Life Sciences Research Articles

Abstract 5201: Characterization of PDLIM2 in non-small cell lung cancer

Abstract Introduction: PDZ and LIM domain protein 2 (PDLIM2) acts as tumor suppressor by downregulating NF-κB and STAT3 signaling, modulating inflammation, immune response, and cell survival. Mouse model studies have demonstrated that downregulation of PDLIM2 leads to PD-1 immune blockade and chemotherapy resistance. We present a large clinical and molecular characterization of PDLIM2 expression in non-small cell lung cancer (NSCLC). Methods: NextGen sequencing of DNA (whole exome)/RNA (whole transcriptome) was performed for NSCLC (Total N = 29126; Adenocarcinoma [-A, N = 15765], Squamous [-S, N = 6416]) patient tumors submitted to Caris Life Sciences (Phoenix, AZ). Mutations were defined as pathogenic SNVs/indels. Samples were stratified by PDLIM2 expression quartiles (in transcripts per million [TPM]) for all NSCLC tumors (Q4: H, Q1: L). PD-L1 expression [22C3; Positive (+): tumor proportion score (TPS) ≥1%]was assessed by IHC. High tumor mutational burden (TMB-high) set as ≥10 mutations per Mb. Cell infiltration in the tumor microenvironment was estimated by QuantiSEQ. Gene expression profiles were analyzed for transcriptional signatures predictive of response to immunotherapy (T cell-inflamed). Real-world overall survival was assessed from insurance claims data and Kaplan-Meier estimates were calculated for molecularly defined subpopulations. Mann-Whitney U and X2/Fisher-Exact tests were applied where appropriate, with P-values adjusted for multiple comparisons (p < 0.05). Results: Median PDLIM2 expression was higher in NSCLC-A compared to NSCLC-S (15.3 vs 13.6 TPM, p < 0.001). PDLIM2-AL had a lower prevalence of EGFR mutations than PDLIM2-AH (15.5% vs 19.4%, p < 0.01), but the opposite pattern was observed for STK11 (21.5% vs 11.2%), RB1 (7.5% vs 3.8%), TP53 (62.9% vs 52.8%), KEAP1 (18.4% vs 10.9%), and SMARCA4 (9.15 vs 4.5%) alterations (all p < 0.001). PDLIM2-AL tumors had decreased PD-L1+ (L: 52.4% vs H: 58.4% p < 0.001) but increased TMB-High status (L: 40.9% vs H: 28.5%, p < 0.001). PD-L1+ and TMB-H frequency did not vary with PDLIM2 expression in NSCLC-S. In both NSCLC-A and -S tumors, PDLIM2H had increased infiltration of NK cells, macrophages, dendritic cells, T cells, neutrophils, monocytes, and B cells compared to PDLIM2L (all p < 0.05), in addition to increased T cell-inflammed scores (p < 0.001). PDLIM2-AH was associated with improved overall survival (OS) (median 21.7 vs L: 15.6 months; p < 0.001; Hazard ratio [HR] = 0.825, 95% Confidence Interval [CI] 0.765 - 0.889) and time on pembrolizumab treatment (median 6.2 vs L: 5.6 months; p = 0.009; HR = 0.825 95% CI 0.714 - 0.953). No difference in OS was observed between PDLIM2-SH vs L tumors (H: 14.8 vs L: 14.0 months; p = 0.37; HR = 0.95, CI 0.85-1.06). Conclusions: NSCLC-A with high PDLIM2 expression have a unique mutational profile, increased immune cell infiltration and favorable OS. Therapeutic strategies for targeting PDLIM2 to modulate NF-κB and STAT3 signaling should be further explored. Citation Format: Karam Ashouri, Harris Krause, Andrew Elliott, Stephen V. Liu, Patrick C. Ma, Balazs Halmos, Zhaoxia Qu, Gutian Xiao, Ari Vanderwalde, Jorge J. Nieva. Characterization of PDLIM2 in non-small cell lung 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 5201.

Abstract 1249: cBioPortal for Cancer Genomics

Abstract cBioPortal for Cancer Genomics is an open-source platform for interactive, exploratory analysis of large-scale clinico-genomic data. cBioPortal provides a suite of user-friendly visualizations and analyses, including OncoPrints, mutation lollipop plots, variant interpretation, group comparison, survival analysis, expression correlation analysis, alteration enrichment analysis, cohort and patient-level visualization. The public site (https://www.cbioportal.org) is accessed by >35,000 unique visitors each month and hosts data from >390 studies spanning individual labs and large consortia. All data is available in the cBioPortal Datahub: https://github.com/cBioPortal/datahub; in 2023 we added 35 studies (~16,000 samples). In addition, >86 instances of cBioPortal are installed at academic institutions and companies worldwide. We also host a dedicated instance for AACR Project GENIE, enabling access to the GENIE cohort of >197,000 clinically sequenced samples from 19 institutions (https://genie.cbioportal.org). The GENIE Biopharma Collaborative (BPC) enables the collection of comprehensive clinical annotations including response, outcome, and treatment history, which can all be visualized in cBioPortal. BPC cohorts for non-small cell lung cancer (~2,000 samples) and colorectal cancer (~1,500 samples) are available, with more cancer types to come. This past year, we significantly enhanced existing features. Group comparison now includes a Mutations tab showing a mirrored lollipop plot and outcomes analysis now supports hazard ratios and landmark analysis. Arm-level copy number can now be compared across groups, as can any data in the generic assay format. We also enhanced the study view. Users can now add charts to summarize the copy number of a specific gene across the cohort. The option to add custom data charts now supports numerical data as well as categorical data. Filters in study view can now be manually submitted, which provides a performance improvement when applying multiple filters in a large cohort. Larger-scale performance improvements are currently underway. We continue to improve support for multimodal datasets incorporating derived data elements, including cell type counts and fractions per sample from imaging or single cell data. The MSK-SPECTRUM ovarian cancer study has samples profiled with bulk sequencing, scRNASeq, H&E and mpIF imaging. Through integrations with CELLxGENE (single cell data) and Minerva (imaging), users can explore these data modalities in detail in isolation and query them jointly in cBioPortal. cBioPortal is open source (https://github.com/cBioPortal/). Development is a collaborative effort among groups at Memorial Sloan Kettering Cancer Center, Dana-Farber Cancer Institute, Childrens Hospital of Philadelphia, Princess Margaret Cancer Centre, Caris Life Sciences, Bilkent University and The Hyve. We welcome open source contributions from others in the cancer research community. Citation Format: Tali Mazor, Ino de Bruijn, Rima AlHamad, Calla Chennault, Corey Dubin, Jeremy Easton-Marks, Zhaoyuan Fu, Benjamin Gross, Charles Haynes, David M. Higgins, Jason Hwee, Prasanna K. Jagannathan, Mirella Kalafati, Karthik Kalletla, James Ko, Tim Kuijpers, Sowmiyaa Kumar, Priti Kumari, Ritika Kundra, Bryan Lai, Xiang Li, James Lindsay, Aaron Lisman, Qi-Xuan Lu, Ramyasree Madupuri, Angelica Ochoa, Yusuf Z. Özgül, Oleguer Plantalech, Matthijs N. Pon, Baby A. Satravada, Jessica Singh, Selcuk Onur Sumer, Pim van Nierop, Floris Vleugels, Avery Wang, Manda Wilson, Hongxin Zhang, Gaofei Zhao, Ugur Dogrusoz, Allison Heath, Adam Resnick, Trevor J. Pugh, Chris Sander, Ethan Cerami, Jianjiong Gao, Nikolaus Schultz. cBioPortal for Cancer Genomics [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 1249.

