bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5367: Single cell resolution spatial transcriptomics reveals genetic driver and TME interaction in clear cell renal cell carcinoma
Authors: Strunilin Ilya , Caravan Wagma , Abedin-Do Atieh , Houston Andrew , Li Yize , Song Yizhe , Cao Song , Mo Chia-Kuei , Chen Siqi , Pachynski Russel , Chen Feng , Ding Li
Published: March 2024

BAP1 mutations are associated with increased T cell and tumor-associated macrophage infiltration in clear cell renal cell carcinoma (ccRCC) and are predictive of poor prognosis. BAP1 mutations have also been found to be associated with T-cell infiltration in peritoneal mesothelioma and uveal melanoma. Because BAP1 mutations are known to be associated with aggressive disease and poor survival, we hypothesize that these mutations lead to a more immunosuppressive microenvironment. Using single nucleus RNA sequencing and high-resolution spatial transcriptomics, we confirmed that BAP1 mutant tumors are immune hot with a high fraction of Treg, exhausted CD8+ T cells, CD4+ T cells, and macrophages. At the same time, these tumors have a lower proportion of endothelial cells and pericytes. Evident of pro-tumor microenvironment we have found BAP1 mutations to be associated with the upregulation of SPP1, CD163, and MARCO immunosuppressive genes in tumor-associated macrophages. Here we propose a mechanism by which BAP1-mutant cancer cells induce an immunosuppressive microenvironment: through activation of the complement system and TGF-β signaling to attract macrophages. Taken together, our results suggest that BAP1 mutations contribute to poor ccRCC outcomes not only through cell-intrinsic effects, but also by promoting infiltration of Tregs and tumor-associated macrophages and polarizing infiltrated macrophages towards immunosuppressive phenotype.

Citation Format: Ilya Strunilin, Wagma Caravan, Atieh Abedin-Do, Andrew Houston, Yize Li, Yizhe Song, Song Cao, Chia-Kuei Mo, Siqi Chen, Russel Pachynski, Feng Chen, Li Ding. Single cell resolution spatial transcriptomics reveals genetic driver and TME interaction in clear cell renal cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5367.
Abstract 5372: KITENIN leads immunosuppression in the glioblastoma microenvironment via CCL20
Authors: Ahn Eun-Jeong , Kim Nah Ihm , Kim Sung Sun , Lee Jae-Hyuk , Rhee Joon Haeng , Kim Kyung-Keun , Kim Hangun , Lee Kyung-Hwa , Moon Kyung-Sub
Published: March 2024

Background: Glioblastoma (GBM) is characterized by an immunosuppressive tumor microenvironment (TME), limiting the effectiveness of existing treatments. The author investigated the role of KAI1 COOH-terminal interacting tetraspanin (KITENIN) in mediating this immunosuppression, focusing on its effect on cytokine secretion and tumor-infiltrating lymphocyte (TIL) profiles in the TME.

Materials and Methods: Using cytokine array and Luminex multiplex assays, the author evaluated cytokine secretion in KITENIN-modulated GL261 cells. The author further isolated TILs from an orthotopic mouse model implanted with these cells and induced in vitro differentiation of myeloid cells. The effect of neutralizing antibody for target cytokine, was assessed in vivo, and clinical relevance was investigated using GBM patient data from our cohort and TCGA.

Results: Employing cytokine array and Luminex multiplex assays, we found increased level of CC chemokine ligand 20 (CCL20) within KITENIN-overexpressed (KIT-HA) GL261 cell supernatants. Immunohistochemical analyses using GBM samples confirmed that both KITENIN and CCL20 expressions co-directionally increased and this was associated with decreased survival by TCGA analysis. Myeloid-derived suppressor cells (MDSCs), macrophages, and CD4+Foxp3+ regulatory T cells (Tregs) were observed to be higher in brain tumor implanted with KIT-HA GL261. Furthermore, KIT-HA GL261 conditioned medium expanded CD45+CD11b+Gr1-Ly6c+ monocytic MDSCs (M-MDSCs), an effect that was abrogated by CCL20 downregulation. In vivo neutralization of CCL20 resulted in reduced tumor volume, prolonged survival, and decreased M-MDSCs, thus affirming the role of CCL20 in mediating immunosuppression.

Conclusion: These findings underscore the KITENIN-CCL20 axis as a promising target for alleviating the immunosuppressive TME in GBM, potentially unlocking new avenues for GBM immunotherapy.

Citation Format: Eun-Jeong Ahn, Nah Ihm Kim, Sung Sun Kim, Jae-Hyuk Lee, Joon Haeng Rhee, Kyung-Keun Kim, Hangun Kim, Kyung-Hwa Lee, Kyung-Sub Moon. KITENIN leads immunosuppression in the glioblastoma microenvironment via CCL20 [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 5372.
Abstract 539: LN18-MRTX1719R cell line, a useful cell model for the discovery of the new PRMT5 inhibitors
Authors: Tang Yao , Wang Liping , Wang Guoqian , Ning Jinying , Hao Feng
Published: March 2024

