumor growth was impossible to achive, since all of the anti-HA antibodies in the blood were captured after 3 injections of HA-pHLIP and additional boosts would be required to restore the level of free anti-HA antibodies to induce a more significant therapeutic efficacy.

Citation Format: Michael DuPont, Hannah Visca, Anna Moshnikova, Donald Engelman, Yana Reshetnyak, Oleg Andreev. Antigen-pHLIP for tumor treatment [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 6759.
Abstract 6801: Cellular senescence involves spatial evolution in colorectal cancer
Authors: Park Tae Jun , Kang Seok Yun , Kim Jang Hee , Lee Hyun Woo , Choi Yong Won , Park Soon Sang , Lee Young-Kyoung
Published: March 2024

Cancer evolution is a continuous process involving cellular changes that promote progression and metastasis. Recent studies demonstrate that cancer evolution is spatially associated: a phenomenon called “the spatial evolution of cancer.” In our data, colorectal cancer cells gradually evolve from an adenoid to a collective invasion morphology, ultimately developing a partial epithelial-mesenchymal transition phenotype as they form an invasive front. Trajectory analysis showed that cancer cells at the invasive front of a tumor exhibit a totally different gene expression pattern from those at the center, and that cellular senescence plays a crucial role in this transition. Epigenetic changes in the promoter/exon1 region of the CDKN2A gene caused by increased reactive oxygen species level are crucial for expression of p16INK4A and induction of senescence in the cancer cells. Two types of senescent tumor cells (type I and type II) were observed, and they played different roles in cancer progression due to different gene expression. Type II senescent tumor cells (p16INK4A+/LAMC2+/MMP7+), a final evolved form of cancer cells, are strongly associated with local invasion and lymph node metastasis of colorectal cancer and worsen the prognosis for patients.

Citation Format: Tae Jun Park, Seok Yun Kang, Jang Hee Kim, Hyun Woo Lee, Yong Won Choi, Soon Sang Park, Young-Kyoung Lee. Cellular senescence involves spatial evolution in colorectal 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 6801.
Abstract 6803: TRIM16 regulates IL-6 secretion in head and neck cancer-associated fibroblasts
Authors: Ly Thuc , Pickard Bailey , Pandey Avisha , Martinez-Rivera Noraida , Rosa-Molinar Eduardo , Washburn Michael , Ding Wen-Xing , Thomas Sufi Mary
Published: March 2024

Despite current therapy, head and neck squamous cell carcinoma (HNSCC), the 7th most common cancer worldwide, is associated with high morbidity and mortality. Response to current therapies could be improved with a better understanding of the tumor microenvironment of HNSCC. We previously reported that cancer-associated fibroblasts (CAFs), the most abundant stromal cell type in the HNSCC microenvironment, have elevated levels of basal autophagy than oral fibroblasts from cancer-free subjects (NF). Disrupting autophagy in CAFs with Beclin-1 siRNA reduced secretion of IL-6, IL-8, and other factors known to promote HNSCC progression. The recent emerging role of autophagy in secretion of tumor promoting factors has high impact because (i) the mechanism can provide a new target to block the secretion of protumor factors from CAFs that contribute to HNSCC progression, and (ii) these cytokines can serve as potential indicators for the efficacy of autophagy-targeted therapies. To this end, we carried out an unbiased assessment of the proteins associated with autophagosomes in primary CAFs from HNSCC samples using mass spectrometry. We performed immunoprecipitation of LC3B from the isolated small vesicles of the CAFs, identified and validated the association of several trafficking proteins in secretory autophagy including VAMP3 and SNAP23. Further, the tripartite motif (TRIM) proteins have been shown to mediate the fate of autophagosomes either for degradation or secretion. We hypothesized that TRIM proteins are involved in transport of secretory autophagosomes. Our data demonstrate CAFs express higher levels of TRIM16 mRNA than NF. We demonstrate immunogold labeling of TRIM16 or IL-6 localized to the autophagosomes using transmission electron microscopy. Further, using immunofluorescence, we demonstrated the colocalization of TRIM16 and IL-6, as well as IL-6 and LC3B in cytoplasmic puncta. In addition, we used proximity ligation assays to confirm the interaction of TRIM16 and IL-6, as well as IL-6 and LC3B. TRIM16 knockdown using siRNA in CAFs reduced the secretion of IL-6 as measured by ELISA. In conclusion, we demonstrate an important role of TRIM16 in secretory autophagy making it a potential therapeutic target to mitigate CAF-mediated HNSCC growth.