Abstract 2567: An analysis of proviral insertion site of Moloney murine leukemia virus, PIM1, kinase expression and clinical outcomes in renal cell carcinoma

Abstract Background: Wild-type Proviral Insertion Site of Moloney Murine Leukemia Virus 1 Kinase (PIM1) is a constitutively active serine/threonine kinase that inhibits apoptosis and promotes cell cycle progression, proliferation, angiogenesis, invasion, and migration. Due to constitutive activation, PIM1 is regulated at transcription and degradation. We have previously shown PIM1 protein levels are elevated in clear cell renal cell carcinoma (ccRCC) compared to normal kidney and that PIM1 inhibition induces tumor regression in pre-clinical models. Current approved therapies for ccRCC do not inhibit PIM1. We hypothesized that high PIM1 expression will adversely affect clinical outcomes in ccRCC. Methods: Whole transcriptome sequencing (WTS) data in the Caris Life Sciences database were examined for PIM1expression and overall survival (OS; defined as time from diagnosis to last contact or death). We assessed PIM1 in primary and metastatic ccRCC, and the impact on survival in ccRCC and papillary RCC. Differentially expressed genes (DEGs) associated with high PIM1 were also determined. Results: PIM1, in the highest quintile, was associated with a shorter OS (869 vs 1738 days; HR 2.629; CI 1.495 - 4.611; p<0.001; N=147) in ccRCC. PIM1 is enriched in metastatic (median 7.62 transcripts per million (TPM), CI 6.35 - 9.26) vs primary (median 5.07 TPM; CI 4.28 - 6.45; p=0.0025) RCC. Upregulated DEGs associated with high PIM1 are involved in the extracellular matrix. IL-6 expression, known to correlate with worse outcomes, was 6.5-fold higher in PIM1 high vs PIM1 low (13.2 vs 2.1 TPM, p<0.0001) RCC. High PIM1 was associated with poor survival following checkpoint inhibition (HR 3.714, 95% CI 1.339 - 10.303, p=0.007) and a trend towards poorer survival following mTOR inhibition and VEGF inhibition. PIM1 did not affect survival in papillary RCC. Conclusions: Real-word data show that high PIM1 correlates with poorer survival in ccRCC but not in papillary RCC. This effect likely persists independent of treatment received. PIM1 enrichment in metastatic ccRCC, and associated DEGs, suggest that PIM1 influences the extracellular matrix to promote metastases. The correlation between IL-6expression and PIM1 supports our prior data that an IL-6/JAK/STAT/PIM1 pathway is involved in ccRCC. Citation Format: Sheldon L. Holder, Galina Lagos, Benedito A. Carneiro, Anthony Mega, Andre De Souza, Rana R. McKay, Andrew Elliott, Chadi Nabhan, Stephanie L. Graff. An analysis of proviral insertion site of Moloney murine leukemia virus, PIM1, kinase expression and clinical outcomes in 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 2567.

Natural occurrence of tomato chlorosis virus on tomatillo (Physalis philadelphica) in the United States.

Tomatillo (Physalis philadelphica L.) is an annual plant native to Mexico and Guatemala, and cultivated in other tropical and subtropical regions. In October 2023, tomatillo plants with interveinal yellowing of leaves, marginal chlorosis, leaf thickening, and leaf rolling symptoms (Figure 1) were observed at Colquitt and Tift County, Georgia, US. The disease incidence ranged from 80-100 % which reduced fruit quality and marketability. Twenty tomatillo leaves exhibiting severe symptoms were collected, and, sub-sampled of the leaves were pooled into microcentrifuge tubes. Further, MagMAX 96 viral RNA isolation kit (Thermo Fisher Scientific, US), was used for the extraction of (n=4) total nucleic acid (TNA) (Kavalappara et al. 2021). Symptomatic leaves were tested for the presence of insect-transmitted viruses such as begomovirus (tomato yellow leaf curl virus, TYLCV), potyvirus (turnip mosaic virus, TuMV), crinivirus (tomato infectious chlorosis virus, TICV; tomato chlorosis virus, ToCV), and tospovirus (orthotospovirus tomatomaculae, TSWV). Polymerase chain reaction (PCR) was performed for detecting TYLCV, using gene-specific primers (Kumar et al., 2023). However, for ToCV, TuMV and TICV detection, cDNA was prepared using 100 ng of TNA as a template, followed by the PCR ( Liu et al., 2012). Moreover, the detection of TSWV was conducted using immuno-strips (Adgia, US) following the manufacturer's instructions. ToCV was detected from all the tested samples, while TuMV, TICV, TYLCV and TSWV were not detected in any symptomatic tissues. In addition, RT-PCR was performed using gene-specific primers targeting the RNA-dependent RNA polymerase (RdRP) gene and the heat-shock protein 70 (Hsp70) gene of ToCV. The PCR amplicon of 439 bp encoding Hsp70 and 643 bp corresponding to RdRP was gel-purified and Sanger sequenced (Azenta Life Sciences, US). BLASTn analysis shows RdRP gene from ToCV-tomatillo (OR905600) has 100 % identity with ToCV of RNA1 segment (RdRP, GenBank accession no. AY903447, Florida, US), while Hsp70 gene (OR900219) has 100 % identity with ToCV of RNA2 segment (Hsp70, GenBank accession no. LC778246, Cairo, Egypt). In addition, the symptomatic tomatillo leaves were studied for transmission assay using tomato, employing non-viruliferous whiteflies (Bemisia tabaci) with 48 h of acquisition access period. Further, two weeks post-infection, the presence of ToCV was detected from the test plants while other whitefly-transmitted viruses remins undetected. In 2023, ToCV is widespread in tomato-growing counties, infecting commercially grown tomato cultivars with intermediate resistance against TYLCV-IL (Israel strain). However, tomatillo plants infected with TuMV in California (Liu et al., 2012), TSWV in Georgia, (Díaz-Pérez and Pappu 2000) and TYLCV in Mexico (Gámez-Jiménez et al. 2009) were reported. This study suggests that tomatillo could be a permissive host for ToCV while restrictive to other prevalent viruses in the region. A recent investigation speculates a potential synergistic interaction between ToCV and TYLCV-IL, exacerbating the breakdown of host resistance in tomato (Fiallo-Olivé et al. 2019, Kumar et al. 2023). To the best of our knowledge, this is the first report for the natural incidence of ToCV on tomatillo within the US. The findings will contribute to developing more effective management strategies against emerging viral threats.