In recent years, the PRMT5-MTA complex has emerged as a "synthetic lethal" novel target for the treatment of MTAP (methylthioadenosine phosphorylase)-deficient cancers. Kryukov et al. first reported the "synthetic lethal" effect of inhibiting PRMT5 in MTAP-deficient tumors, and they identified MTAP as the synthetic lethal partner of PRMT5 in 2016. MTAP is a tumor suppressor gene frequently lost in various cancers, including pancreatic cancer, non-small cell lung cancer, and mesothelioma. Furthermore, the loss of MTAP leads to the accumulation of MTA in cancer cells, which competes with PRMT5's co-factor SAM for binding and forms the PRMT5-MTA complex. Cancer cells lacking MTAP show enhanced sensitivity to PRMT5 loss. Therefore, this complex has emerged as a potential therapeutic target, particularly for the treatment of MTAP-deficient cancers. MRTX1719 is a highly effective selective inhibitor of the PRMT5-MTA complex, capable of selectively inhibiting PRMT5 activity in MTAP-deficient cells. As the most promising candidate drug, MRTX1719 is undergoing Phase I/II clinical evaluation for patients with MTAP-deficient cancers. Drug resistance is an inevitable problem with small molecule inhibitors. Currently, the mechanism of resistance to MRTX1719 is not clear, as MTAP/CDKN2A loss is an early driver event in most cancers. To further explore the resistance mechanism of MRTX1719, our group used the MTAP-deficient LN-18 cell line and induced mutagenesis using the mutagen ENU. We then used a drug low-concentration gradient method to establish the LN-18-MRTX1719R-resistant cell line. Specifically, LN-18 cells were treated with increasing doses of MRTX1719 for 6 months, and mixed clones that tolerated MRTX1719 were selected. Subsequently, single clones were selected, and a stable MRTX1719-resistant clone was obtained, named LN-18-MRTX1719R. Cell proliferation inhibition assay showed that the IC50 of LN-18-MRTX1719R was over 1000nM, which is 100-fold more resistant than the parental LN-18 cells (9.87 nM). We explored the resistance mechanisms using RNA-Seq and WES methods. The resistance to MRTX1719 of the LN-18-MRTX1719R cell line was highly stable after 20 cell passages. The LN-18-MRTX1719R cell line can be a useful cell line mode in the development of future PRMT5·MTA inhibitors.

Citation Format: Yao Tang, Liping Wang, Guoqian Wang, Jinying Ning, Feng Hao. LN18-MRTX1719R cell line, a useful cell model for the discovery of the new PRMT5 inhibitors [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 539.
Abstract 54: Engineering chimeric antigen receptor (CAR) T cells using autoantigen binding domains for treatment of B cell malignancies with increased specificity
Authors: Cheever Abiagil , Lindsay Hunter , Hansen MacKenzie , Kang Chloe , Weber K. Scott , O'Neill Kim
Published: March 2024

Introduction: Chronic inflammation often leads to cancer, and many autoimmune diseases lead to a significantly increased risk of developing cancer. Though many autoimmune diseases, like Graves’ Disease (GD), don’t have treatments that address this risk. Chimeric antigen receptor (CAR) T cells are an effective treatment for hematological cancers by eliminating all B cells, but a more targeted form of CAR T cells could eliminate specific B cell populations in B cell malignancies, including GD.

Methods: To allow our CAR T cells to target specific B cell populations that cause GD, we engineered chimeric autoantigen receptors (CAAR) with epitopes of thyroid stimulating hormone receptor (TSHR) as the binding domain. This acts as bait for the anti-TSHR B cells. Jurkat T cells were lentivirally transduced with these CAAR plasmids. Anti-TSHR B cell lines were engineered by transducing Nalm6 cells with lentivirus for anti-TSHR B cell receptors. Cytotoxicity assays were performed on primary human TSHR CAAR T cells incubated with the anti-TSHR B cell lines. Cytokine production, proliferation, and cytotoxicity assays were performed on these coincubated cells to determine the efficacy of our TSHR CAAR T cells.

Results: Two of the three Jurkat TSHR CAAR T cells showed significant binding to anti-TSHR antibodies (TRAbs). They also showed significant activation, measured by CD69 expression, when incubated with plate bound TRAbs, but not soluble TRAbs. The B cell lines were validated by binding to TSHR. The primary TSHR CAAR T cells showed significantly increased proliferation and production of TNF, IFN-y, and IL-2, when compared with controls. The TSHR CAAR T cells also had significant cytotoxic action against the anti-TSHR B cell lines, but not Nalm6 B cells. The cytotoxic function was comparable to our CD19 CAR T cell with the same activation domains, though it demonstrated specificity to only anti-TSHR B cells.

Conclusion: TSHR CAAR T cells are a potentially curative treatment for GD that will stop chronic inflammation and could reduce GD patient’s risk of developing thyroid cancer. This is also an important development in CAR T cell therapies that allows for specific targeting of certain B cell populations.

