Citations

Abstract Background: Lung adenocarcinoma (ADC) and squamous cell carcinoma (SCC) are the most common lung cancer subtypes. SCC is traditionally resistant to anti-angiogenic therapies, though the mechanisms remain unclear. Nintedanib, a multi-angiokinase receptor inhibitor, is approved for treating ADC in combination with docetaxel, based on selective efficacy in the LUME-1 trial. It is also FDA-approved for idiopathic pulmonary fibrosis due to its antifibrotic properties. Both ADC and SCC feature fibrotic stroma rich in activated tumor-associated fibroblasts (TAFs). Our prior work showed Nintedanib has stronger antifibrotic effects on ADC-TAFs than SCC-TAFs. However, its impact on TAF-driven angiogenesis and the role of TAFs in SCC resistance to multi-angiokinase inhibitors remain unexplored. Methods: The pro-angiogenic activity of TAFs secretome from ADC and SCC patients was assessed. TAFs were stimulated with TGF-β1 to mimic their activated phenotype, then treated with Nintedanib. Conditioned media from treated TAFs was used to assess endothelial cell migration and vascular network formation. Pro-angiogenic factor secretion was analyzed using antibody arrays, and functional roles of selected proteins were validated in vitro and in vivo using genetic approaches. Results: Nintedanib inhibited the pro-angiogenic effects of ADC-TAF secretome but paradoxically enhanced angiogenesis in SCC-TAFs, increasing endothelial cell migration in vitro and in vivo. Notably, Nintedanib selectively upregulated CXCL-1 secretion in SCC-TAFs. Silencing CXCL-1 using siRNA in SCC-TAFs abrogated the Nintedanib-induced angiogenesis. Similar findings with other multi-angiokinase inhibitors suggest aberrant CXCL-1 production in SCC-TAFs may drive universal resistance to these therapies. Conclusion: SCC-TAFs are resistant to the anti-angiogenic effects of Nintedanib, which paradoxically promotes angiogenesis via CXCL-1 upregulation. This study provides the first mechanistic insight into SCC resistance to anti-angiogenic therapies and underscores the critical role of TAFs in modulating therapeutic responses. Targeting CXCL-1 or TAF-mediated pathways may inform alternative anti-angiogenic strategies in SCC, potentially in combination with immune checkpoint inhibitors. Citation Format: Natalia Isabel Diaz Valdivia, Paula Duch, Alejandro Bernardo, Marselina Arshakyan, Noemi Reguart, Derek Radisky, Oriol Casanovas, Jordi Alcaraz. Intrinsic resistance mechanism of squamous cell lung carcinomas to the multiangiokinase receptor inhibitor Nintedanib through CXCL-1 upregulation in tumor associated fibroblasts [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5554.

Abstract Background: Diffuse pleural mesothelioma (DPM) is a rare and aggressive cancer arising from mesothelial cells of the pleura. Over 2, 500 new cases are diagnosed annually in the United States, mainly attributable to asbestos exposure. The prognosis is poor, and while nivolumab (PD-1 inhibitor) and ipilimumab (CTLA-4 inhibitor) have been approved for first-line treatment of unresectable DPM, the three-year survival rate is only 23%. DPM tumors express high levels of epidermal growth factor receptor (EGFR) and mesenchymal-epithelial transition (MET), but targeted therapies like erlotinib (EGFR tyrosine kinase inhibitors) have shown no clinical benefit. We hypothesized that amivantamab, a bispecific antibody that targets both EGFR and MET, may have activity against DPM through antibody-dependent cellular cytotoxicity (ADCC). Objective: The purpose of this study was to assess the potential of amivantamab to mediate ADCC in tissue culture experiments with DPM cell lines. Methods: We assessed DPM for the expression of EGFR and MET using available data from bulk RNA sequencing. We also examined protein expressions of EGFR and MET by Western blot analysis in five epithelial (EM-Meso, H2452, H2461, Mero-41 and Mero-95), three sarcomatoid (H2373, H2596 and ONE58), and one normal mesothelial (MeT-5A) cell lines. Co-culture experiments were performed with the DPM and normal cell lines and NK cells in the presence of amivantamab to evaluate ADCC. Results: We identified high levels of EGFR and MET expression in both epithelial and sarcomatoid cases of DPM from 183 patients. This finding was corroborated by Western blot analysis of all eight DPM cell lines. In vitro experiments demonstrated that amivantamab induced dose-dependent ADCC in DPM cell lines, with robust NK cell-mediated ADCC observed across all tested DPM lines. Notably, sarcomatoid-type DPM cells were more sensitive to amivantamab-induced ADCC than epithelial-type cells. These findings suggest that amivantamab may offer therapeutic benefits for DPM tumors with high EGFR and MET expression. Conclusion: This study provides preclinical evidence supporting the potential of amivantamab as a novel therapeutic option for DPM, especially in sarcomatoid tumors. Ongoing in vivo studies including in patient-derived xenograft (PDX) models will further evaluate the efficacy of amivantamab in DPM. Citation Format: Shinichiro Suzuki, Lin Yang, Jennifer Ayers-Ringler, Ezequiel J. Tolosa, Kuan-Li Wu, Janet L. Schaefer-Klein, Farhad Kosari, Aaron S. Mansfield. Antitumor activity of amivantamab in diffuse pleural mesothelioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7331.

Abstract Squamous cell carcinoma (SCC) development in mammals is thought to require months or years from the time that initial changes occur. We show that SCCs induced by the mouse papillomavirus MmuPV1, which are generally diagnosed 4-6 months post infection, can be found as soon as 2 weeks post infection. SCCs developed within 2 weeks in immune-competent, adult FVB (FVB/NTac) mice at sites prone to human papillomavirus (HPV)-induced tumors and cancers in human patients: the base of the tongue, the anus, and the skin. The anterior tongue, less susceptible than the base of the tongue to HPV-induced cancer in humans, was resistant to rapid-onset cancer. The presence of invasive cancer in histological tissue sections was verified by multiple pathologists. These diagnoses were corroborated by the results of a classic test of malignancy: skin tumors obtained two weeks post infection grew ectopically as subcutaneous grafts in immune-deficient hosts. Preventative treatment with a novel BRD4 inhibitor dramatically reduced the frequency of ear skin SCCs present 2 weeks post infection and significantly reduced overall disease severity. Like many human papillomaviruses, MmuPV1 lacks an E5 gene. Expression of HPV16 E5 driven by the Keratin 14 promoter in FVB mice (FVB/NTac-Tg(KRT14-HPV16E5*)33Plam/Plam) significantly increased the frequency of rapid-onset invasive cancers caused by MmuPV1 at the base of the tongue and in the anus. In an E5-transgenic mouse with a persistent MmuPV1-induced ear skin lesion, we detected a lymph node metastasis at 38 weeks - further demonstrating a cancer phenotype. Cancers and less severe dysplasia induced by infection at the base of the tongue or the anus spontaneously regressed within 4 to 8 weeks of infection; however, ear lesions persisted. When CD4+ and CD8+ T cells were depleted, lesions at the base of the tongue and anus also persisted, indicating that spontaneous regression involves a T cell-mediated immune response. These results indicate that papillomavirus infection alone is sufficient to induce rapid onset of invasive cancer, and that persistence of disease depends on the tissue-specific immune microenvironment. Citation Format: Andrea Bilger, Ella T. Ward-Shaw, Denis L. Lee, Renee E. King, Shwu-Yuan Wu, Michael A. Newton, Cheng-Ming Chiang, Darya Buehler, Kristina Matkowskyj, John P. Sundberg, Rong Hu, Paul F. Lambert. Rapid-onset squamous cell carcinoma and spontaneous regression following papillomavirus infection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7237.