Abstract 317: Using the Biomek NGeniuS Next Generation Library Prep System to automate the Illumina TruSight Oncology 500 assay

Abstract Cancer researchers using next generation sequencing (NGS) need reliable sample preparation methodologies that generate high-quality, sequence-ready libraries from often-challenging samples. This process can be aided using automated liquid handling systems. The Biomek NGeniuS Next Generation Library Prep System from Beckman Coulter Life Sciences is a reliable, easy-to-use, purpose-built liquid handler for NGS library preparation ideal for laboratories requiring more flexibility and less hands-on time. The user-friendly software requires no programming skills, making it easy to set up and run. Biomek NGeniuS Next Generation Library Prep System’s hardware features include an on-deck thermal cycler, gripper, temperature-controlled reagent storage, and high-density tip storage which reduces user interventions and enables more walkaway time. An extensive and growing library of complimentary demonstrated NGS sample prep protocols allows the user to select from a variety of applications for NGS workflows. One such workflow is Illumina TruSight Oncology 500 assay, which represents a comprehensive NGS assay targeting the full coding regions of 523 genes implicated in the pathogenesis of solid tumors. Using enrichment-based library preparation techniques for use with formalin-fixed, paraffin-embedded (FFPE) samples, the Illumina TruSight Oncology 500 assay, when running the DNA Only workflow, can detect single nucleotide variants (SNVs), indels, amplifications, and multinucleotide variants (MNVs) in a single sequencing run. The Illumina TruSight Oncology 500 assay also detects immunotherapy biomarkers for tumor mutational burden (TMB) and microsatellite instability (MSI) in DNA samples. The Illumina TruSight Oncology 500 assay also offers a combined RNA/DNA workflow that also allows for the assessment of fusion events. In this poster, we show the utility of the Biomek NGeniuS Next Generation Library Prep System to perform the Illumina TruSight Oncology 500 assay, which saves researchers valuable hands-on time and offers a highly streamlined setup process for users new to liquid handling automation. Confidential - Company Proprietary Citation Format: Zach Smith. Using the Biomek NGeniuS Next Generation Library Prep System to automate the Illumina TruSight Oncology 500 assay [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 317.

Abstract 3890: Comprehensive molecular and immunological characterization of early-onset esophagogastric cancer

Abstract Background: The incidence of early-onset esophagogastric cancer (EOEGC), defined as age of diagnosis <50, has increased in recent decades. Compared to average-onset esophagogastric cancer (AOEGC), EOEGC are more likely to be genomically stable, have diffuse histology, and present with more advanced disease. Here, we utilized a large real-world data cohort to characterize EOEGC with molecular and immune signatures. Methods: 5,175 esophagogastric cancer (EGC) samples (<50 yrs, n=530; ≥50 yrs, n=4645) were tested by next generation sequencing (NGS) (592, NextSeq; WES, NovaSeq) and whole transcriptome sequencing (WTS) (NovaSeq) (Caris Life Sciences, Phoenix, AZ). EGC patients with age <50 and ≥50 were classified as EOEGC and AOEGC, respectively. Deficient mismatch repair (dMMR)/microsatellite-instability (MSI) was tested by IHC and NGS. Tumor mutational burden (TMB) totaled somatic mutations per tumor (high>10 mt/MB). Pathway enrichment determined by Gene Set Enrichment Analysis (Broad Institute, Cambridge, MA). Immune cell fractions were calculated by deconvolution of WTS: Quantiseq. Statistical significance was determined using chi-square and Mann-Whitney U test with p-values adjusted for multiple comparisons (q < 0.05). Results: EOEGC had higher frequency of CDH1 (18.9% vs 6.5%) but lower frequency of TP53 (65.9% vs 74.4%) mutations compared to AOEGC (all q<0.05). EOEGC had higher frequency of CCNE1 (6.8% vs 4.4%), MYC (6.1% vs 4.1%) and FGFR2 (3.45% vs 1.87%) copy number alterations and ARHGAP26 (5.67% vs 2.2%) fusion. EOEGC had lower frequency of TMB high (3.8 vs 10.1%) and dMMR/MSI-H (1.3% vs 5.7%) (all p<0.05). EOEGC had pathway enrichment of epithelial mesenchymal transition (EMT) (NES: 1.8, FDR: 0.0) and angiogenesis (NES: 1.3, FDR: 0.24). EOEGC had lower median MAPK pathway activity score (MPAS) (-0.16 vs 0.13, p<0.05). Analysis of inferred immune cell infiltrates showed that EOEGC had increased infiltration of B cells (4.7% vs 3.9%) and M2 macrophages (3.6% vs 3%) but decreased infiltration of M1 macrophages (3.7% vs 4%) (all q<0.05). EOEGC had increased T cell inflamed signature (31.3% vs 26.1%, p<0.05) and increased expression of HAVCR2 (TIM3) but decreased expression of FOXP3 and CD274 (PD-L1) (FC: 0.84-1.09, all p<0.05). There was no difference in LAG3, CTLA4, and IDO1 gene expression. Conclusion: EOEGC has several unique molecular and tumor immune microenvironment features. In this real world cohort, EOEGC has increased frequency of CDH1 mutations, ARHGAP26 fusions, enrichment of EMT and angiogenesis pathways, decreased MAPK pathway activity, decreased frequency of TMB high and dMMR/MSI-H, and a unique immune cell infiltrate with decreased M1 macrophages and increased M2 macrophages. These unique differential characteristics present therapeutic opportunities but also demonstrate the limitations of currently approved therapies in this subset of patients. Citation Format: Lawrence W. Wu, Sachin K. Deshmukh, Sharon Wu, Joanne Xiu, Alex Farrell, Vincent Lam, Emil Lou, Sanjay Goel, Chadi Nabhan, Ryan H. Moy. Comprehensive molecular and immunological characterization of early-onset esophagogastric 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 3890.

