bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3642: The aged lung promotes reactivation from metastatic melanoma dormancy
Authors: Fane Mitchell , Weeraratna Ashani
Published: June 2022

Our previous data have indicated that while tumors grow more slowly in aged mouse skin, they form more lung metastases. We queried whether the lung microenvironment undergoes age-related changes that provide a permissive niche for metastatic outgrowth. Our data showed that secreted factors by lung fibroblasts changed during aging that promoted proliferation of melanoma cells via expression of the tyrosine kinase receptor MER. Further investigation of this MER-high phenotype revealed that these melanoma cells induce dramatic changes to the lung microenvironment via secretion of its primary ligand PROS1, which creates a more permissive niche for metastasis. It first acts in a paracrine manner to induce proliferation of previously dormant melanoma cells in the lung. MER induced secretion of PROS1 by melanoma cells also regulates the production of extracellular matrix (ECM) from lung fibroblasts. Specifically, healthy human lung fibroblasts produce thick, unified cellular derived matrices (CDMs) in vitro that are growth-restrictive to melanoma cells. Treatment of lung fibroblasts with conditioned media from MER-High melanoma cells or with recombinant-PROS1 alters CDM density and orientation, which resulted in a CDM that dramatically increased melanoma proliferation. Finally, we found that MER induced metastatic outgrowth of melanoma cells within the lungs of mice decreased CD4 and CD8 T-cells whilst increasing infiltration of and immunosuppressive T-regulatory cells (Tregs) and Myeloid Derived Suppressor Cells (MDSCs).

Overall, we find that age-induced activation of melanoma cells towards a MER-high state within the lung can induce a cancer phenotype which dramatically alters fibroblast ECM production and immune cell infiltration to promote aggressive and efficient metastatic outgrowth and a growth permissive microenvironment for dormant melanoma cells.

Citation Format: Mitchell Fane, Ashani Weeraratna. The aged lung promotes reactivation from metastatic melanoma dormancy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3642.
Abstract 3653: The moderating role of dietary patterns on physical health and risk of cancer mortality in the REasons for Geographic and Racial Differences in Stroke (REGARDS) cohort
Authors: Aduboffour Akwasi O. , Carter Stephen J. , Howard George , Khan Saira , Lewis-Thames Marquita , Moore Justin Xavier , Moore MPH X.
Published: June 2022

Diet and physical activity share an association with cancer mortality. We investigated whether empirically-defined dietary patterns modified the association between a composite of physical health and cancer mortality.

METHODS: We performed analysis among participants from the Reasons for Geographic and Racial Differences in Stroke (REGARDS) cohort. Physical health (PH) composite was assessed by the product of baseline heart rate and systolic blood pressure, a reliable measurement of myocardial oxygen demand, and self-reported physical activity. Participant adherence was assessed with empirically-defined dietary patterns including: Convenience (including Chinese and Mexican foods, pasta, pizza), Plant-based (including fruits, vegetables), Southern (including added fats, fried foods, sugar-sweetened beverages), Sweets/Fats (including sugary foods) and Alcohol/Salads (including alcohol, green-leafy vegetables, salad dressing) was examined as an effect modifier. We performed comparisons across PH composite risk groups (i.e., severe, high, moderate, and low risk). We estimated hazard ratios (HRs) and associated 95% confidence intervals (CIs) by PH composite groups, stratified by dietary patterns.

RESULTS: Among 20,857 participants, 18% were defined as severe-risk PH, 30% were high-risk, 15% were moderate-risk PH, and 38% were low-risk PH. The most common dietary pattern adhered to by severe-risk PH participants was the southern based diet (30% adhering, p value <0.01). Severe-risk PH participants generally had higher risks for cancer mortality, corresponding to a 2.4-fold increased risk of cancer mortality (adjusted HR: 2.42; 95% CI: 1.52 - 3.85) even with limited southern diet consumption (1st quartile).

CONCLUSIONS: Dietary patterns influence cancer mortality risk; however cardiovascular health is independently associated with greater long-term risk of cancer death.