Pancreatic cancer ranks third in cancer related deaths in the USA, however, no reliable theragnostic biomarker exists for the disease. The community of bacteria within the body can become dysbiotic and promote cancer through inflammation and other means. The objectives of this study were to determine the identity and abundance of bacterial species in the gut of mice before and after development of pancreatic cancer. As there can be significant variation in the genomes of different strains of bacterial species, a metagenome analysis was also performed. We hypothesized that there would be significant variation in species and genes of the gastrointestinal tract of mice before and after xenografting with pancreatic cancer. A decrease in ecological diversity was also predicted. Fifteen female SCID mice were divided into three cages. To develop the SubQ pancreatic cancer model, 5 X 106 Panc1 cells were injected at the flank. Fecal materials were collected prior to tumor cell implantation and after tumor development. DNA was extracted from the fecal material. For phylogenetic analysis the seven variable regions of the 16S rRNA gene were amplified and sequenced. Metagenomic analysis was performed as well. Average tumor size in mice after 40 days was ∼ 32 mm3. Microbial diversity was decreased in two out of the three groups of mice. Principle component analysis showed major differences between all samples. There were at least 15 OTUs in each set of samples specific to pretumor mice and 15 OTUs specific to tumor bearing mice. One genus of bacteria, Alistipes, was specific to each set of tumor bearing mice. Taxa at significantly increased abundances in tumor bearing mice included Prevotellaceae and Porpyromonadaceae. Moreover, there were several gene functions that were enriched in tumor bearing mice. This study produced data that demonstrates that there is a link between health status and the gut microbiome. However, this is not a new idea. This study could serve as foundation to establish a gut microbiota profile of pancreatic cancer. Precision medicine is becoming more common due to the decreasing costs of DNA sequencing. The findings of our investigation strengthens the hypothesis that there is an interaction between gut microbiome and pancreatic cancer development and bacteria or bacterial genes could serve as theragnostic biomarker for pancreatic cancer. Acknowledgement: This research was supported by NIH Grant# T34-GM100831, NSF-NOYCE Graduate student training award, a US Department of Education Graduate student training award to Elizabeth City State University Campus of The University of North Carolina and a University of North Carolina Collaboratory Grant award to Dr. H. Banerjee and Dr. Colby Hunter. Citation Format: Gabrielle Morgan, Colby Hunter, Michael Hall, Narendra Banerjee, Erik Armstrong, Kanisha McNeil, Karington Perry, Santanu Bhattacharya, Hirendra N. Banerjee, Krishna K. Rachamala, Shamit Dutta, Naga M. Nakka. Phylogenetic and metagenomic analysis of the human pancreatic cancer microbiome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6526.

Abstract Breast cancer affects 1 in 8 women and often results from mutations caused by oxidative DNA damage from reactive oxygen species. Two key lesions, 8-hydroxy-adenine (8-OH-Ade) and 4, 6-diamino-5-formamidopyrimidine (FAPY-Ade), exhibit two distinct patterns: FAPY-Ade lesions are predominant in normal tissues, while 8-OH-Ade lesions are abundant in cancerous tissues. These shifts reflect tissue redox status, which may provide a pathway to assess future breast cancer risk. Quantitation of these lesions by chemical detection methods is precise but time-intensive and impractical for high-throughput clinical applications. To address this, monoclonal antibodies have been developed and offer a rapid, scalable, and clinically practical alternative for efficiently detecting and quantifying these biomarkers. This innovation can potentially support personalized breast cancer screening strategies. BALB/c mice were immunized with 8-OH-Ade or FAPY-Ade haptens coupled to Keyhole Limpet Hemocyanin (KLH). After three monthly immunizations, the serums of IgG and IgM titers were evaluated. The mice having the highest IgG titer for each antigen were given an intravenous boost, and the spleens were removed three days later for fusions according to standard methods. Using hapten-conjugated BSA-coated 96-well Pro-Bind plates, the ELISA results demonstrated 30-40 positive wells for each antigen. Up to ten positive wells for each antigen were selected and cloned to yield monoclonal antibody-producing hybridomas. The ELISA of the anti-FAPY-Ade antibody, FA5, showed concentration-dependent