Citation Format: Thuc Ly, Bailey Pickard, Avisha Pandey, Noraida Martinez-Rivera, Eduardo Rosa-Molinar, Michael Washburn, Wen-Xing Ding, Sufi Mary Thomas. TRIM16 regulates IL-6 secretion in head and neck cancer-associated fibroblasts [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 6803.
Abstract 6808: Investigating the role of CXCR3+ monocytes on CAR T cell function
Authors: Ong Claire Victoria , Kaczanowska Sabina , Ju Wei , Cronk James , Ramakrishna Sneha , Kaplan Rosandra
Published: March 2024

Chimeric antigen receptor T cells (CAR-Ts) are an emerging immunotherapy that have remarkable efficacy against leukemias and lymphomas but limited success against solid tumors. We conducted a phase I clinical trial (NCT 02107963) of GD2 CAR-T (GD2-CAR.OX40.28.z.ICD9) administration to children and young adults with neuroblastoma and, for the first time, osteosarcoma. While all patients progressed and were switched to alternative therapy, we observed a significant association between good CAR-T expanders and expression of C-X-C Motif Chemokine receptor 3 (CXCR3) on monocytes detected in pretreatment apheresis. This study seeks to uncover the possible interactions between CXCR3 and CAR-Ts that may alter CAR-T function and ultimately lead to better CAR-T expansion. We hypothesize that CXCR3+ monocytes induce functional change in CAR-Ts resulting in improved expansion.

To recapitulate the interaction between tumor, CAR-Ts and monocytes, we took 143Bs (osteosarcoma cell line) and GD2 CAR-Ts and co cultured them with THP-1s (a monocytic leukemia cell line) that were either untransduced (UTD) or transduced via lentivirus to express CXCR3 (CXCR3+). The ratio of UTD or CXCR3+ THP-1s to CAR-Ts and 143Bs was varied between co cultures. Enzyme-linked immunoassay (ELISA) for interferon gamma (IFNy) expression was run as a proxy measurement for CAR-T cell activity. Flow cytometry was used to assess for expression of activation and exhaustion markers. Differences between UTD and CXCR3+ co cultures were evaluated for statistical significance via unpaired t-test.

We observed that CXCR3 was elevated on GD2 CAR-Ts co cultured with CXCR3+ monocytes. This pattern was seen most prominently in CD4+ CAR-Ts, but also subtly in CD8+ T-cells. Exhaustion markers CD39 and LAG3 were elevated in CAR-Ts co cultured with CXCR3+ monocytes. Additionally, CAR-Ts co cultured with CXCR3+ monocytes displayed greater activity via elevated IFNy expression.

We conclude that GD-2 CAR-T cells co cultured with CXCR3+ monocytes express elevated exhaustion markers due to possible increased activity when in the presence of CXCR3+ monocytes. Additionally, we suspect that elevated CXCR3 expression on CAR-Ts co cultured with CXCR3+ monocytes may be helping CAR-Ts better target tumor cells, thus leading to improved expansion. If CXCR3 can improve CAR-T cell function, this could lead to better optimization of CAR-Ts against solid tumors and create a viable treatment option for neuroblastoma and osteosarcoma patients.

Citation Format: Claire Victoria Ong, Sabina Kaczanowska, Wei Ju, James Cronk, Sneha Ramakrishna, Rosandra Kaplan. Investigating the role of CXCR3+ monocytes on CAR T cell function [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 6808.
Abstract 6844: Discovery of a novel sialic acid O-acetyltransferase in colon tissue
Authors: Lee Bum Seok , Cook Ashley , Zeineldin Maged , Sur Surojit , Dobbyn Laura , Popoli Maria , Khalili Sana , Vogelstein Bert , Papadopoulos Nickolas , Zhou Shibin , Bettegowda Chetan , Buckhaults Phillip , Larman Tatianna , Gabrielson Kathy , Kinzler Kenneth W. , Wyhs Nicolas
Published: March 2024