Citation Format: Akwasi O. Aduboffour, Stephen J. Carter, George Howard, Saira Khan, Marquita Lewis-Thames, Justin Xavier Moore, PhD, MPH X. Moore. The moderating role of dietary patterns on physical health and risk of cancer mortality in the REasons for Geographic and Racial Differences in Stroke (REGARDS) cohort [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3653.
Abstract 3674: Differential expression of stress-survival pathway genes related to Hispanic colorectal cancer disparities
Authors: Kulkarni Aditi , Basnet Urbashi , Patil Abhijeet R. , DeJesus Rebecca , Roy Sourav
Published: June 2022

Colorectal cancer (CRC) accounts for 12% and 8% of all estimated new cases of cancer and 11% and 9% of all cancer deaths in Hispanic men and women, respectively. Stress-survival pathways have been implicated in tumorigenesis of CRC and in providing resistance to chemotherapy-induced cell-death by suppressing reactive oxygen species (ROS), repairing DNA damage, and by regulating cell signaling pathways. However, studies related to the role of these pathways in CRC from the Hispanic health disparities perspective, are limited. Essential to improving treatment, prognosis, and detection strategies is the identification and validation of new ethnicity-specific transcriptomic markers within the stress-survival pathways as effective predictors of CRC. In this study, we have explored the role of stress-survival pathway genes in Hispanic and Non-Hispanic White (NHW) CRC tissues. We have used microarray and RNA-seq datasets obtained from the gene expression omnibus (GEO) and the cancer genome atlas (TCGA) to identify 28 genes associated with CRC. These genes were screened for transcript level expression in one normal colon and two CRC cell lines, and in cDNA arrays containing tumor (n=40) from different stages, and control (n=8) samples. The protein level expressions were evaluated by immunohistochemistry (IHC) in CRC tissue microarrays (TMAs) containing different stages of tumor (n = 108) and control (n = 12) tissues. All 28 genes were also analyzed for their transcript level expressions in Hispanic and NHW tissues. Genes related to apoptosis like FOXM1, CDK1, CDK4, Chek1, and MCM10 showed consistent upregulation in CRC cell lines, cDNA arrays and TMAs when compared to normal samples. We could also detect a correlation between the differential expression of these genes and the tumor stages. CDK4, Chek1, FOXM1, and CDC25, were observed to be downregulated in the Hispanic CRC tissues. On the other hand, oxidative stress markers were not seen to be differentially expressed in the two CRC cell lines when compared to the normal cell line. However, some of the known oxidative stress markers like PRDX4, GPX1, and SOD2 were found to be upregulated in Hispanics when compared to NHW tissues. Our results suggest that some of these stress-survival pathway genes could be explored as interesting candidates for ethnicity-specific CRC biomarkers and as novel therapeutic targets in the future.

Citation Format: Aditi Kulkarni, Urbashi Basnet, Abhijeet R. Patil, Rebecca DeJesus, Sourav Roy. Differential expression of stress-survival pathway genes related to Hispanic colorectal cancer disparities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3674.
Abstract 3680: Distinctive metabolomics profiles associated with African American current smokers who have high aggressive prostate cancer
Authors: Jun Se-Ran , Su L. Joseph , Matich Eryn , Hsu Ping-Ching
Published: June 2022

Background: Smoking has not been an established risk factor for prostate cancer (PCa), and has not been emphasized in PCa prevention. However, recent studies have shown increasing evidence that there is a higher risk of biochemical recurrence, PCa mortality, and metastasis among current smokers, presenting an urgent need in re-evaluating the association between smoking and aggressive PCa. This study aimed to determine whether smoking increase the likelihood of developing a more aggressive prostate cancer.

Methods: Equal numbers of African Americans (AAs) and European Americans (EAs) by smoking status (never/former/current) matched with PCa aggressiveness, BMI, 5-year age group, and year of baseline recruitment, totaling 480 participants, were included in the metabolomics study. For metabolomics analysis, fold change and BH-adjusted p-value from t-test adjusted for age for univariate analysis, and PCA adjusted for age and PLS-DA supervised statistical analysis for multivariate analysis were employed to decipher the underlying metabolomic patterns, and identify significantly dysregulated metabolites for the variables of interest.