reduced binding to immobilized BSA-FAPY-Ade hapten conjugate in increasing FAPY-Ade, confirming its FAPY-Ade binding specificity. Immunohistochemistry using normal and cancerous breast tissue revealed significant staining of normal breast tissue compared to cancerous breast tissue using the FA5 antibody. Conversely, the anti-8-OH-Ade antibody, 8A6, displayed weak staining of normal breast tissue with higher staining of cancerous breast tissues. The results demonstrate that monoclonal antibodies specific for 8-OH-Ade and a novel antibody specific for the ring-opening reactive oxygen product, FAPY-Ade, are practical and rapid alternatives to chemical detection methods for their analysis. Immunohistochemistry mirrored past results using chemical detection methods for these oxidized bases, with high FAPY-Ade compared to 8-OH-Ade in normal tissues and the opposite result in cancer-derived tissues. The potential for high-throughput analysis of 8-OH-Ade and FAPY-Ade levels in the DNA of breast tissues may provide information on future breast cancer risk. Further research is underway to evaluate this hypothesis. Citation Format: Zenzeale A. Hudson, Sarah McLaughlin, Alfredo A. Paredes, Gary K. Ostrander, Eric H. Holmes. Monoclonal antibody evaluation of oxidative DNA base damage in normal and neoplastic breast tissues: development of antibodies specific for 8-OH-adenine and 4, 6-diamino-5-formamidopyrimidine (FAPY-adenine) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7063.

Abstract Approximately 80% of breast cancers express estrogen receptor alpha (ERα, ESR1), which drives cancer progression. Despite the array of targeted therapies, ERα+ breast cancer exhibits the highest recurrence and late distant relapse rates among subtypes. (Z)-Endoxifen (ENDX), an active metabolite of tamoxifen, is a potent, orally bioavailable selective estrogen receptor modulator with a favorable safety profile now under development for breast health applications. Its synthesis yields endoxifen-related byproducts that are new chemical entities (NCEs), including AT416E, AT416Z, AT402E and AT402Z. This study compared the anti-estrogenic effects of these compounds to ENDX in ERα+ breast cancer cell lines (MCF7, T47D, UCD12), including those with ESR1 mutations (Y537S, D538G) linked to endocrine resistance. In 2D cultures with estrogen (E2), ENDX and all NCEs showed significant anti-proliferative effects across all lines, achieving 20-90% inhibition at 0.02-5 μM (p<0.0001). ENDX was the most effective in MCF7; ENDX, AT416Z, and AT402Z in T47D; and ENDX, AT416E, and AT416Z in UCD12. Under E2-deficient conditions, all NCEs had lower IC50 values than ENDX in MCF7 cells, while AT416Z and AT402Z outperformed ENDX in UCD12. In T47D D538G mutants, ENDX, AT416E, and AT402Z exhibited the highest potency, while all NCEs showed comparable efficacy in Y537S mutants. In MCF7 cells, ENDX, AT402E, and AT402Z demonstrated similar activity against Y537S mutants, while AT402Z was the most potent in D538G mutants. In 3D cultures at a clinically relevant concentration (∼1 μM), all compounds significantly reduced proliferation (∼75-90%; p<0.0001), with ENDX and AT402Z showing superior potency. AT416E outperformed ENDX in wound-healing assays (p<0.0001). All NCEs induced comparable G1 cell cycle arrest in MCF7 (p<0.0001), while AT402E and AT402Z showed stronger promotion of G1-to-S phase arrest and S/G2/M inhibition compared to ENDX in T47D cells (p<0.05). These NCEs also significantly induced apoptosis in MCF7 cells at 1-5 μM (p<0.0001), with AT416Z, AT402E, and AT402Z surpassing ENDX (p<0.0001). In T47D cells, apoptosis was significantly induced by ENDX, AT416E, and AT416Z (p<0.0001 vs. vehicle), with ENDX showing superior activity. RT-qPCR analysis at 0.1 μM revealed ENDX and AT416Z as the most potent inhibitors of E2-induced ERα target gene expression in MCF7 cells, with ENDX surpassing all NCEs in T47D cells. Synergy assays indicated that the NCEs do not synergize with ENDX, supporting their potential use as stand-alone agents. These findings highlight four NCEs with notable anti-cancer activity, with some matching or surpassing ENDX in several cancer-relevant cellular readouts, marking them as novel candidates for ERα+ breast cancer therapy. Further studies will advance one or more of these NCEs into in vivo models to confirm their therapeutic efficacy and potential for clinical application. Citation Format: Lena Batoon, Rajeev S. Muthyala, Sandra S. Hammer, Natalie Farris, Steven C. Quay, John R. Hawse. Novel (Z)-endoxifen-related new chemical entities exhibit potent anti-cancer activity in ERα+ breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4742.