Abstract 7092: Cancer biomarker discovery with scalable biofluid proteomics

Abstract Biofluid proteomics opens opportunities for the discovery of early detection, diagnostic, and prognostic cancer biomarkers through non-invasive or minimally invasive screening. Plasma and serum are accessible sample types, however, the high dynamic range of proteins creates analytical challenges for the sensitive detection of potential biomarkers. In this work, we compare two proteomic approaches for large-scale cancer biomarker discovery: the Olink Proximity Extension Assay and the Seer Proteograph Product Suite. Olink Explore 3072 is an immunoassay that gains specificity by using dual antibody recognition to measure approx. 3,000 proteins in each sample. Antibody pairs are tagged with oligonucleotides, which hybridize and are amplified before measurement by next generation sequencing. Seer Proteograph is an enrichment strategy that uses a proprietary mix of functionalized magnetic nanoparticles with affinity for certain amino acids and functional groups in native state proteins. The resulting peptides are measured using untargeted mass spectrometry to identify proteins at the peptide level, paving the way for the identification of proteoforms and peptide variants. Aliquots of human plasma from 78 samples representing a range of cancer indications were obtained through the Discovery Life Sciences biobank and used for analysis by both the Olink and Proteograph workflows. Olink analysis achieved Pearson correlation scores of greater than or equal to 0.8 when correlating each of seven process controls to each other for all approx. 3000 proteins. On average, greater than or equal to 75% of the proteins were detected across all panels in all sample groups except for Normal and Benign groups (64% and 74% respectively). After stringent FDR filtering to ensure only high-quality data was analyzed, the Seer Proteograph workflow quantified 4,323 proteins across all 78 samples. Pearson correlation scores of greater than or equal to 0.96 were achieved when comparing digestion controls, process controls, and sample cleanup controls. Both platforms achieved detection of proteins across the dynamic range of plasma, including low abundance proteins. The Olink Explore 3072 and Seer Proteograph platforms were found to be complementary, with approx. 15% overlap in measured proteins. Cancer biomarker discovery at the proteomic level is bolstered by recent advances in analytical capabilities, as revealed by the Olink Explore and Seer Proteograph platforms. Importantly, complementary implementation of these two workflows will enable comprehensive biomarker analysis. Citation Format: Danielle B. Gutierrez, Jessica L. Moore, Jaison Arivalagan, Tiffany Louie, Leslie Wilkinson, Ray Clemens. Cancer biomarker discovery with scalable biofluid proteomics [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 7092.

Abstract 802: Molecular profile of gastric cancer in Hispanics living in Puerto Rico

Abstract Gastric cancer (GC) is the 4th leading cause of cancer death globally. The incidence and mortality rates of GC differ dramatically between racial/ethnic groupings. In the United States (U.S.), Hispanics, non-Hispanic Blacks, and Asian or Pacific Islanders are more likely to be diagnosed with GC and have a higher mortality rate than non-Hispanic Whites. Among the Hispanic population living in Puerto Rico (PRH), GC ranks among the top ten leading causes of cancer death, whereas this malignancy is not a major cause of cancer mortality in the mainland U.S. Tumor profiling approaches have resulted in the discovery of actionable gene alterations, which serve as a guide for treatment strategies and ultimately enhance the overall survival rates of individuals diagnosed with cancer. However, descriptive information regarding the genetic mutational landscape of GC tumors from PRH is limited. This study aims to describe the genomic profile of GC tumors in PRH and to identify the most prevalent genetic mutations. We retrospectively examined GC tumor mutational profiles from 106 PRH that underwent NGS testing from 2015 to 2022 (provided by CARIS Life Sciences and the Precision Oncology Alliance). We compared PRH GC tumor somatic mutation prevalence to TCGA and the AACR Project Genomics Evidence Neoplasia Information Exchange (GENIE), both available through the cBioPortal for Cancer Genomics. Descriptive statistics were performed to characterize the database. Among the top mutated genes for GC tumors in PRH were TP53 (54.1%, n=85), ARID1A (34.2%, n=73), KMT2D (30.5%, n=82), CDH1 (29.4%, n=85) and ZFHX3 (27.3%, n=55). The most frequent gene amplifications were ERBB2 (7.1%, n=84), KRAS (6.0%, n=84), and CCNE1 (4.9%, n=81). Compared to other datasets, PRH had significantly different mutational frequencies of GC driver genes like TP53, ARID1A, CDH1, and KMT2D. This study is the first to report PRH GC tumor mutational profiles and compare the mutational frequencies to other non-Hispanic and U.S. mainland Hispanic populations using TCGA and GENIE data sets. Furthermore, our study provides data on the specific mutational landscape for Hispanics with GC and the implications on therapeutic options and clinical outcomes. Developing new treatments for genomically diverse populations requires understanding the most common carcinogenic molecular pathways that affect Hispanics with GC. Citation Format: Ingrid Montes-Rodriguez, Hilmaris Centeno-Girona, Noridza Rivera, Marcia Cruz-Correa. Molecular profile of gastric cancer in Hispanics living in Puerto Rico [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 802.

Abstract 3362: Describing the molecular landscape of cervical cancer metastases: Implications for future therapeutic targets

Abstract Introduction: Cervical Cancer (CC) is a heterogenous disease with multiple histological sub-types. Early screening has led to a reduction in disease related mortality, but a significant number of patients present with or progress to advanced disease. Studies between the molecular and immune alterations in CC primaries (CCP) and CC metastases (CCM) are needed to explore potential therapeutic strategies. Methods: Relationships of CCM and alterations detected by NGS (592, NextSeq; WES, NovaSeq) were investigated in 2,668 (1,393 primary Cervix samples, 383 local metastatic GYN samples, and 892 distant metastatic samples) CC samples (Caris Life Sciences, Phx, AZ). PD-L1+ was tested by IHC (22c3, >1%). Tumor mutational burden (TMB) was measured by summing somatic mutations per tumor (H: >10 mt/MB). Immune infiltrates estimated by deconvolution of WTS data (NovaSeq) using Quantiseq. Statistical significance determined by chi-square and Mann-Whitney U and adjusted for multiple comparisons (q<0.05). Results: Hierarchical clustering of CCM sites by alterations (mutations, amplifications and fusions) frequency revealed CCM to Liver (n=77) and Lymph Nodes (n=265) as the most distinct from CCP while Vagina/Vulva (V/V) (n=196) and Lung (n=115) CCM were most similar. Lymph Nodes had decreased TP53-mt (6.6% vs 14.7%) and ARID1A-mt (4.53% vs 8.27%) but higher BRCA1-mt (3.03% vs 0.87%) compared to CCP (p<0.05). Liver had higher BRCA1-mt (6.67% vs 0.87%), ASXL1-mt (8.47% vs 2.96%) and APC-mt (4% vs 0.65%) compared to CCP (p<0.05). CCM to Lymph Nodes had higher median TMB (median [range]: 6 [0-101] vs 5 [0-460], q<0.05) and HPV16/18+ rate compared to CCP (81.2% vs 66.8%, p<0.05). CCM to Liver had lower expression of CTLA4 compared to CCP (1.67-fold decrease), similarly, Ovary/FT had significantly lower expression of immune checkpoint (IC) gene expression (CD80, CD86, CD274, PDCD1, PDCD1LG2, CTLA4, HAVCR2, IDO1, LAG3, IFNG: 2.23-4.66-fold decrease) (q<0.05). While Lymph Nodes had higher expression of the IC genes compared to CCP (1.30-1.49-fold, q<0.05). CCM to Liver and Ovary/FT had lower median infiltration of immune cells while Lymph Nodes had higher compared to CCP of Macrophage M1 (2.7% vs 1.03% vs 3.35% vs 3.1%), B cells (3.7% vs 3.5% vs 5.6% vs 4.3%), CD8 T cells (0% vs 0% vs 1.01% vs 0.75%), and Tregs (1.6% vs 0.74% vs 2.86% vs 2.5%) (q<0.05). CCM to Liver and Ovary/FT had lower median IFN score while Lymph Nodes had higher median IFN score (q<0.05) compared to CCP, similarly, Ovary/FT had a lower % of T-cell inflamed tumors with Lymph Nodes having higher T-cell inflamed tumors compared to CCP (7.89% vs 47.4% vs 29.3%, q<0.05). Discussion: CCM to Lymph Nodes and Liver had the most distinct molecular and immune landscape compared to CCP while Ovary/FT had a similar molecular profile but a distinctively cold immune profile compared to CCP. Additional studies to evaluate the potential therapeutic targets are warranted. Citation Format: Matthew J. Hadfield, Sharon Wu, Alex Farrell, Matthew Oberly, Grace Sun, Premal Thaker, Jody Wellcome, Matthew Anderson, Don Dizon. Describing the molecular landscape of cervical cancer metastases: Implications for future therapeutic targets [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 3362.