Sialic acids have critical functions relating to host-pathogenic interactions and the immune microenvironment. Subsequent post-translational O-acetyl modifications add an extra layer of control by modulating many of these interactions in both normal and transformed colorectal cells. The processes through which this modification is added to sialic acids are not completely understood. To date only one gene, CASD1, has been identified to encode a sialic acid O-acetyltransferase in human cells. We used whole-genome sequencing on normal human colon tissue to identify a haplotype strongly associated with colorectal sialic acid O-acetylation, as determined by histopathological mPAS staining. Only SNP’s of a single candidate gene within the haplotype correlated perfectly in a 91-sample validation set. CRISPR-Cas9 editing of this gene in 2D cell line and 3D normal colon organoid models yielded changes to modified sialic acid levels. These findings suggest that this candidate gene functions as a novel sialic acid O-acetyltransferase in colorectal tissue and could have implications as a diagnostic tool for patients with inflammatory diseases of the bowel or as an indicator for therapeutic targets in cancer.

Citation Format: Bum Seok Lee, Ashley Cook, Maged Zeineldin, Surojit Sur, Laura Dobbyn, Maria Popoli, Sana Khalili, Bert Vogelstein, Nickolas Papadopoulos, Shibin Zhou, Chetan Bettegowda, Phillip Buckhaults, Tatianna Larman, Kathy Gabrielson, Kenneth W. Kinzler, Nicolas Wyhs. Discovery of a novel sialic acid O-acetyltransferase in colon tissue [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 6844.
Abstract 6845: MAGEA3 expression and secretion in MMR-MSH6 and MMR-PMS2 microsatellite-stable colorectal cancer
Authors: Yom Jina , Gonzalez Rachel , Gilmore Bailey , Guo Zhaoying , Han Andy , Zewdu Eden , Kong Dehe , Qu Tianli , Guo Hailey , Strom Alex , Zhao Zoe , Hu Xiaomin , Ren Qi , Ma Yan , Zhang Ranran , Wu Zhaohui , Liu Xuan , Fu Wei
Published: March 2024

Despite the breakthrough of PD-1/PD-L1 blockade therapy in the recent years, mixed responses have been shown across multiple tumor types, including in colorectal cancer (CRC), the second leading cause of death worldwide. This immunotherapy has proven to be successful for a subtype of CRC that carries mismatch repair deficiency (dMMR) and high microsatellite instability (MSI-H), despite a lack of response by microsatellite-stable CRC. For these types of non-responsive cancers, new immunotherapy targets are currently being investigated for alternative therapy such as the MAGE-A cancer testis antigen family whose expression is restricted to germline cells in normal tissue but is overexpressed in many cancer cells. In this study, we examine the relationship between MAGEA3 and MMR proteins MSH6 and PMS2. Immunohistochemical staining of CRC tissues demonstrated co-expression between MAGEA3 with both MSH6 and PMS2. Immune cells were observed to express MAGEA3 in CRC as well. To investigate the possibility of secreted MAGEA3 from tumor cells, we performed western blot analysis of CRC cell lines, ELISA, and binding assay on cell lines and the positive results suggest that MAGEA3 protein is secreted. The results suggest that MAGE-A3 may be a promising target for microsatellite-stable colorectal cancer immunotherapy.

Citation Format: Jina Yom, Rachel Gonzalez, Bailey Gilmore, Zhaoying Guo, Andy Han, Eden Zewdu, Dehe Kong, Tianli Qu, Hailey Guo, Alex Strom, Zoe Zhao, Xiaomin Hu, Qi Ren, Yan Ma, Ranran Zhang, Zhaohui Wu, Xuan Liu, Wei Fu. MAGEA3 expression and secretion in MMR-MSH6 and MMR-PMS2 microsatellite-stable colorectal 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 6845.
Abstract 6847: IGF2BP3 promotes esophageal squamous cell carcinoma progressions via Wnt/CTNNB1 pathway
Authors: Tian Zhongxian , Zhou Yongjia , Ding Yi , Wu Zhengwen , Zhao Xiaogang
Published: March 2024