Citation Format: Abiagil Cheever, Hunter Lindsay, MacKenzie Hansen, Chloe Kang, K. Scott Weber, Kim O'Neill. Engineering chimeric antigen receptor (CAR) T cells using autoantigen binding domains for treatment of B cell malignancies with increased specificity [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 54.
Abstract 5420: NLRX1 involves in prostate cancer cell growth, survival and migration
Authors: Lin Wan-Wan , Rathore Varsha
Published: March 2024

Prostate cancer (PCa) is one of the most common male cancer and is the second leading cause of death in men. The basic working components about its cause and progression are not surely known in the research as well as the practitioner community. The current curing therapies such as surgery, radiation therapy, cryotherapy, and hormone therapy are not very encouraging. Subsequently, there is a need to investigate new potential targets for focused treatment. NLRX1 (NOD9 or NLR family member X1) is one member among the NLRs and is a non-inflammasome forming NLR sub-group localized in mitochondria. Previous studies showed that NLRX1 is involved as a tumor suppressor in colorectal cancer and hepatocellular carcinoma. However, NLRX1 serves as tumor promoter in breast cancer and head and neck cancer sarcoma. The present study focuses on the role of NLRX1 in prostate cancer cells under stress conditions including serum-free and hydrogen peroxide (H2O2) to understand cell regulation and functions of NLRX1. We used human prostate cancer PC3 and LNCaP cell lines as the models and the following methods, including Annexin V/PI staining for cell viability, BrdU incorporation for cell proliferation, mitoSOX staining for mitochondrial ROS, wound healing assay for cell migration and Boyden chamber for cell invasion. First, we assessed the TCGA database via UCSC Xena and found that prostate cancer tumors have high NLRX1 expression than normal tissues. Next, we knock down the NLRX1 through lentivirus transduction. We found that silencing NLRX1 expression can decrease cell proliferation in PC3 cells. Furthermore, silencing NLRX1 can significantly decrease cell viability under H2O2 in PC3 cells and serum-free condition in both cancer cell lines. In addition, we found that H2O2-induced cell death is reversed by necroptosis inhibitor Nec-1. Moreover, NLRX1 silencing reduces cell migration in both PC3 and LNCaP cells and can inhibit cell invasion in both cell lines at 24 h. These results suggest that NLRX1 is a positive regulator of cell growth, migration and invasion in prostate cancer cells, and NLRX1 provides a cell-protective role in response to H2O2 and serum-free stress. Thus, this study will provide a potential target for the development of a therapeutic strategy for prostate cancer.

Citation Format: Wan-Wan Lin, Varsha Rathore. NLRX1 involves in prostate cancer cell growth, survival and migration [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 5420.
Abstract 5436: A therapeutically targetable dysadherin/YAP axis promotes cancer stem features and tumor aggressiveness in hepatocellular carcinoma
Authors: Jeon So-El , Jang Tae-Young , Lee Choong-Jae , Yun Hyeon-Ji , Cho Yeong-Hoon , Lim Da-Ye , Nam Jeong-Seok
Published: March 2024

Hepatocellular carcinoma (HCC) remains one of the most challenging tumors to treat. Cancer stem-like cells (CSCs) are believed to be important in HCC development, immune evasion, and treatment resistance. Dysadherin is a cell membrane glycoprotein linked to cancer progression by endowing HCC with CSC features. However, the underlying mechanism of increasing CSC characteristics in refractory advanced HCC is not entirely understood. A comprehensive bioinformatics study revealed that dysadherin-expressing HCC cells were associated with cancer stemness and the YAP signaling activation. Dysadherin knockdown or overexpression in liver cancer cell lines attenuated or increased cancer stemness and aggressiveness via YAP signaling, respectively. Correspondingly, dysadherin deficiency lowered cancer stemness and aggressiveness in the HCC mouse model by blocking the YAP pathway. Mechanistically, dysadherin promoted YAP nuclear localization, which activated stem cell transcription factors such as OCT4, KLF4, and MYC. Of note, our comprehensive gain and loss of function studies in CSCs demonstrated that targeting the dysadherin/YAP axis could boost sorafenib’s therapeutic effect and suppress PD-L1 expression, which helps tumor cells escape from immune surveillance. Thus, our results highlight that the dysadherin/YAP axis drives the aggressive phenotype of HCC, providing a potential therapeutic strategy for HCC.

Citation Format: So-El Jeon, Tae-Young Jang, Choong-Jae Lee, Hyeon-Ji Yun, Yeong-Hoon Cho, Da-Ye Lim, Jeong-Seok Nam. A therapeutically targetable dysadherin/YAP axis promotes cancer stem features and tumor aggressiveness in hepatocellular 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 5436.
Abstract 544: Reverse engineering strategy for new targets identification
Authors: Xie Bin , Zhang Taimei , Li Le , Chen Hongkui , Liu Yang , Bi Lintao , Wen Danyi
Published: March 2024

The issue of drug resistance in cancer is very similar to the field of infectious disease, either existing before treatment (intrinsic) or generated during therapy (acquired), highly affected by cell endogenous factors and/or the surrounding environment. Acquired resistance can be occurred several months after targeted therapy via alternations of drug targets due to the genomic mutations of the targets, epigenetic transcriptional or translational changes, for example the C797S mutation impairs AZ9291 binding to the EGFR kinase domain. To overcome this, novel well-designed chemical compounds targeting new mutations would be urgently needed, as well as the potential new oncogenes/susceptible genes driving the resistance. Considering about the heterogeneity of primary tumor tissues, LIDE has established one functional cost-effective in vitro and in vivo screening method, named reverse engineering strategy for targets identification by using patients-derived conditional reprogramed cancer cell lines (CRs) and IO-FIVE (Immuno-Oncology drugs Five In Vivo Efficacy test). Basically, pro-siRNA library or CRISPR/sgRNA library is carried out in CRs for candidates hunting, alternatively drug-response evaluation among multiple clinical biopsies in mice cancer trials, after filtered with bioinformatic scanning from large groups of patients, and finally it is validated in vivo by CRISPR, shRNA silence or targeted compounds. In addition to these known factors (EGFR C797S, Her2/Met amplification, Ras and MAPK activation), we have discovered that epigenetic factors for DNA modification, protein translation kinases, mitochondrial/peroxisomal transporters and cell membrane lectins may contribute to AZ92921 resistance in vivo. And one previously unknown acyltransferase member has been identified as key driver gene for pan-cancer development by IO-FIVE, and patients with high expression level of this enzyme have had poor survival outcomes, accompany with uncontrolled tumor growth and immunotherapy resistance, which is becoming a promising therapeutic target.