Abstract 3361: Surfaceome and cancer testis antigen profiling of lung adenocarcinoma by large-scale transcriptomic analysis

Abstract Background: The emergence of improved modalities to target tumor specific and tumor associated antigens in solid tumors (e.g. antibody-drug conjugates, bi-specific antibodies, and adoptive cell therapy) underscores the need to efficiently identify and validate new therapeutic targets. Methods: We examined differential expression of surfaceome genes (Fonseca et al., 2016) and cancer testis (CT) antigen genes (Almeida et al., 2009) among major lung adenocarcinoma genomic subtypes; conducting comprehensive surfaceome/CT antigen profiling of 9002 NSCLC adenocarcinoma samples using data generated by RNA sequencing (whole transcriptome) at Caris Life Sciences (Phoenix, AZ). NSCLC tumors were stratified into 5 subgroups based on the presence of specific driver mutations: ALK fusion (N=325), EGFR mutant (N=1739), KRAS mutant (no STK11 or KEAP1 mutation; N=2561), KRAS + STK or KEAP1 mutation (N=1264), and Pan-WT (no mutation in ALK, EGFR, KRAS, STK11, or KEAP1 detected; N=3113). Fold-change in expression was calculated by dividing the median value for a specific subgroup by the median of all other groups combined. Significance was tested by Mann-Whitney U test, with p-values adjust for multiple hypothesis testing (Benjamini-Hochberg). Results: Surfaceome genes with the highest median expression overall included CD74, TMBIM6, and CD44, while the highest median expression among CT antigens was observed for SPAG9, KIAA0100, and RQCD1, yet the expression of these genes across the 5 driver mutation-based subgroups was similar (fold-changes ranged 0.93-1.06). To identify driver-associated candidate genes with relatively high expression, genes with subgroup-specific median expression >75th-percentile and with significant differences when compared to all other groups were ranked by fold-change. While several candidate surfaceome genes were associated with a specific subgroup, CLDN10 was identified among the top 3 candidates for both ALK fusion and KRAS + STK11/KEAP1 mutant subgroups: ALK fusion (GRIN2A, 3.03-fold; CLDN10, 2.53-fold; SLITRK6, 1.86-fold), EGFR mutant (SFTPC, 3.82-fold; MUC21, 1.99-fold; ATP13A4, 1.60-fold), KRAS mutant (VSIG1, 1.86-fold; AREG, 1.47-fold; C15orf48, 1.25-fold), and KRAS + STK11/KEAP1 mutant (ABCC2, 4.03-fold; MUC13, 3.21-fold; CLDN10, 3.00-fold). The top driver-associated CT antigen candidates included LEMD1 (1.42-fold) in ALK fusion, TMEM108 (1.27-fold) in EGFR mutant, XAGE1D (1.7-fold) in KRAS mutant, and CABYR (1.52-fold) in KRAS + STK11/KEAP1 mutant subgroups. Conclusion: Our analysis revealed several surfaceome and CT antigen candidate genes with relatively high expression across lung adenocarcinoma, as well as differential expression of candidate genes within specific genomic subtypes. These findings can help prioritize the validation and development of novel therapeutic targets in lung adenocarcinoma. Citation Format: Swartz Andrew, Andrew Elliott, George W. Sledge, Stephen V. Liu, Balazs Halmos, Vincent K. Lam, Taofeek K. Owonikoko. Surfaceome and cancer testis antigen profiling of lung adenocarcinoma by large-scale transcriptomic analysis [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 3361.

Abstract 6423: KRAS mutation detection by Sanger sequencing using the SeqStudio Flex capillary electrophoresis instrument

Abstract The detection of mutations in the KRAS gene highly associated with cancer is one of the most widely used Sanger sequencing capillary electrophoresis (CE) applications in molecular diagnostics. In addition to their traditional role in cancer diagnostic, KRAS mutations have recently gained additional relevance as pharmacogenomics (PGx) markers for a significant number of new oncology therapeutics. KRAS mutation detection assays are used as companion diagnostics (CDx) for these cancer treatments. Analysis by Sanger Sequencing of KRAS mutations is cost effective and lends itself for use in resource limited healthcare settings. While there are currently no commercial kits for targeted sequencing of the KRAS mutation hotspots available, the online Applied Biosystems Primer Designer™ Tool (thermofisher.com) allows standardization of the PCR primer design between labs and repeat orders of the KRAS assays by using these commercially available assay ID numbers. Here we demonstrate the use of Primer Designer KRAS mutation detections assays for mutation in codon 12, 13 and 61 totaling to 8 allele variants. The data was generated on the SeqStudio Flex CE instrument using reference standard DNA as well as DNA extracted from FFPE solid tumor samples. We show accuracy of variant call (OPA 100% (64/64)) and concordance (OPA 98.3% (59/60)) in a side-by-side comparison of the SeqStudio Flex with the legacy Model 3500 Genetic Analyzer instrument using a set of commercially available (Horizon Discovery) KRAS reference DNA samples. Primary solid tumor samples are processed to Fresh Frozen Paraffin Embedded samples (FFPE) for further disease marker evaluation. These samples are stable for long term at ambient temperatures and provide the opportunity to evaluate prospective biomarkers at large scale to understand their potential utility. Discovery Life Sciences has generated a large-scale biorepository of human biospecimen, including FFPE, plasma/sera, buffy coat, and PBMCs. Archival FFPE samples from oncology indications associated with KRAS mutations, such as non-small cell lung cancer and colorectal cancer, were analyzed and identified samples with KRAS mutations, demonstrating the potential diagnostic utility of these assays. Finally, cell-free (cf) DNA from double spun plasma from relevant indications was also successfully evaluated for KRAS mutations, highlighting the potential for these assays in liquid biopsy studies. The study demonstrates the robustness of the Primer Designer KRAS mutation detection sequencing assays using reference standard DNA as well as DNA derived from FFPE DNA and cfDNA from patient derived tumor samples. For Research Use Only. Not for use in diagnostic procedures. Citation Format: Achim Karger, Ami Patel, Kaylee Tseng, Camila Ornelas, Mary Napier, Rod Beck, Kerri Collwell, Shawn Fahl, Carole Bornarth. KRAS mutation detection by Sanger sequencing using the SeqStudio Flex capillary electrophoresis instrument [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 6423.