Results: AA participants were significantly younger (mean=61.4, SD=7.7) compared with EAs (mean=63.5, SD=7.5). Current smokers had a 2.4 times higher risk of high aggressive PCa. When stratified by race, the risk diminished for EAs but increased for AAs. Global metabolic profiles detected a total of 1,487 compounds of known identity. After excluding metabolites with missing values in more than 20% of the samples and with small standard variation, we observed a distinct cluster of participants from AA aggressive PCa patients and current smokers that were separated from EAs and never smokers. With BH-adjusted p-value < 0.05 and fold change > 2, we identified 10 significantly dysregulated metabolites between AA and EA among high aggressive PCa and current smokers. Further, 36 metabolites between current and never smokers among AA high aggressive PCa were significantly dysregulated, but none of them are annotated as tobacco metabolites.

Conclusion: Our study presented distinctive metabolomics profiles specific to AA current smokers who had high aggressive PCa. Furthermore, the distinctive patterns were not driven by the tobacco metabolites, with the potential to identify metabolites that might help to understand the relationships between smoking and aggressive PCa in AA.

Citation Format: Se-Ran Jun, L. Joseph Su, Eryn Matich, Ping-Ching Hsu. Distinctive metabolomics profiles associated with African American current smokers who have high aggressive prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3680.
Abstract 37: Understanding the social determinants of guideline-based mammography adherence across race/ethnicity: Results from the 2016 California Health Interview Survey
Authors: Johnson Roman , Thomeer Mieke Beth
Published: June 2022

Purpose: There is not a consensus concerning recommendations about breast cancer screening which has generated confusion about when and how often to undergo mammography screening in the population, yet there is limited population-based data about the extent to which patients adhere to various mammographic screening guidelines in practice. Our purpose was to evaluate population-based adherence to mammographic screening using criteria from major guideline-producing organizations across race/ethnicity to understand what social determinants of mammography adherence are associated with each guideline producing institution.

Methods: Women aged 45-79 in the 2016 California Health Interview Survey were included. Self-reported mammographic screening within 1 or 2 years, according to major guideline-producing organizations (American Cancer Society [ACS], US Preventative Services Task Force [USPSTF], American College of Radiology [ACR], American College of Obstetricians and Gynecologists [ACOG]) was calculated with logistic regression, adjusting for demographics, health care, insurance status, and BMI.

Results: 7,551 women were included in this study. By age category, cross-sectional adherence to ACR/ACOG (annual screening) (65 to 67%) and USPSTF guidelines (biennial screening) ranged from 82-83% and with increasing age being generally associated higher adherence. The highest proportions of women undergoing mammographic screening were seen in women ages 60-69 (67% within last year, 85% within last 2 years). Statistically significant predictors of adherence to mammography screening included Latina race (OR 1.62, 0.18 SE), full time employment (OR 0.73, 0.09 SE), and not having a usual healthcare provider (OR 0.50, 0.12 SE), adjusted for demographics, health care, insurance status, and BMI.

Conclusion: Frequency of screening increases with age with highest screening proportions in women ages 65-69 (66% within last year, 81% within last 2 years). For ACR/ACOG screening guidelines, adherence to mammography screening remains poorer in women with no usual source of care, given that their odds of exhibiting adherence was half that of women who did have a regular source of care and nearly 65 percent lower odds of those women with USPSTF mammography adherence.

Key Words: Breast cancer, screening, race/ethnicity, guideline-based care

Citation Format: Roman Johnson, Mieke Beth Thomeer. Understanding the social determinants of guideline-based mammography adherence across race/ethnicity: Results from the 2016 California Health Interview Survey [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 37.
Abstract 3707: Moonlighting function of mitochondrial protein VDAC2 provides a potent pharmacological strategy against selective killing of hepatocellular carcinoma
Authors: Mishra Piyush
Published: June 2022