The M6A RNA modification plays a crucial role in the progression of tumors. Simultaneously, IGF2BP3 is primarily expressed in the tumor cells of squamous cell carcinoma, rather than in stromal cells, including esophageal squamous cell carcinoma (ESCC), a subtype mainly prevalent in East Asia. However, the mechanism of IGF2BP3 in the progression of esophageal squamous cell carcinoma and its role in the tumor microenvironment (TME) remain unknown and require further exploration. Our research reveals that IGF2BP3 not only plays a significant oncogenic role in tumor cells, including the activation of the Wnt pathway, PI3K-AKT pathway, and MAPK pathway, but also, the high expression of IGF2BP3 is positively correlated with the enrichment of M2 macrophages in ESCC tissues, in the TME, IGF2BP3 promotes the infiltration of M2 macrophages, ultimately facilitating the progression of ESCC. Mechanistically, IGF2BP3 enhances the translation of USP36 mRNA by directly binding to its mRNA, rather than increasing the stability of mRNA. The IGF2BP3-mediated USP36 protein elevation enhances the binding capacity with CTNNB1 protein, reducing its degradation through the ubiquitin-proteasome pathway, and following improved the Wnt/CTNNB1 pathway activating, Ultimate, IGF2BP3 aggravated the malignancy of the tumor. Additionally, the activation of the Wnt pathway promotes TGFB1 transcription and secretion, subsequently activating the polarization of M2 macrophages, infuriating the formation of the tumor immune-suppressive microenvironment, ultimately promoting the progression of esophageal squamous cell carcinoma. Our findings strongly emphasize the oncogenic role of IGF2BP3 in the progression of ESCC both in vitro and in vivo, providing new insights and perspectives for the treatment of esophageal squamous cell carcinoma.

Citation Format: Zhongxian Tian, Yongjia Zhou, Yi Ding, Zhengwen Wu, Xiaogang Zhao. IGF2BP3 promotes esophageal squamous cell carcinoma progressions via Wnt/CTNNB1 pathway [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 6847.
Abstract 6858: PPARδ and type 1 interferon collaboratively regulate CIITA-MHCII pathway: Unveiling novel mechanisms of tumor Tregs
Authors: Yang Qiyuan , Liang Yuqiong , Zheng Ye
Published: March 2024

Regulatory T cells (Tregs) play an essential role in suppressing anti-tumor immunity, often resulting in unfavorable clinical outcomes across numerous cancers. However, systemic Treg depletion, while augmenting anti-tumor responses, also triggers detrimental autoimmune disorders. Thus, dissecting the mechanisms by which Tregs navigate and exert their functions within the tumor microenvironment (TME) is pivotal for devising innovative Treg-centric cancer therapies. Our study highlights the role of peroxisome proliferator-activated receptor β/δ (PPARδ), a nuclear hormone receptor involved in fatty acid metabolism. Remarkably, PPARδ ablation in Tregs escalated tumor growth and augmented the immunosuppressive characteristics of the TME. This absence of PPARδ spurred an increased expression of genes central to antigen presentation, notably CIITA and MHCII. Our results showcase a novel association where the absence of CIITA in PPARδ-deficient Tregs bolsters anti-tumor responses, casting CIITA as a pivotal downstream regulator of PPARδ within Tregs. In vitro assays demonstrated that elevated CIITA levels enhance the suppressive capacity of Tregs, facilitated by an antigen-independent interaction between Treg-MHCII and Tconv-TCR/CD4/Lag3. A significant revelation was the role of type 1 interferon as a TME signal that promotes the genesis of MHCII+ Tregs; this process is accentuated by PPARδ deficiency. In conclusion, the co-regulation between TME cues and PPARδ signaling shapes the adaptive and suppressive roles of Treg cells through the CIITA-MHCII pathway. Strategically targeting the potent MHCII+ Treg population could open a new avenue for cancer therapies by boosting anti-tumor defenses while curbing autoimmune threats.