Abstract 6851: The genomic, transcriptomic, and immunologic landscape of TEM8 (ANTXR1) in neuroendocrine neoplasms (NENs)

Abstract Introduction: TEM8 (coded by ANTXR1) is overexpressed in malignant tissues, with oncolytic viruses such as SVV-01 uniquely binding to this receptor on tumor-associated angiogenic endothelial cells, pericytes, fibroblasts, and immune inflammatory cells. Recent pre-clinical data suggest that TEM8-targeting therapies may convert immunologically “cold” tumor microenvironments (TME) into “hot” milieu more amenable to treatment with immune checkpoint inhibitors (ICIs). Methods: NextGen sequencing of DNA (592 genes or WES) and RNA (WTS) was performed on neuroendocrine neoplasms (NENs; N = 1724), excluding small-cell lung cancer, submitted to Caris Life Sciences (Phoenix, AZ). Mutations were defined as pathogenic SNVs/indels. ANTXR1 expression was divided by ANTXR1-expression quartiles (transcripts per million [TPM];Q4:H;Q1:L). PD-L1 expression (SP142; Positive (+): ≥2+,≥%5) was assessed by IHC. High tumor mutational burden (TMB-H) was defined as ≥10 mutations per MB. Cell infiltration in the TME was estimated by QuantiSEQ. Gene expression profiles were analyzed for transcriptional signatures predictive of response to immunotherapy (T cell-inflamed) and MAPK pathway activation score (MPAS). Overall survival (OS) data was obtained from insurance claims, and Kaplan-Meier estimates were calculated for molecularly defined subpopulations of patients. Mann-Whitney U and X2/Fisher-Exact tests were applied where appropriate, with p-values adjusted for multiple comparisons (p<0.05). Results: NENs from the adrenal gland had the highest median ANTXR1 expression (50 TPM), followed by those from the colon and rectum (43.5), genitourinary organs (43.3), and small bowel (excluding appendix) (39.2). The lowest median ANTXR1 expression was observed in NENs from the pancreas (31.5). No pathogenic mutations were significantly associated with ANTXR1H vs L tumors (p>0.05 for all), along with no differences in the prevalence of TMB-H (8.6% vs 12.6%, p=1) or PD-L1+ (5.8%: vs 3.1%, p=1). A greater proportion of B cells (5.34 % vs 3.78%, p<0.001), M1 (1.46% vs 0.15%, p<0.001) and M2 macrophages (3.92% vs 2.67%, p<0.001), and T-regulatory cells (1.77% vs 1.08%, p<0.001) was observed in ANTXR1H TMEs, which were more frequently classified as T cell-inflamed compared to ANTXR1L (42% vs 8%, p<0.001). ANTXR1H tumors also had higher MPAS compared to ANTXR1L (1.16 vs -1.61 arbitrary units, p<0.001). There was no significant difference in median OS between ANTXR1H vs L tumors (18.8 mo. vs 21.2 months, Hazard Ratio:1.18, p=0.114). Conclusion: The increased immune cell infiltrate and prevalence of T-cell inflamed status among ANTXR1H NENs suggests these tumors might be more responsive to treatment with ICIs. As trials incorporating intratumoral injections of SVV-01 in combination with ICIs are underway, further investigation of clinical and molecular associations with ANTXR1 expression in NENs is warranted. Citation Format: Samuel A. Kareff, Harris Krause, Andrew Elliott, Peter Hosein, Emil Lou, Heloisa Soares, Matthew Oberley, Aman Chauhan. The genomic, transcriptomic, and immunologic landscape of TEM8 (ANTXR1) in neuroendocrine neoplasms (NENs) [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 6851.

Abstract 6750: Validation of a platform to develop new dendritic cell vaccines for cancer immunotherapy

Abstract Osteosarcoma is the most common malignant form of bone cancer that generally causes patients to die of pulmonary metastasis. If detected before the tumor has metastasized, osteosarcoma has a 75% survival at 5 years after the conventional therapy with surgery and chemotherapy. However, most osteosarcomas are high-grade tumors that grow quickly and spread before being detected, lowering patient survival to 20%. Our unique protocol using the human platelet lysate PLTMax® allows establishing tumor cell lines without genetically modifying cells and with minimal genetic drift. The use of PLTMax® and allogeneic tumor cell cultures has proven to be a major advantage for the generation of large amounts of rich tumor antigen libraries for dendritic cell (DC) vaccines. This protocol was used in an autologous dendritic cell/allogeneic tumor lysate vaccine developed at the Mayo Clinic and licensed to Mill Creek Life Sciences (MCLS) to treat newly diagnosed glioblastoma (GBM), which normally has a poor survival rate at only 2%. The technology was used in a phase I clinical trial (MC1272) performed at the Mayo Clinic. The clinical outcomes proved over a 10 time increase in survival rate of patients that received the DC vaccine after the conventional therapy, providing over 20% three-year survival rate. Our technology differs from other clinically used DC vaccines in three primary ways: a) Highly efficient cell line establishment: The unique method for generating tumor cell lines uses Mill Creek Life Sciences’ PLTMax®, a media supplement derived from human platelets. PLTMax® more closely mimics the native microenvironment of tumors and is used to reduce genetic drift and senescence. b) High genetic fidelity: GBM tumor cells expanded via PLTMax® maintain genetic fidelity and TAA expression—a feat that has not been previously achieved by any other GBM cell culture method. c) Reduction of treatment delays and inclusion of all patients that qualify for treatment: Because we will use allogeneic tumor cell lines, we will be able to avoid delays in treatment caused by the time required to culture autologous tumor cell lines, and all patients may receive treatment without exclusion. Through our preliminary work we transferred the technology from a labor-intensive, small-scale approach, to an automated scalable system. Our ultimate goal is to apply this strategy to not only osteosarcoma, but also several other tumor types to validate it as a platform. Like osteosarcoma, there are other therapy-refractory cancers that we believe can benefit from the use of our platform technology, through the use of DC vaccines that offer an immune response against a wider panel of tumor antigens. Citation Format: Vanesa Alonso-Camino, William Mirsch. Validation of a platform to develop new dendritic cell vaccines for cancer immunotherapy [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 6750.