Citation Format: Bin Xie, Taimei Zhang, Le Li, Hongkui Chen, Yang Liu, Lintao Bi, Danyi Wen. Reverse engineering strategy for new targets identification [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 544.
Abstract 5441: Down regulation of cysteine glutamate antiporter in ALDH1A1 expressing oral and breast cancer stem cells induced oxidative stress triggered ferroptosis
Authors: Bellala Ravishankar
Published: March 2024

Background: Sulfasalazine, an xCT inhibitor, is being used as a repurposed antineoplastic drug to induce ferroptosis. Ferroptosis is a regulated necrotic cell death pathway that is dependent on iron reserves. Interestingly, cancer stem cells (CSCs) that are regarded as major drivers of resistance to conventional therapies accompanied with tumor relapse and recurrence have bulk amount of iron reserves in the form of ferritin. This suggests that inducing ferroptosis might disrupt stemness and drug resistant mechanisms in cancer stem cells, thereby reducing the risk of drug resistance, cancer recurrence, and relapse.

Materials & Methods: In the present study, ALDH1A1 expressing oral (OCSCs) and breast (BCSCs) cancer stem cells were sorted and used to investigate the role of sulfasalazine to induce ferroptosis. To check the self-renewability of CSCs spheroid formation assay was performed and the resultant CSCs were treated with sulfasalazine (SAS) and subjected to gene expression analysis RT PCR and flow cytometry. FACS was performed to check stem cell marker expression, cell cycle arrest, and apoptosis.

Results: Our results suggest that the cells showed a gradual increase in sphere formation till S3 in the case of OCSCs and S2 in the case of BCSCs, with a gradual decrease in sphere forming efficiency from the respective generations. When treated with 0.6mM SAS, these cells induced ferroptosis by downregulating stem cell markers like ALDH1A1, SLC7A11, ferritin, and GPx 4 with a concomitant increase in transferrin and STEAP 3. Flow cytometry studies revealed that the cells have undergone mitochondrial dysfunction characterized by loss of membrane potential and the cell cycle progression was halted in the G2/M phase.

Conclusion: In the present study, we demonstrate that SAS potentially induced ferroptosis accompanied with oxidative stress in both OCSCs as well as BCSCs by lowering GPx 4 activity, a key enzyme that scavenges the products produced as a result of oxidative stress.

Citation Format: Ravishankar Bellala. Down regulation of cysteine glutamate antiporter in ALDH1A1 expressing oral and breast cancer stem cells induced oxidative stress triggered ferroptosis [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 5441.
Abstract 5449: Exploring signaling pathways essential for stemness and metastasis of colorectal cancer cells
Authors: Fujishita Teruaki , Taketo Makoto M. , Aoki Masahiro
Published: March 2024

The prognosis of colorectal cancer (CRC) patients with distant metastasis remains poor, requiring elucidation of molecular mechanisms and identification of therapeutic targets. While cancer stem cells have been implicated in metastasis, recurrence, and drug resistance, our understanding of the regulatory mechanisms governing the stemness of CRC stem cells remains largely incomplete. In this study, we utilized an innovative autochthonous metastatic mouse model of CRCs generated by the introduction of sporadic mutations in Ctnnb1, Kras, Trp53, and Smad4 genes (CKPS mice) and CRC cell lines derived from the mice (CKPS cells). Our investigation revealed that the cAMP/PKA/CREB and TGFβ/SMAD4 pathways play a positive and negative role, respectively, in the sphere-forming and metastasis-initiating ability of CRC cells. Treatment with the CREB inhibitor 666-15 or the genetic knockout of CREB significantly reduced the spheroid-forming and metastasis-initiating potentials of CKPS cells. Intriguingly, we observed that the irinotecan treatment induced CREB phosphorylation in liver metastases, and co-treatment of 666-15 greatly potentiated the inhibitory effect of irinotecan on liver metastasis. Furthermore, we found that the transition from 2D to 3D spheroid culture of CKPS cells led to the induction of RhoC expression. Notably, RhoC knockout in CKPS cells effectively suppressed spheroid formation in vitro and inhibited metastasis formation in an intrasplenic injection liver metastasis assay. Potential therapeutic implications of these findings will be discussed.