Abstract Hepatocellular carcinoma (HCC) mortality is increasing globally, partly due to the growing prevalence of non-viral liver diseases. Genome-wide association studies (GWAS) have identified genetic variants associated with HCC development. Leveraging GWAS summary statistics and linkage disequilibrium score regression (LDSR), we investigated disease co-development with non-viral HCC to provide unique insights into HCC etiology and prioritize relationships for further causal inquiry. We utilized the LDSR statistical framework to estimate the genetic correlation and heritability between non-viral HCC with 923 epidemiologic, behavioral, and clinical traits from the United Kingdom Biobank (UKBB) cohort. We observed a SNP-array heritability (h2 obs) of 0.20 for non-viral HCC. Next, we estimated the pairwise genetic correlation between non-viral HCC and heritable (h2 obs > 0.02) UKBB traits, identifying significant positive genetic correlations between non-viral HCC and blood-based biomarkers of liver injury (ALT, GGT) and allostatic load (including glycated hemoglobin, blood pressure, and total albumin). We also identified a positive genetic correlation between non-viral HCC and diseases associated with metabolic dysfunction-associated steatotic liver disease (MASLD), including diabetes, hypertension, chronic ischemic heart disease, and others. Non-viral HCC was negatively associated with filtered coffee intake but positively associated with instant coffee intake. In addition, smoking behavior and the amount of alcohol consumption in the UKBB cohort showed a positive association with non-viral HCC. Our study provides an atlas of cross-trait genetic correlations in non-viral HCC. Taken together, our results demonstrate possible polygenic pleiotropy between non-viral HCC and different phenotypic traits. The results support further exploration of the predictive power of blood-based biomarkers and selected behaviors for inferring HCC development among non-viral individuals. Citation Format: Younghun Han, Vikram Shaw, Jinyoung Byun, Rikita Hatia, Katherine McGlynn, Lewis Roberts, Manal Hassan, Christopher I. Amos. Linkage disequilibrium score regression identifies genetic correlations between hepatocellular carcinoma and clinically relevant traits [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2284.