Voltage dependent anion channels (VDACs) family of proteins are the most abundant proteins in the outer mitochondrial membrane (OMM). There are three isoforms of VDAC, that are structurally and functionally conserved as far as their ion transport function is concerned. However, there are isoform-specific functions that support distinct pathways. For VDAC2 isoform, an established isoform-specific function is the targeting of the pro-apoptotic Bak protein to the OMM, which stabilizes Bak, leading to its elevated cellular levels. On the surface of OMM, activation and oligomerization of Bak and Bax, another pro-apoptotic protein leads to OMM perforation and release of pro-apoptotic proteins from intermembrane space to the cytoplasm. This is an irreversible step that stimulates the downstream events of apoptotic pathway. Hepatocellular carcinoma (HCC) ranks third in cancer mortality and in the advanced stages is associated with poor prognosis. We observed that unexpectedly, HCC cells are vastly more sensitive to tBid induced apoptosis compared to normal hepatocytes. tBid is a BH3-only pro-apoptotic Bcl-2 family protein, the effect of which is primarily mediated by Bak. We have shown that, in the absence of VDAC2, the Bid-Bak apoptosis pathway is suppressed. VDAC2 protein expression is the lowest in liver and lung among different tissues. We also determined the protein abundances in the HCC lines and normal hepatocytes and found elevated VDAC2 and Bak levels in HCC. Our findings are in line with multiple datasets that show an elevation of VDAC2/Bak expression in human HCC suggesting a potential clinical relevance. We found that combination of an inhibitor of the anti-apoptotic proteins (S63845, a selective Mcl-1 inhibitor), which normally suppresses Bak activation, with an activator of the tBid pathway (TRAIL) enhanced the death of hepatocarcinoma cells with little effect on normal hepatocytes. This treatment strategy was also effective at reducing tumor growth in vivo, but only in tumors expressing VDAC2. Thus, we postulate that increased VDAC2 expression in HCC leads to elevated Bak recruitment to the OMM, which in turn sensitizes HCC to tBid. Therefore, this differential expression provides an excellent opportunity for selective elimination of HCC cells.

Citation Format: Piyush Mishra. Moonlighting function of mitochondrial protein VDAC2 provides a potent pharmacological strategy against selective killing of hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3707.
Abstract 3796: Deeper insight into intratumoral heterogeneity by MRI and PET-guided stereotactic biopsies from glioblastoma patients
Authors: Anand Atul , Petersen Jeanette Krogh , Burton Mark , Larsen Martin Jakob , Andersen Lars van , Harwood Dylan Scott , Pedersen Christian Bonde , Poulsen Frantz Rom , Grupe Peter , Kruse Torben A. , Thomassen Mads , Kristensen Bjarne Winther
Published: June 2022

Glioblastoma is one of the most aggressive cancers, but the molecular evolution is not fully understood. We used PET imaging combined with deep sequencing of glioblastoma biopsies at both the RNA and DNA levels to get a deeper insight into molecular evolution. In the clinical setting, PET imaging provides information about metabolically active tumor areas, but the molecular interpretation is unclear. Our primary objective was to perform an intratumoral spatial comparison of biopsies from potentially aggressive and less aggressive areas in glioblastomas according to PET scans. We used MRI co-registered with 11C-MET-PET (amino acid) and 18F-FDG PET (glucose) in order to obtain representative neurosurgical stereotactic biopsies from tumor areas with high accumulation of these two metabolites (hotspot), low accumulation (coldspot), and no accumulation as the periphery of six glioblastoma patients that were processed for whole genome, exome, and transcriptome sequencing. Differential gene expression and gene ontology analysis showed that hotspots were enriched in gene sets associated with DNA replication, cell cycle, and ligand-receptor interaction. Genome and exome analysis suggested hotspots and coldspots have similar mutational profiles. However, a limited number of hotspot-specific mutations and novel fusion transcripts indicated that hotspot-associated tumor cells developed from coldspot-associated tumor cells and point at the potential role of hotspot driver genes in glioblastoma evolution. Our findings reveal that hotspots in glioblastomas represent a more advanced stage of molecular evolution than coldspots.