Citation Format: Teruaki Fujishita, Makoto M. Taketo, Masahiro Aoki. Exploring signaling pathways essential for stemness and metastasis of colorectal cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5449.
Abstract 5452: AXIN2 differentially modulates stem cell subpopulations providing a potential therapeutic opportunity for targeting colorectal cancer stem cells
Authors: Zhang Chi , Facey Caroline O. , Opdenaker Lynn M. , Boman Bruce M.
Published: March 2024

Constitutively activated WNT-signaling promotes stem cell (SC) overpopulation which drives most colorectal cancer (CRC) development. It is not known, however, how WNT-signaling activity regulates different SC sub-populations. We previously reported that SC population size decreases as AXIN2 is knocked down, and increases when AXIN2 is overexpressed. Changes in cell cycle and apoptotic rate were also observed. Here, we present our recent results on changes between different SC subpopulations (ALDH+ vs. LGR5+) that occur due to AXIN2 modulation. We hypothesize that modulation of AXIN2 promotes differential expansion between different CRC SC subpopulations (ALDH+ and LGR5+). We observed that AXIN2 not only increases LGR5+ SC population size, but also expands the ALDH+ SC population to a greater extent as shown by an increased ALDH+-to-LGR5+ SCs ratio. This indicates that AXIN2, via WNT signaling, regulates SC population size, particularly ALDH+ SCs. Since the ALDH SC marker is also a key component in the retinoic acid (RA) signaling pathway, we surmised that increased AXIN2 would not only lead to elevated ALDH levels, but also upregulate the RA-signaling pathway components in CRC cells. To this end, we conducted NanoString mRNA profiling to study the effects of AXIN2 knockdown and overexpression in HT29 and SW480 CRC cells. The results showed that increased AXIN2 significantly up-regulated several key components of the RA-signaling pathway. Taken together, our results indicate that AXIN2 drives a shift in SC subpopulation sizes favoring the ALDH+ SCs. It also promotes the RA-signaling component expression - a pathway known to regulate SC differentiation. As a potential treatment strategy for CRC, therapeutically inducing high levels of AXIN2 could shift the balance between SC populations towards ALDH+ SCs, which may provide a way to target cancer SCs with retinoids to induce their differentiation.

Citation Format: Chi Zhang, Caroline O. Facey, Lynn M. Opdenaker, Bruce M. Boman. AXIN2 differentially modulates stem cell subpopulations providing a potential therapeutic opportunity for targeting colorectal cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5452.
Abstract 5471: Ribosome heterogeneity and RNA polI inhibition in osteosarcoma heterogeneity
Authors: Baud'huin Marc , Colleville Elvina , Thepault Rose Anne , Orgebin Emilien , Lamoureux Francois , Ory Benjamin
Published: March 2024

Osteosarcoma is the most common primary bone tumour in adolescents and young adults, with an incidence rate of 3.8 per million. These tumors affect the long bones of the body such as femur. Current standard therapy includes tumour resection associated with multi-drug chemotherapy. The 5-year survival rate can reach 65% but is significantly reduced to 30% when pulmonary metastases are detected at the time of diagnosis. As this rate has not improved in the last 50 years, it is necessary to identify new therapeutic strategies. Inhibition of ribosomal biogenesis is one of the strategies developed in many cancer studies. This can be obtained by inhibition of RNA pol I, and in particular by BMH-21. The objective of this study was to analyze the complexity and heterogeneity of osteosarcomas through the ribosomal spectrum. The effect of BMH-1 on survival was also studied in osteosarcoma cell lines. The data obtained showed that there was no difference in ribosome protein composition in the “core” ribosome in different osteosarcoma cell lines. In addition, BMH-21 decreased cell viability in all osteosarcoma and chemoresistant cell lines. However, any activation of the apoptotic pathway by caspases was detected, nor any impact on the formation of new colonies after treatment with BMH-21. These results suggest a rather cytostatic mechanism. Moreover, a G0/G1 arrest in cell cycle was shown for U2OS cells. Finally, BMH-21 does not appear to have any impact on migration in HOS and U2OS cells. The different molecular mechanisms involved in the regulation of proliferation need to be further investigated.

Citation Format: Marc Baud'huin, Elvina Colleville, Rose Anne Thepault, Emilien Orgebin, Francois Lamoureux, Benjamin Ory. Ribosome heterogeneity and RNA polI inhibition in osteosarcoma heterogeneity [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 5471.
Abstract 5479: Spatially resolved molecular features in the tumor-immune microenvironment of germline BRCA1/2 mutated versus BRCA wild type high grade serous ovarian cancers
Authors: Qian Jing , Roman Lynda D. , Bassiouni Rania , Rajpara Seeta , Neuman Monica , Khetan Varun , Craig David W. , Carlson Joseph W. , Carpten John D.
Published: March 2024