Observational studies have demonstrated that aspirin and/or PTSG2 inhibitor (e.g., celecoxib) use, before or after colon cancer (CC) diagnosis, is associated with a lower risk of recurrence. However, a randomized trial of adjuvant celecoxib therapy in stage III CC did not show improved survival compared to placebo treatment. We therefore hypothesized that immune microenvironment features may identify patient subgroups with stronger survival benefits from anti-inflammatory celecoxib treatment. Drawing from an NCI-Intergroup trial (Cancer and Leukemia Group B [now Alliance for Clinical Trials in Oncology]/SWOG 80702) for patients with stage III resected CC that compared 3 versus 6 months of adjuvant fluorouracil, leucovorin, and oxaliplatin ± 3 years of celecoxib, we conducted tissue-based tumor microenvironment profiling on 1,098 resected tumors using a custom 9-plex, T-cell multiplex immunofluorescence assay coupled with digital image analysis and supervised machine learning. The Kaplan-Meier method was used to describe disease-free survival, based on T cell density, and log-rank testing was performed. Cox proportional hazards models were used to examine unadjusted and adjusted associations between T cell density and disease-free survival (DFS). Two-sided P values ≤0.05 were considered statistically significant. We found that higher CD3+ T cell density was associated with longer DFS with an adjusted hazard ratio (HR) of 0.69 (95% confidence interval [CI] 0.52-0.91) for patients with densities in the highest tertile as compared the lowest tertile (P=0.01). T cell subset analysis showed that stromal regulatory T helper cells (HR 0.42 comparing highest to lowest tertile, CI 0.31-0.58, P<0.001) and stromal memory cytotoxic T cells (HR 0.55 comparing highest to lowest tertile, CI 0.42-0.74, P<0.001) were most strongly associated with longer DFS. Compared to the placebo group, adjuvant celecoxib use associated with improved DFS for patients with low but not high stromal CD3+ T cell density (adjusted HRs of 0.45 [CI 0.31-0.65] for lowest tertile and 1.13 [CI 0.73-1.74] for highest tertile; Pinteraction=0.01). This association was also seen when the analysis was restricted to microsatellite stable tumors (HR 0.50 [CI 0.32-0.80] for lowest density tertile and 1.50 [CI 0.87-2.59] for highest density tertile; Pinteraction=0.01). T cell subset analysis showed that DFS benefit for celecoxib use most strongly associated with stromal memory helper T cell density (HR 0.38 [CI 0.26-0.56] for lowest density tertile and 1.26 [CI 0.80-1.97] for highest density tertile; Pinteraction<0.001). Together, these results show that while higher T cell infiltration is associated with improved DFS, lower T cell infiltration is associated with improved DFS for patients treated with celecoxib. This unexpected finding may inform immunomodulatory treatment strategies for CC. Citation Format: Yasutoshi Takashima, Chao Ma, Qian Shi, Andressa Dias Costa, Tyler Twombly, Juha P. Väyrynen, Melissa Zhao, Ardaman Shergill, Pankaj Kumar, Felix Couture, Philip Kuebler, Smitha Krishnamurthi, Benjamin Tan, Eileen M. O'Reilly, Anthony F. Shields, Shuji Ogino, Charles S. Fuchs, Jeffrey A. Meyerhardt, Jonathan A. Nowak. Prognostic and predictive role of T cell infiltration in stage III colon cancer (CC) treated with celecoxib: CALGB/SWOG 80702 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5858.

Breast and ovarian cancer represent the most common and lethal cancers in women. Despite chemotherapy and targeted agents, recurrence is frequent and outcomes for patients with advanced disease are dismal, underscoring the need for novel therapeutic strategies. Using an unbiased approach with DepMap CRISPR screens across more than 1000 cancer cell lines, we identified six genes (CTPS1, RHOA, PRKRA, RAD9A, HUS1, and RAD1) that were essential for triple negative breast cancer (TNBC) cells, but not ERα+ breast cancer cells, that were also associated with poor outcomes. Validation of these effects by siRNA knockdown studies revealed that suppression of CTPS1 elicited the most potent inhibitory effects, characterized by S phase cell cycle arrest and apoptosis. Similar effects were also found in ovarian cancer cells which share many molecular features. CTPS1, along with CTPS2, is responsible for enzymatic conversion of UTP to CTP, an essential nucleoside triphosphate required for multiple cellular processes including DNA/RNA synthesis, phospholipid synthesis and glycosylation. CTPS1 was found to be highly expressed in tumor tissues, for both breast and ovarian cancer. RT-PCR assessment of CTPS1 expression in a panel of over 40 breast and ovarian cancer lines confirmed variable, but elevated expression