Citation Format: Atul Anand, Jeanette Krogh Petersen, Mark Burton, Martin Jakob Larsen, Lars van Andersen, Dylan Scott Harwood, Christian Bonde Pedersen, Frantz Rom Poulsen, Peter Grupe, Torben A. Kruse, Mads Thomassen, Bjarne Winther Kristensen. Deeper insight into intratumoral heterogeneity by MRI and PET-guided stereotactic biopsies from glioblastoma patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3796.
Abstract 3821: Rapid acceleration of KRAS-mutant pancreatic carcinogenesis via remodeling of tumor immune microenvironment by PPARD
Authors: Liu Yi , Deguchi Yasunori , Wei Daoyan , Moussalli Micheline J. , Liu Fuyao , Deguchi Eriko , Li Donghui , Wang Huamin , Valentin Lovie Ann , Colby Jennifer K. , Wang Jing , Zheng Xiaofeng , Ying Haoqiang , Gagea Mihai , Ji Baoan , Shi Jiaqi , Yao James C. , Zuo Xiangsheng , Shureiqi Imad
Published: June 2022

Pancreatic intraepithelial neoplasia (PanIN) is a precursor of pancreatic ductal adenocarcinoma (PDAC), which commonly occurs in the general populations with aging. Although most PanIN lesions (PanINs) harbor oncogenic KRAS mutations that initiate pancreatic tumorigenesis, PanINs rarely progress to PDAC. Critical factors that promote this progression, especially targetable ones, remain poorly defined. We show that peroxisome proliferator-activated receptor-delta (PPARD), a lipid nuclear receptor, is upregulated in PanINs in humans and mice. Furthermore, PPARD ligand activation by a high-fat diet or GW501516 (a highly selective, synthetic PPARD ligand) in mutant KRASG12D (KRASmu) pancreatic epithelial cells strongly accelerates PanIN progression to PDAC. This PPARD activation induces KRASmu pancreatic epithelial cells to secrete CCL2, which recruits immunosuppressive macrophages and myeloid-derived suppressor cells into pancreas via the CCL2/CCR2 axis to orchestrate an immunosuppressive tumor microenvironment and subsequently drive PanIN progression to PDAC. Our data identify PPARD signaling as a potential molecular target to prevent PDAC development in subjects harboring PanINs.

Citation Format: Yi Liu, Yasunori Deguchi, Daoyan Wei, Micheline J. Moussalli, Fuyao Liu, Eriko Deguchi, Donghui Li, Huamin Wang, Lovie Ann Valentin, Jennifer K. Colby, Jing Wang, Xiaofeng Zheng, Haoqiang Ying, Mihai Gagea, Baoan Ji, Jiaqi Shi, James C. Yao, Xiangsheng Zuo, Imad Shureiqi. Rapid acceleration of KRAS-mutant pancreatic carcinogenesis via remodeling of tumor immune microenvironment by PPARD [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3821.
Abstract 3839: RASSF1C and tumor microenvironment
Authors: Amaar Yousef G. , Reeves Mark E.
Published: June 2022

Introduction: Tumor microenvironment (TME) plays a vital role in tumor invasion and metastasis and provides a rich environment for identifying novel therapeutic targets. The TME landscape consists of extracellular matrix (ECM) and stromal cells. ECM is a major component of TME that mediates interaction between cancer cells and stromal cells to promote invasion and metastasis. We have shown in published work that RASSF1C promotes cancer stem cell development, migration, and drug resistance, in part, by promoting EMT through a mechanism that involves up-regulation of the PIWIL1-piRNA gene axis. Interestingly, one of the target genes identified by our microarray study to be upregulated by RASSF1C is P4HA2. In cancer, P4H2A is vital for collagen posttranslational modification and folding. This leads to formation of a stiff ECM and induction of cancer stem cell marker gene expression, resulting in metastatic dissemination. High expression of P4HA2 is associated with significantly lower survival of lung cancer patients. Thus, RASSF1C could promote tumor cell ECM remodeling to induce lung cancer cell stemness, invasion and metastasis by up-regulating a novel PIWIL1-P4HA2 gene axis expression.

Methods: Analysis of a microarray study using H1299 cells over-expressing RASSF1C or control vector backbone identified P4HA2 as a RASSF1C gene target. RT-PCR, immunoblots, and immunofluorescence techniques were used to confirm P4HA2 gene expression using specific primers and antibodies in cells over-expressing RASSF1C or PIWIL1 along with control cells. Kaplan-Meier analysis was performed to determine the relationship of P4HA2 expression to lung adenocarcinoma patient survival using data from The Cancer Genome Atlas (TCGA).