High-grade serous ovarian cancer (HGSC) is the most common and aggressive subtype of epithelial ovarian cancer. Importantly, germline mutations in BRCA1/2 give rise to HGSC. Because most HGSC are diagnosed at stage III/IV, outcomes are poor and more effective therapies are needed. Although immune checkpoint inhibitors show promise for a number of cancer types, efficacy has been variable in HGSC. To help provide new clues to the molecular features of HGSC, we set out to elucidate the tumor microenvironment of HGSC using spatial transcriptomics approaches. Specifically, we applied 10x Genomics Visium Spatial Transcriptomics (ST) technology to 48 samples representing primary HGSC tumors from 24 patients, including 15 BRCA1/2 mutated cases and 9 BRCA non-mutated cases. In addition to individual sample analysis, we normalized data from 64,784 total features across all sections and performed an integrated cluster analysis to get an overall picture of the cellular architecture of HGSC, but to also directly investigate differences in the tumor microenvironment based on BRCA mutation status. The resulting 17 integrated clusters were subjected to molecular annotation using GSEA, ESTIMATE, and CIBERSORTx, which revealed feature clusters representing important cell types defined by marker genes. Analysis of the entire dataset identified a primary tumor cluster defined by the expression of known ovarian cancer genes including WFDC2 and “don’t eat me” marker genes CD24 and CD47. However, the predominant tumor cell population in BRCA mutated cases was marked by the expression of MUC1, EPCAM, CLDN3, and CD24, while BRCA non-mutated tumor cell cluster was marked by EFNA1, ID1, LSR, and CD9. Importantly, both groups revealed the presence of a specialized monocyte derived macrophage cluster defined by the expression of LYZ and genes involved in cholesterol metabolism including APOE and APOC1. These lipid-associated macrophages are M2 polarized and shown to be associated with poor outcome. Additional analyses are underway to determine spatial dependencies of various tumor and immunological cell compartments. These data provide novel insights into the HGSC tumor microenvironment that may influence HGSC’s vulnerability to immunological therapies.

Citation Format: Jing Qian, Lynda D. Roman, Rania Bassiouni, Seeta Rajpara, Monica Neuman, Varun Khetan, David W. Craig, Joseph W. Carlson, John D. Carpten. Spatially resolved molecular features in the tumor-immune microenvironment of germline BRCA1/2 mutated versus BRCA wild type high grade serous ovarian cancers [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 5479.
Abstract 5504: Integrating ultrahigh-plex spatial phenotyping: From discovery to clinical applications
Authors: Pratapa Aditya , Ma Ning , Jhaveri Niyati
Published: March 2024

Background The journey from discovering predictive spatial biomarkers (known as spatial signatures) for immunotherapy responses to their clinical application necessitates a cohesive approach bridging ultrahigh-plex discovery experiments with high-throughput translational studies. This study focuses on integrating Akoya Biosciences spatial multiplexed imaging technologies with advanced data analysis techniques for comprehensive spatial phenotyping across the discovery-to-clinical continuum.

Methods Human formalin-fixed, paraffin-embedded (FFPE) cancer tissues were profiled using ultrahigh-plex PhenoCode™ Discovery panels to examine cell lineage, immune activation, and checkpoint markers via the PhenoCycler®-Fusion spatial biology platform. The subsequent use of PhenoCode™ Signature panels targeted specific biomarkers of immune profile, immune contexture, tumor-infiltrating lymphocytes (TIL), macrophage polarization, and T cell status via the PhenoImager® HT platform. Open-source whole slide image analysis software QuPath was employed for precise image analysis, including ROI segmentation, cell detection, classification, exploration of spatial interactions, and identification of distinct spatial signatures.

Results Our analysis revealed distinct spatial relationships within various tumor types, quantifying immune cell distributions and their interactions. The ultrahigh-plex data correlated with high-throughput signature panel analyses, thus paving a new simplified way for targeted identification and development of predictive spatial signatures for immunotherapy outcomes.

Conclusions The combined use of ultrahigh-plex discovery panels, high-throughput signature panels, and deep-learning image analysis provides a comprehensive understanding of cellular interactions in the tumor microenvironment. This integrated approach, utilizing Akoya's end-to-end workflows, accelerates the identification of predictive spatial biomarker signatures across various human tissue samples.

Citation Format: Aditya Pratapa, Ning Ma, Niyati Jhaveri. Integrating ultrahigh-plex spatial phenotyping: From discovery to clinical applications [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 5504.
Abstract 5515: Low body mass index (BMI) induces neuronal NPY and promotes brain metastasis of lung cancer
Authors: Tyagi Abhishek , Wu Shih-Ying , Deshpande Ravindra , Wu Kerui , Liu Liang , Watabe Kounosuke
Published: March 2024

Brain metastases (BrMs) are a common occurrence in lung cancer with a dismal outcome. To develop a more effective therapeutic strategy, it is crucial to understand the underlying pathological mechanism of brain metastasis. Recent clinical data has unveiled a significant association between low body mass index (BMI) and an increased risk of brain metastasis in lung cancer patients, leading to reduced survival rates compared to other primary tumors. However, the molecular mechanism underlying this paradoxical relationship, often referred to as the "obesity paradox," in the context of lung cancer mortality presents a complex and intriguing puzzle and warrants in-depth investigation. Here, we examined biological mechanism of how low BMI promotes lung cancer brain metastasis. In this study, we conducted a retrospective pan-analysis of 5,516 patient cohorts with brain metastasis originating from lung, breast, melanoma, and renal cancers with their BMI status. We found that lung cancer patients with low BMI have significantly higher brain metastatic incidence compared to the high BMI patients, in contrast to breast, melanoma, and renal cancers. Furthermore, we found that low BMI plays a pivotal role in mediating neuron activation within the brain and that the secretion of neuronal neuropeptide Y (NPY), a protein involved in appetite and energy homeostasis via GHSR-receptor manner, promotes metabolic switching through NPY/NPY5R axis in the tumor cells in the brain thereby enabling metastasis. Elevated levels of neuronal NPY in the brains of cancer-free subjects with low BMI suggest its potential utility as a promising prognostic biomarker for identifying the increased risk of metastatic disease in lung cancer patients with low BMI. We also demonstrated that reversing low BMI or blocking the NPY/Y5R interactions effectively abrogated brain metastasis in our animal model. Our findings suggest a novel pro-metastatic role of low BMI-induced NPY in the progression of brain metastasis. We emphasize the importance of elucidating the clinical implications of this relationship to develop an updated intervention strategy for the clinical care of patients with lung cancer and low BMI.