Abstract 1833: Antioxidant and antiproliferative activities of Ipomoea cairica extracts on three breast cancer cell lines

Abstract Introduction: Breast cancer was the top cause of cancer deaths in 2020 with a death toll of 685 000 worldwide. Breast cancer treatments are based on drug specificity for unique receptors on the surface of some breast cancer types, there is a search for alternative treatments that are inclusive of different breast cancer types. Evidence suggests that antioxidants offer a protective effect against certain cancers. Studies on Ipomoea cairica have reported potential anticancer activity, hence this study aimed to assess the antioxidant and antiproliferative potentials of the extracts of Ipomoea cairica on three breast cancer cell lines. Materials and Methods: The study protocol was approved by the University of the Free State Health Sciences Research Ethics Committee (HSREC), with approval number: UFS-HSD2022/0353/2607-0001. Ipomoea cairica stem and leaves were harvested and identified by Prof. H. Baijnath, Department of Conservation Science, School of Life Sciences, University of KwaZulu-Natal, Durban, South Africa, with specimen number 29 30 DD housed at the Ward Herbarium. The dried plant was extracted sequentially using organic solvents ranging from non-polar to polar and water. The extracts were filtered and concentrated using a rotary evaporator and then evaporated to dryness/lyophilized. Cells were cultured according to standard established procedure. The extracts were subjected to phytochemical analyses. FRAP, DPPH, and ABTS assays were done to examine antioxidant activity and MTT assays were used to evaluate the antiproliferative/cytotoxicity of the extracts against normal (Vero) and breast cancer cell lines (MCF-7, MDA-MB-231, and 4T1). Results and Discussion: Extracts of Ipomoea cairica possess alkaloids, saponins, phenols, flavonoids, phytosterols, tannins, triterpenoids, and anthraquinones. The results from the DPPH and ABTS assays revealed that the methanol and ethyl acetate fractions had significantly the highest antioxidant potentials while the methanol stem extract exhibited the most significant antioxidant activity in the FRAP assay. The DCM stem extract showed the highest antiproliferative activity against all tested cell lines. Conclusion: Extracts of I. cairica possess significant antioxidant properties and antiproliferative activities against a wide array of breast cancer types including the most challenging triple-negative breast cancer. The plant would make for a strong herbal candidate for adjuvant therapy with breast cancer medication. Citation Format: Ayodeji Mathias Adegoke, Doreen Kulabako Kyagaba, Nomonde Hadebe, Nandi Mabaso, Relebohile Patricia Lefojane, Himansu Baijnath, Oyeronke Adunni Odunola, Mamello Patience Sekhoacha. Antioxidant and antiproliferative activities of Ipomoea cairica extracts on three breast cancer cell lines [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 1833.

Abstract 5163: Determining the role of septin expression in response of breast cancer patients to microtubule targeted chemotherapy

Abstract Drugs that disrupt the microtubule cytoskeleton, including both microtubule stabilizing taxanes and destabilizers like eribulin, are regularly used in the treatment of metastatic breast cancer. However, there are currently no molecular biomarkers to guide their use. Septins, a class of small GTPases, interact with both microtubules and actin to functionally link these cytoskeletal components and regulate cell biological processes shown to be critical for the invasion and migration of cancer cells. In particular, septin 9 (SEPT9) directly interacts directly with microtubules through an N-terminal domain on the longest isoform, SEPT9i1, which is sufficient to promote migration and invasion of breast cancer cells in vitro. We hypothesized that the relative expression of septin9 isoforms in breast tumors could play a role in their response to treatment with microtubule-targeted chemotherapy, particularly in the metastatic setting. A retrospective investigation of the relationship between septin expression and response to taxane chemotherapy was conducted utilizing full transcriptome sequencing data from biopsies of 75 breast cancer patients treated at our NCI-Designated Mays Cancer Center who received molecular profiling through Caris Life Sciences. Levels of total SEPT9 expression as well as the expression of specific variants that encode mechanistically distinct septin9 isoforms, SEPT9v1, v2, and v3, were determined for each sample and correlated with demographic, treatment, and outcome data for these patients. The oncogenic sept9_v1 variant (which produces the septin9_i1 isoform) was detected in over half of tumors and those with the highest total septin9 expression also expressed significantly higher levels of this isoform. Strikingly, the expression of total septin9, as well as the v3 variant, was significantly increased in taxane-treated patients who survived over a year after diagnosis with metastatic breast cancer, suggesting that septin9 expression could be a molecular correlate of response to taxane chemotherapy. We found that the expression of total septin9 and its variants was not significantly correlated with race, ethnicity, or cancer type suggesting that our findings would apply to our patient population as a whole, which is over 50% Hispanic. These data were used to support an ongoing prospective study to determine effect of septin expression on response to taxane chemotherapy in the neoadjuvant setting and determine the effect of taxane treatment on septin expression by comparing samples pre and post treatment. Together this work supports our overall goal of identifying predictive biomarkers to facilitate the use of microtubule targeted chemotherapy in a more personalized manner. Citation Format: Tamarah Aldawoodi, Jacob Nathaniel Essif, Amna Naqvi, Lauren D. Boyle, Daniela Urueta Portillo, Kate Lathrop, April L. Risinger. Determining the role of septin expression in response of breast cancer patients to microtubule targeted chemotherapy [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 5163.

Abstract 7037: Comprehensive molecular and immune profiling of triple negative invasive lobular carcinoma

Abstract Background: Triple-negative invasive lobular carcinoma (TN-ILC) is a rare (0.1-1.4%) breast cancer with prognosis worse than ER positive ILC. Currently there are no targeted therapies or clinical trials specifically for TN-ILC. A comprehensive analysis of the molecular and immune landscape can help identify novel targets and pathways for TN-ILC to improve patient outcomes. Here, we characterized the molecular and immune signature of TN-ILC. Methods: 11,760 breast cancer samples (Invasive ductal (ID) TNBC, n=364; TN-ILC, n=31) were analyzed (592, NextSeq; WES, NovaSeq), (WTS; NovaSeq) (Caris Life Sciences, Phoenix, AZ). Tumor mutational burden (TMB) totaled somatic mutations per tumor (high ≥10 mt/MB) was tested by NGS. Microsatellite-instability (MSI) was tested by NGS. Immune cell fractions were calculated by deconvolution of WTS. Statistical significance was determined using chi-square and Mann-Whitney U test with p-values adjusted for multiple comparisons (q < 0.05). Results: TN-ILC had higher frequency of CDH1 (0.83% vs. 66.67%), ERBB2 (0.84% vs.28.57%), ARID1A (1.65% vs. 23.33%), CBFB (0.55% vs.16.67%), AKT1 (2.22% vs.16.13%) and KMT2C (3.83% vs.13.79%), but lower frequency of TP53 (89.7%5 vs. 38.71%) mutations (all p<0.05) compared to ID-TNBC. TN-ILC had higher frequency of TMB high (23.3% vs. 5.2%, p<0.05) but there was no difference in dMMR/MSI-H (0% vs 1.1%, p=1). TN-ILC had higher AR RNA (FC: 14.2), protein expression (80.6% vs. 24.8%) and higher frequency of fusion variant-AR (16.1% vs 4.9%) (all p < 0.05) compared to ID-TNBC. Analysis of inferred immune cell infiltrates showed that TN-ILC had higher infiltration of M2 macrophages (5.34% vs. 2.94%) and neutrophils (4.83% vs. 2.65%) but lower infiltration of M1 macrophages (3.46% vs 2.17%) and CD8 T cells (0.58% vs. 0.11%) (all p<0.05). TN-ILC had lower T cell inflamed signature (16.1% vs 35.7%, p<0.05), decreased immune checkpoint genes (CD274, CTLA4, FOXP3, LAG3, IDO, FC: 1.2-2.7, all p < 0.05), PD-L1 protein expression (SP142: 30.4% vs. 50.2%, p=0.06; 22c3: 15.4% vs. 42.6%, p = 0.05) and differential expression of BCL2 family genes (upregulation: BIK, FC: 2.2; downregulation: BAX, BAK1, BID, PMAIP1, MCL1, BCL2A1, BCL2L10, FC: 1.0-6.3, all p < 0.05) compared to ID-TNBC. Conclusion: These data suggest that TN-ILC had higher frequency of CDH1, ERBB2, AKT1, ARID1A mutations, higher M2 macrophages and neutrophils and lower M1 macrophages and CD8 T cells infiltration and, lower T cell inflamed signature. High TMB and AR expression can translate into use of immunotherapy (ICI) and AR antagonists in these patients. Additonal analysis to determine the optimal biomarker for ICI response in TN-ILC is needed. Results of our study need to be validated in larger studies with survival correlation. Citation Format: Pooja Advani, Sachin Deshmukh, Sharon Wu, Jacob Andring, Joanne Xiu, Jose Leone, Priya Jayachandran, Stephanie Graff, Mathew Oberley, George Sledge, Asher Chanan-Khan. Comprehensive molecular and immune profiling of triple negative invasive lobular 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 7037.