Results: Analysis of microarray data from H1299 cells over-expressing RASSF1C or control vector backbone shows that RASSF1C up-regulates P4HA2 gene expression 2-fold. The up-regulation of P4HA2 expression by RASSF1C was confirmed by RT-PCR, immunoblots, and immunofluorescence using P4HA2 gene specific primers and antibodies. Further, H1299 cells over-expressing PIWIL1 show increased P4HA2 gene expression on immunoblots. Kaplan-Meier analysis shows that high P4HA2 expression in lung adenocarcinoma patients negatively correlates with patient survival. Thus, our findings suggest the hypothesis that RASSF1C may promote lung cancer cell ECM remodeling to induce lung cancer cell stemness, invasion and metastasis, in part, by up-regulating a novel pathway involving a PIWIL1-P4HA2 gene axis.

Conclusions: Investigating the role of a RASSF1C-PIWIL1-P4HA2 gene axis in ECM remodeling in lung cancer cells will provide important mechanistic information that could, in turn, lead to identification of useful biomarkers for lung cancer prognosis and targets for therapy.

Citation Format: Yousef G. Amaar, Mark E. Reeves. RASSF1C and tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3839.
Abstract 3843: TIMP1 and TIMP2 re-educate TGFβ induced pro-inflammatory/pro angiogenic decidual-like NK cells phenotype
Authors: Albini Adriana , Gallazzi Matteo , Palano Maria Teresa , Carlini Valentina , Calabrone Luana , Ricotta Riccardo , Stetler-Stevenson William G. , Noonan Douglas M.
Published: June 2022

Background: Tissue inhibitors of metalloproteases (TIMPs) have been reported to exert dual roles in cancer, based on the different cancer type and interactions with the tumor microenvironments. As inhibitors of Matrix metalloproteases (MMP) they downregulate invasion, metastasis and angiogenesis. However, TIMP1 levels have been found increased in patients with different types of cancer. Natural Killer cells have been found to be phenotypically and functionally compromised in several solid and hematological cancers. We were the first in characterizing NK pro-angiogenic phenotype in the peripheral blood and tumor tissues of patients with Non-small Cell Lung cancer, colorectal cancers, prostate of patients. Also, we found that pro-angiogenic NK cells in the peripheral blood of colorectal cancer patients release of TIMP-1 and TIMP-2, in a STAT3/STAT5-independent manner. Here, we investigated the effects of TIMP-1 and TIMP-2 recombinant proteins, using a TGFβ-mediated NK cell polarization model, in vitro.

Methods: We investigated the effects of TIMP1 and TIMP2 recombinant proteins in hindering decidual-like markers in NK cells, generated by polarizing cytolytic NK cells with TGFβ. The effects of TIMP1 or TIMP2 on NK cell surface antigens were determined by multicolor flow cytometry.

Results: We found that TIMP1 and TIMP2 were effective in interfering with TGFβ induced NK cell polarization towards a decidual-like-phenotype. TIMP1 and TIMP2 counteracted the effect of TGFβ in increasing the percentage of CD56bright, CD16-, CD9+ and CD49a+, and restoring normal levels for TIMP 1 and 2 also inhibited decrease levels of the activation marker NKG2D induced by TGFβ and decreased the TGFβ upregulated exhaustion marker TIM-3. NK cell degranulation capabilities against K562 cells were also decreased by TGFβ and not by TIMP1 or TIMP2. TIMP1 treatment could partially restore degranulation marker CD107a expression.

Conclusions: Our results suggest a potential role of TIMPs in controlling the tumor-associated cytokine TGFβ-induced NK cell polarization. TGFβ stimulation represents a model to prove TIMP's new properties, further studies will help defining TIMPs immunomodulatory activities of NK cells in cancer, and their possible future diagnostic-therapeutic roles.