Citation Format: Abhishek Tyagi, Shih-Ying Wu, Ravindra Deshpande, Kerui Wu, Liang Liu, Kounosuke Watabe. Low body mass index (BMI) induces neuronal NPY and promotes brain metastasis of 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 5515.
Abstract 5524: Identification of PAEP as an immune activity modulator to mediate the immune resistance in tumor metastasis
Authors: Buenaventura Rand Gabriel M. , Chen Weiping , Zhang Shuling , Mock Beverly , Merlino Glenn , Yu Yanlin
Published: March 2024

Most cancer-related deaths are associated with the complex phenotypic behavior of metastasis, a process in which tumor cells spread from their initial site to distant sites. To successfully metastasize, the tumor cell will be required to make numerous adjustments and survive a series of stages involving intravasation, circulation in the blood or lymph system, extravasation, and growth at distant organs, and overcome the challenges of immunosurveillance. The tumor microenvironment is the ecosystem that surrounds and feeds the tumor. The molecular mechanism by which the metastatic tumors survive and escape from immune attack in the tumor microenvironment remains unclear. To study the molecular mechanism of interaction between metastatic tumors and the immune system in metastasis, we established immune-resistant metastatic models. We identified that PAEP, a progestogen-associated endometrial protein, is significantly upregulated in immune-resistant metastatic tumors compared with non-immune-resistant tumors. PAEP is a glycoprotein that inhibits cell immune function and plays an essential role in the pregnancy process. To examine the function of PAEP in tumor metastasis, we first introduced PAEP into non-immune resistant and poorly metastatic melanoma and rhabdomyosarcoma (RMS) cells. We found that overexpression of PAEP significantly promotes tumor metastatic potential of both melanoma and RMS cells in immunocompetent mice. Interestingly, there was no difference in metastasis between the control and overexpression groups in immunodeficient mice. These data suggest that PAEP may mediate tumor metastasis relating to host immunity. To further confirm the function of PAEP in tumor metastasis, we use CRISPR/Cas9 technology to knockout endogenous PAEP and will test the metastatic potential in different hosts. We will also investigate the molecular mechanisms using RNA sequencing and Digital Spatial Profiling (DSP) to analyze the metastatic samples. Our study will uncover a new mechanism for how tumors survive in the microenvironment and identify a novel target for the treatment of metastatic diseases.

Citation Format: Rand Gabriel M. Buenaventura, Weiping Chen, Shuling Zhang, Beverly Mock, Glenn Merlino, Yanlin Yu. Identification of PAEP as an immune activity modulator to mediate the immune resistance in tumor metastasis [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 5524.
Abstract 5527: Spatiotemporal analysis of growth factor regulation of osteosarcoma AKT and ERK signaling dynamics in an ex vivo lung metastasis model
Authors: Makkawi Rawan , Roberts Ryan D. , Copperman Jeremy , Davies Alexander E.
Published: March 2024

In the present study, we investigated the relationships between spatiotemporal distribution of growth factors in the lung microenvironment, metastatic osteosarcoma cell signaling, and intratumoral gene expression heterogeneity. Osteosarcoma is an aggressive bone malignancy that commonly metastasizes to the lungs, resulting in significant patient mortality. During metastasis, disseminated cancer cells are exposed to microenvironment-derived growth factors that result in signaling pathway activation. Two such pathways are MAPK/ERK and PI3K/AKT, which regulate complex transcriptional programs underlying cell proliferation and survival decisions. In this study, we utilized precision cut lung slices (PCLS) obtained from mice with established osteosarcoma metastases to ex vivo model associations between the spatiotemporal distributions of growth factors in the lung, osteosarcoma signaling dynamics, and intratumoral gene expression heterogeneity, with single cell precision. To accomplish this, we engineered osteosarcoma cells that co-expressed genetically-encoded ERK and AKT sensors, engrafted them into PCLS tissues, then recorded sensor dynamics via live-cell microscopy. These tissues were subsequently fixed and immuno-stained for measurement of growth factor expression in resident lung cells and spatial correlation with observed tumor cell ERK-AKT signaling dynamics. Using this approach, we found that growth factor expression was diffusely increased within areas bordering osteosarcoma metastases. FGF2 and IGF1, which we have previously demonstrated to potently induce ERK signaling in osteosarcoma cells, were both significantly upregulated in these regions. However, despite relatively uniform overexpression of growth factors adjacent to tumors, individual osteosarcoma cells displayed stochastics patterns of ERK and AKT signaling, and downstream gene targets. These findings revealed that upregulation of lung growth factors, and the inherent systems-level properties of osteosarcoma signaling networks, can converge to promote intratumoral signaling and gene expression heterogeneity. Together, our results, using a novel PCLS and live-cell biosensor imaging approach, provide new insight into how spatial and temporal changes in microenvironmental signaling composition are integrated by metastatic osteosarcoma cells to promote heterogeneity.