Citation Format: Qiyuan Yang, Yuqiong Liang, Ye Zheng. PPARδ and type 1 interferon collaboratively regulate CIITA-MHCII pathway: Unveiling novel mechanisms of tumor Tregs [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 6858.
Abstract 6916: InnoPanelTMand InnoPediaTM: resources of engineered cell lines and xenografts for drug discovery
Authors: Li Siyu , Liu Chang , Wang Na , Hu Yuqing , Ning Jinying , Hao Feng
Published: March 2024

The Development of precision medicine relies on well-characterized experimental models that carry naive, induced, or engineered genomic and epigenetic features targetable by certain treatments. Patient-derived xenografts (PDX) and cell lines can serve as such in vivo and in vitro models by harboring abnormalities such as somatic and driver mutations, dysregulated gene expression and silencing, microsatellite instability, gene fusions, copy number amplification and loss of heterozygosity. In addition, cell lines can also be engineered to introduce sometimes rare-occurring abnormalities that the engineered cell line shares an identical genetic background with its parental cell line, thus providing a clearly contrasting pair to evaluate treatment specificity. With the advancement of disease research and drug discovery, existing cell lines can no longer meet the needs of various types of drug discovery. To evaluate drug efficacy more quickly and accurately, Kyinno has constructed and identified more than 4,000 engineered cell lines, including overexpressed cell lines constructed through transposon systems or lentivirus infection, gene-edited cell lines constructed through CRISPR-Cas9 technology, and reporter cell lines targeting multiple signaling pathways and membrane receptors. Besides, InnoPanelTM, a panel of over 1000 engineered cell lines was established and characterized to cover a wide range of heavily targeted genes such as CLDN18.1, CLDN18.2, LAG3, OX40, PD-L1, PD-L2, and TIM3. Cell line families were also developed for genes with multiple known driver mutations. All cell lines are annotated with background information, engineering details, functional characterization, and efficacy studies. Furthermore, we developed xenografts for several engineered cell lines and showed that the combination of cell lines and xenografts can accelerate drug development. Additionally, all of this information can be accessed in our free database, InnoPediaTM. The database also offers browsing, searching, and analytical capabilities for over 600 commonly used tumor cell lines and thousands of engineered cell lines with essential genomic and pharmacological data, cell line efficacy data for nearly 2000 common anti-tumor drugs, and comprehensive details for over 100 standard antibodies. Our engineered cell lines are now registered in the online cell line database Cellosaurus, which is available on the ExPASy server.

Citation Format: Siyu Li, Chang Liu, Na Wang, Yuqing Hu, Jinying Ning, Feng Hao. InnoPanelTMand InnoPediaTM: resources of engineered cell lines and xenografts for drug discovery [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 6916.
Abstract 6917: Ba/F3 AKT engineering cell lines, a useful platform for novel drug discovery
Authors: Wang Guoqian , Tang Yao , Xu Jingxiao , Peng Yao , Wang Yu , Ning Jingying , Hao Feng
Published: March 2024

AKT, also known as phosphoinositide 3-kinase (PI3K), plays a key role in various signaling cascades involved in cell growth and division, apoptosis inhibition, and angiogenesis. For AKT, which is at the central node of the PI3K/AKT/mTOR signaling pathway, the loss of PTEN, mutation, or amplification of AKT/PIK3CA can lead to the overactivation of the AKT signaling pathway, resulting in the occurrence and development of tumors. Recent studies have also found that AKT activation is associated with drug resistance in tumor therapy. Therefore, AKT has become a hot target for tumor therapy. The AKT family consists of three highly homologous isoforms, AKT1 (PKBα), AKT2 (PKBβ), and AKT3 (PKBγ), with more than 85% homology in their kinase regions. E17K is the most common and well-characterized activating mutation, accounting for approximately 15% of all AKT mutations. Rare mutations mainly include L52R, Q79K, and D323H. The failures of AKT inhibitors LY2780301 and Bayer's BAY1125976 demonstrate the significant challenges and complexities in developing inhibitors of the AKT pathway. Currently, the most advanced AKT inhibitor globally is AstraZeneca's capivasertib, which is primarily indicated for HR+ breast cancer and triple-negative breast cancer. Our group has generated 21 Ba/F3-AKT engineering cell lines, as useful models for novel drug discovery in vitro and in vivo, almost all clinically significant mutations were included, such as common mutations AKT1/2-E17K as well as rare mutations L52R, Q79K, D323H, and others. The Ba/F3-AKT engineering cell lines can be useful for developing and evaluating next-generation AKT inhibitors and exploring newly acquired resistance mutations in AKT.

Citation Format: Guoqian Wang, Yao Tang, Jingxiao Xu, Yao Peng, Yu Wang, Jingying Ning, Feng Hao. Ba/F3 AKT engineering cell lines, a useful platform for novel drug discovery [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 6917.