Abstract 5627: PPARG-high circulating monocytes exhibit an immunosuppressive phenotype in urothelial cancer patients treated with anti-PD1

Abstract Background: PPARG is expressed in a variety of immune cells such as monocytes, macrophages, and lymphocytes, where it plays a role in their maturation and function. PPARG has been implicated in immune mediated resistance but to date, a comprehensive analysis of PPARG expression in peripheral blood mononuclear cells (PBMCs) in urothelial cancer (UC) patients is lacking. In this study, we identified the immunophenotypes associated with PPARG expression in UC patients treated with anti-PD1 therapy. Methods: We performed single-cell RNA sequencing of PBMCs from 5 UC patients and 3 normal healthy volunteers (NHV). Real-world samples (Discovery Life Sciences) were collected from patients who received prior Pembrolizumab. Time post-treatment to collection ranged from 46 to 307 days. An average of 9,500 cells were recovered per donor reflecting an ~80% QC pass rate. Count matrices were generated using 10x Genomics Cell Ranger pipeline (v7.0.0; GRCh38 reference). Downstream data processing, analysis and visualization were done on Cellenics software. Results: We evaluated 75,725 cells and identified 9 distinct cell populations. The proportion of T-cells was significantly decreased in UC samples compared to NHVs (40% vs 60%, p=0.03). Highest PPARG expression was observed on Classical Monocytes (CMs) cell population. PPARG expression was higher on CMs in UC compared to NHV samples (1.74 vs 0.11 logcounts; padj<0.001). Unsupervised clustering of all monocytes revealed two distinct CM clusters. In cluster A, 95% of the cells originated from UC samples, while most cells in cluster B (81%) originated from NHV samples. PPARG was the top differentially expressed gene upregulated in cluster A (PPARG-high) vs cluster B (PPARG-low) (1.89 vs 0.23 logcounts; padj<0.001). PPARG-high CMs are characterized by a gene signature inclusive of GPRIN3, RGCC, MAFB and IRF2BP, which have been shown to be upregulated in M2 polarized macrophages [1]. Moreover, pathway analysis using GSEA highlighted that PPARG-high CMs are enriched for TREM1 signaling genes (padj<0.001), whose high expression has been associated with immunosuppressive phenotypes in myeloid cells [2] and shorter survival of patients across other cancers [3]. Conclusion: PPARG-high circulating CMs in UC patients exhibit a transcriptomic profile associated with immunosuppression and M2 macrophage polarization. FX-909, a first-in-class PPARG inverse agonist, is being evaluated in the clinic and the study includes an exploratory biomarker approach to assess the potential effect of PPARG inhibition on the CM immune-profile of advanced UC patients enrolled in the Ph1 study [4].

Abstract 6091: Multi-cancer early cancer detection and cancer-type identification with high sensitivity non-invasive using urine: Pancreatic lung cancer early detection

Abstract N-NOSE, a breakthrough cancer detection test from HIROTSU BIO SCIENCE, utilizes the unique chemotactic behavior of the nematode C. elegans. This nematode is drawn to the urine of individuals with 15 different types of cancers (including stomach, colon-rectum, lung, breast, pancreas, liver, prostate, uterus, esophagus, gallbladder, bile duct, kidney, urinary bladder, ovary, oropharynx) while avoiding urine from healthy persons, making N-NOSE a comprehensive primary screening tool. It has a high accuracy rate, with 87.5% sensitivity and 90.2% specificity for early-stage cancers. Being non-invasive and reasonably priced, N-NOSE has been a significant success in Japan since its 2020 launch, with over 500,000 screenings.This presentation delves into the technology aspect of N-NOSE in Japan, comparing its clinical research and key findings with competitors to underscore its effectiveness in early cancer detection. Furthermore, N-NOSE now advances to not only detect cancer but also identify its type in a noninvasive way, using urine and with high PPV.With pancreatic cancer being particularly lethal, we outline “N-NOSE plus pancreas,” the world’s first pancreatic cancer detection test at an early stage. This involves genetically modifying C. elegans to react specifically to early-stage pancreatic cancer urine samples, a development centered on the newly identified cr-4 GPCR in AWC neurons. Next, we profile our newest liver-cancer-specific test, “N-NOSE plus liver,” which offers early liver cancer detection with high sensitivity and specificity using urine.Finally, we highlight how N-NOSE bridges the gap in cancer screening and type identification, offering a complete, non-invasive solution. Citation Format: Eric Di Luccio, Takaaki Hirotsu. Multi-cancer early cancer detection and cancer-type identification with high sensitivity non-invasive using urine: Pancreatic lung cancer early detection [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 6091.

Procedural norms and their significance in the implementation of the competence of representative bodies

On the eve of the centenary of the birth of Professor E. I. Kozlova, who devoted her scientific life to the study of the problems of democracy and the forms of its implementation, attention should be paid to the problem of improving the mechanisms of parliamentary activity, which she repeatedly raised in her writings of the soviet period. Revealing the legal nature and essence of the representative bodies of the soviet period — the soviets of workers’ deputies, later — the Soviets of People’s deputies, in the post-Soviet period — parliaments, E.I. Kozlova emphasized the enduring importance of the procedures of the representative bodies. Analyzing the scientific heritage of Professor E. I. Kozlova, the author substantiates the relevance of the scientist’s conclusions and proposals for modern representative bodies, defines some directions for the development of the scientist’s ideas.

Bioethics-CSR Divide

Bioethics-CSR Divide