Citation Format: Rawan Makkawi, Ryan D. Roberts, Jeremy Copperman, Alexander E. Davies. Spatiotemporal analysis of growth factor regulation of osteosarcoma AKT and ERK signaling dynamics in an ex vivo lung metastasis model [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 5527.
Abstract 553: Assessment of CNTN4 expression and its association with PD-L1 across 18 various cancers using an artificial intelligence-powered immunohistochemistry analyzer
Authors: Shin Seunghwan , Moon Jimin , Cho Soo Ick , Ock Chan-Young , Jeon Bu-Nam , Kim Gyeongyeon , Yu Hyunkyung , Cha Miyoung
Published: March 2024

Background: Contactin-4 (CNTN4) has been identified as an inhibitory immune checkpoint protein and mainly expressed on tumor cells. It interacts with APP present in T cells, leading to a reduction in T cell proliferation, activation, and responsiveness to tumors. In this study, we applied artificial intelligence (AI)-powered analyzers to examine the expression of CNTN4 and its association with PD-L1 in various solid tumors.

Methods: The AI-powered immunohistochemistry (IHC) analyzers were developed using pan-cancer whole slide images (WSIs) stained with 18 different IHC types. Pathologists annotated cells and tissues on the WSIs. The universal model was trained by annotations from IHC WSI, which could detect tumor cells with four grade intensities (strong, moderate, weak, and negative) and segment invasive cancer area. Finally, the AI analyzer can count the tumor cells in an invasive cancer area and calculate the ratio based on intensity. The AI analyzer was applied to the pan-cancer tissue microarray dataset (n=795) consisting of 18 cancers to validate the analyzer. The dataset was stained with rabbit monoclonal antibody (clone EPR 8735) and PD-L1 22C3 antibody, respectively. The expression levels of CNTN4 and PD-L1 were calculated with H-score (range 0 to 300) or tumor proportion score (TPS).

Results: When positivity was defined as moderate-to-strong staining intensity in ≥ 30%, 50%, 75%, and 90%, 42.4% (337/795), 25.5% (203/795), 11.4% (91/795), and 5.2% (41/795) of samples were CNTN4 positive, respectively. In case of moderate-to-strong staining in ≥ 50% tumor cells, CNTN4 expression were most frequently observed in hepatocellular carcinoma (63.9%), endometrial carcinoma (50.0%), stomach cancer (43.9%), pancreatic cancer (40.0%), and prostate cancer (35.4%). Notably, 10% of cases with an H-score < 200 (75/757) showed a range of PD-L1 expression (PD-L1 TPS ≥ 1%), while all of the samples with an H-score ≥ 200 (n=38) were PD-L1 TPS <1%.

Conclusions: This study describes the CNTN4 expression levels in various tumors using an AI-powered IHC analyzer. CNTN4 tends to be expressed in opposition to PD-L1, making it a viable immune therapeutic target in cancers that do not conventionally express PD-L1.

Citation Format: Seunghwan Shin, Jimin Moon, Soo Ick Cho, Chan-Young Ock, Bu-Nam Jeon, Gyeongyeon Kim, Hyunkyung Yu, Miyoung Cha. Assessment of CNTN4 expression and its association with PD-L1 across 18 various cancers using an artificial intelligence-powered immunohistochemistry analyzer [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 553.
Abstract 5541: The unique metabolite landscape of the aged microenvironment dictates melanoma migratory and invasive capacity
Authors: Alicea Gretchen Marie , Patel Payal , Wei Meihan , Rebecca Vito , Wirtz Denis
Published: March 2024

Metastatic burden and organ failure are the root causes of mortality in patients with advanced melanoma; however, little is known of the metabolic drivers that enable organ colonization and the formation of overt metastases. Recent studies have drawn attention to the importance of metabolism in the survival and colonization of cancer cells. In melanoma, there have been studies focusing on the invasive capabilities of cells; however, not much is known regarding metabolic plasticity in melanoma migratory, invasive and ultimately metastatic capabilities.We have previously shown changes in the lipid landscape of the aged microenvironment promotes melanoma resistance to targeted therapy. Here, we find a distinct metabolite landscape in the aged microenvironment elevates melanoma metastatic capacity. We utilized a panel of well characterized melanoma models and tested their invasiveness and motility under different metabolite levels modeled after the aged microenvironment. Transcriptomic and proteomic analysis identifies distinct signatures that explain the migratory and invasive differences in cells cultured in varying metabolic environments. We hypothesize that different nutrient availabilities melanoma cells are exposed to during the metastatic phase affect the migratory and invasive capacity of the cell. We tested this by performing 3D migration assays, organoids and transwell assays of cells grown in media with varying nutrient levels. Further analysis is currently underway to better understand the mechanisms in which these metabolic differences facilitate cell migration, invasion and eventually colonization. This work has important implications for our understanding of the metabolic determinants of melanoma migration and metastatic colonization, and we hypothesize restraining tumor metabolic plasticity is vital to prevent and treat metastases in melanoma.

Citation Format: Gretchen Marie Alicea, Payal Patel, Meihan Wei, Vito Rebecca, Denis Wirtz. The unique metabolite landscape of the aged microenvironment dictates melanoma migratory and invasive capacity [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 5541.