Citation Format: Adriana Albini, Matteo Gallazzi, Maria Teresa Palano, Valentina Carlini, Luana Calabrone, Riccardo Ricotta, William G. Stetler-Stevenson, Douglas M. Noonan. TIMP1 and TIMP2 re-educate TGFβ induced pro-inflammatory/pro angiogenic decidual-like NK cells phenotype [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3843.
Abstract 385: iRGD mediated delivery of neoantigens to enable immunotherapy in integrin b5-rich tumors
Authors: de Mendoza Tatiana Hurtado , Mose Evangeline S. , Botta Gregory P. , Braun Gary B. , Kotamraju Venkata R. , French Randall P. , Suzuki Kodai , Miyamura Norio , Sun Siming , Patel Jay , Teesalu Tambet , Ruoslahti Erkki , Sugahara Kazuki N. , Lowy Andrew M.
Published: June 2022

Pancreatic duct adenocarcinomas are known for their abundant desmoplastic stroma that acts as a barrier for drug penetration and reduces treatment efficacy. iRGD is a tumor-penetrating peptide that initially targets αv integrins expressed on tumor vasculature with its RGD motif and then is proteolytically processed to expose a CendR motif (R/KXXR/K) that interacts with neuropilin-1 (NRP-1), leading to extravasation. We investigated the mechanism of iRGD tissue penetration and found that iRGD initially targets Carcinoma Associated Fibroblasts (CAFs) and then spreads to the tumor cells in a time dependent manner. CAF targeting was dependent on integrin β5 expression and CAFs induced upregulation of integrin β5 in the adjacent tumor cells in a TGF- β dependent manner. Drugs conjugated or co-administered with iRGD can penetrate deep into tumor tissue, significantly increasing the efficacy of chemotherapeutic agents in a variety of solid tumors. Our recent data shows that KrasLSL-G12D/+ Trp53LSL-R172H/+ Pdx1-Cre (KPC) mice treated with iRGD co-administered with Gemcitabine increased survival compared to drug alone. In addition, we are using iRGD to deliver neoantigens to breast and pancreatic cancers to enable immunotherapy. These tumors have a low mutational burden and a very immunosuppressive microenvironment, rendering them resistant to such therapies. We have used iRGD, to deliver the ovalbumin 257-264 (OVAI) peptide to triple negative breast tumors, followed by adoptive T cell transfer of OT1 CD8 T cells, in order to elicit an antitumor immune response. Our preliminary data showed tumor regression in 70%, and complete response in 42% of the mice treated with iRGD plus OVA1. We are now working on adapting this strategy to pancreatic cancer.

Citation Format: Tatiana Hurtado de Mendoza, Evangeline S. Mose, Gregory P. Botta, Gary B. Braun, Venkata R. Kotamraju, Randall P. French, Kodai Suzuki, Norio Miyamura, Siming Sun, Jay Patel, Tambet Teesalu, Erkki Ruoslahti, Kazuki N. Sugahara, Andrew M. Lowy. iRGD mediated delivery of neoantigens to enable immunotherapy in integrin b5-rich tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 385.
Abstract 3861: Microscope-based sorting of highly motile cancer-associated fibroblasts using a photoconvertible fluorescent protein
Authors: Lee Sieun , Ahn Young-Ho
Published: June 2022

In the tumor microenvironment, cancer-associated fibroblasts (CAFs) are known to play key roles in promoting cancer cell invasion and cancer progression. CAFs are also highly invasive compared with normal fibroblasts in a 3-D co-culture system with lung cancer cells. Due to CAF heterogeneity, there are various differences in cell motility even within a CAF population. To classify CAFs according to their motility, we named less motile CAFs as “static CAFs” and more motile CAFs as “motile CAFs”. To separate and isolate these static CAFs and motile CAFs from a 3-D co-culture system, CAFs were transfected with a green-to-red photoconvertible fluorescent protein, Dendra2. Dendra2-transfected CAFs at specific areas (i.e., static or motile CAFs) were UV-irradiated using a confocal microscope. CAFs with red fluorescence were then isolated by flow cytometry and were subjected to RNA sequencing. In gene ontology and gene set enrichment analyses, two types of CAFs showed significant differences in global gene expression patterns. These data suggest that heterogeneous CAFs with different motility have distinct roles in the regulation of various signaling pathways in the tumor microenvironment.