relative to normal models. Notably, chemotherapy-resistant and PARP inhibitor (PARPi)-resistant models exhibited the highest levels of CTPS1 expression among all cells analyzed. In partnership with Step Pharma, which has developed a first-in-class highly selective CTPS1 inhibitor (STP938), we have demonstrated that STP938 elicits potent anti-proliferative effects at nM concentrations across many cell line and patient derived models, both in 2D and 3D culture systems. Furthermore, STP938 exhibited strong anti-cancer activity in chemotherapy- and PARPi-resistant ovarian cancer models, both as a monotherapy and in combination with standard-of-care treatments. RNA-seq analysis in cell models following STP938 treatment or CTPS1 knockdown/knockout showed significant dysregulation of DNA replication and cell cycle related genes, suggesting potential synergy with drugs targeting replication stress or DNA damage response pathways. Single-cell sequencing further revealed specific cell-cell interactions/cross talk/pathways that are disrupted by STP938 in vivo using ovarian cancer patient-derived xenograft. The first-in-human clinical trial for STP938 began in 2022 for relapsed/refractory B- and T-cell lymphomas (NCT05463263). In August 2024, a second phase I trial opened for advanced solid tumors, including an expansion cohort for CTPS2-null ovarian cancer (NCT06297525) at Mayo Clinic. These laboratory discoveries, and ongoing clinical studies, are expected to pave the way for a broader uptake of this novel therapeutic approach aimed at improving outcomes in patients with highly aggressive forms of these diseases. Citation Format: Xiyin Wang, Michael Emch, Lauren Voll, Rebecca Epp, Esther Rodman, Noa Odell, Nicole Pearson, Xiaonan Hou, Matthew Goetz, Scott Kaufmann, John Weroha, Philip Beer, John Hawse. Exploiting CTPS1 dependency for the treatment of breast and ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6388.

Abstract Background: The tumor microenvironment (TME) plays a pivotal role in ovarian cancer (OC) progression and prognosis, while also contributing to biological differences between primary and metastatic tumors. This study examines OC TMEs to uncover associations with clinical prognosis and tumor biology by analyzing paired primary and metastatic tumors from the same patients. Methods: This study focused on 57 stage-IIIC/IV OC patients treated at the Mayo Clinic, all of whom underwent surgical debulking and adjuvant therapies. All patients provided informed consent, and the study received IRB approval. A tissue microarray (TMA) was specifically designed and constructed, containing paired primary and metastatic tissue cores from the same patients. 147 tissue cores from the constructed TMA were profiled using a 35-antibody panel, with 17 antibodies for cell phenotyping and 18 for functional state characterization. Profiling was performed using Cell DIVE, a multiplexed imaging solution at the single-cell level developed by Leica Microsystems. After cell segmentations and classifications, detailed spatial niche and regional analyses were performed at the single-cell level. Immune cell populations, as well as their TME distributions, were characterized to conduct prognostic associations and contrast primary and metastasis differences. Results: A total of 1,292,988 total cells (after excluding low-density signals and ambiguous classifications) were analyzed, with a median of 8,033 cells per core. Using spatial neighborhood analysis, 12 distinct spatial niches were identified with specific cell types and/or unique functional states. Among these, a macrophage- and fibroblast-enriched niche at the tumor-stroma interface was associated with improved overall survival (p=0.008). Another rare Treg-enriched niche, along with other immune cells, demonstrated protective effects, particularly in patients with residual macroscopic tumors after debulking surgery (p=0.05), while no significant effects were observed in those without visible residual disease (p=0.8). When comparing primary and metastatic tumors, metastatic lesions exhibited significantly increased tumor mass (p=2e-4) and a notable reduction in helper T cells (p=0.04) in tumor areas, as well as significantly increased fibroblast cells in tumor-stroma interfacing regions (p=0.002), reflecting distinct immunosuppressive and tumor-promoting shifts. Conclusion: This study reveals spatial and cellular differences in primary and metastatic OC TMEs, distribution differences linked to prognosis and surgical outcomes. These findings emphasize the potential of spatially resolved TME profiling to refine adjuvant and neoadjuvant therapeutic strategies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6266.