Citation Format: Sieun Lee, Young-Ho Ahn. Microscope-based sorting of highly motile cancer-associated fibroblasts using a photoconvertible fluorescent protein [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3861.
Abstract 3869: A human in-vitro T cell exhaustion model for assessing immuno-oncology therapies
Authors: Goral Paulina , Brackenbury Louise , Nunan Robert , Jenkinson Rhiannon
Published: June 2022

Chronic stimulation of CD8 T cells either as a result of viral infection or within the tumor microenvironment can lead to a state of exhaustion, whereby T cells no longer make a productive effector response. Remobilisation of exhausted tumor specific CD8 T cells (TEX) presents a possible therapeutic strategy for driving anti-tumor T cell responses. The success of biologics targeting receptors and ligands associated with exhausted T cells such as PD-1/PD-L1 and CTLA-4 has spurred an acceleration of research to unravel the molecular processes that drive or maintain the exhausted state with the goal of developing treatments that target them. Effective therapeutic development necessitates reliable human in-vitro models that closely mimic disease conditions. To achieve this we adapted an in-vitro TEX model via application of flowing cytometry and time-lapse imaging techniques to kinetically assess TEX cell phenotype and function. Persistent and repeated rounds of polyclonal human CD8 T cell stimulation recapitulated distinctive characteristics of exhausted T cells including high expression of checkpoint receptors and upregulation of transcription factors known to suppress immune activation. Over 8 days and 4 rounds of stimulation TEX cells lost effector capabilities, producing lower levels of pro-inflammatory cytokines and became refractory to re-stimulation through the TCR. In functional cytotoxicity assays TEX cells exhibited substantial deficits in target killing compared to donor matched Teff cell counterparts. Checkpoint receptor blockade partially restored Teff function by enhancing immune mediated target cell death. Overall, this in-vitro TEX assay models key aspects of CD8 T cell exhaustion and offers a versatile and accessible platform to test novel therapeutics targeting exhaustion pathways either individually or in combination with checkpoint inhibitors to test if they fully restore effector function.

Citation Format: Paulina Goral, Louise Brackenbury, Robert Nunan, Rhiannon Jenkinson. A human in-vitro T cell exhaustion model for assessing immuno-oncology therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3869.
Abstract 3873: Profiling the tumor microenvironment within clear cell renal cell carcinoma by single-nuclei RNA-Seq
Authors: Liu Ruiyang , Strunilin Ilya , Wu Yige , Karpova Alla , Chen Siqi , Mo Chia-Kuei , Li Yize , Chen Feng , Ding Li
Published: June 2022

Clear cell renal carcinoma (ccRCC) appears as one of the most immune infiltrated tumors in pan-cancer comparisons. A systematic characterization of the immune microenvironment of ccRCC is important for understanding the disease progression and would be beneficial to predicting disease outcomes. To better understand the cellular components within the tumor microenvironment (TME), the interactions among different cell types, and the contribution to disease outcomes, we collected 34 ccRCC tumor samples for single-nuclei RNA-Seq (snRNASeq) profiling. We obtained 161312 cells in total, including 6791 stroma cells (endothelial cells, fibroblasts and myofibroblasts) and 38918 immune cells (lymphoid and myeloid populations).Analysis of cell-cell-interactions suggests tumor cells interact most frequently with stroma and immune cells. Of interest, we noticed some ligand-receptor pair interactions commonly found among all samples, such as VEGFA from tumor cells with VEGFR1/2 from endothelial cells, suggesting the presence of angiogenesis. Within endothelial populations, we found a common interaction between ANGPT2 and TEK, whose interaction scores are moderately correlated with lymphocyte percentages across samples, suggesting this interaction could contribute to endothelium leakage and lymphocytes recruitment. To better understand the immune cell subtype/states within each immune cell type, we further distinguished 14 different subpopulations within macrophages according to key marker expressions. The presence of the subpopulations suggest diverse states and heterogeneous functions within macrophages. Of note, the expression of iron uptake receptor TFRC is elevated in higher-grade tumors. Higher TFRC expression is associated with poorer prognosis, which is also validated by the CPTAC discovery cohort using bulk proteomics data. In summary, our study showcases the potential of using snRNA-Seq to profile the TME of ccRCC, in identifying important cell-cell interactions and predicting disease outcomes.

Citation Format: Ruiyang Liu, Ilya Strunilin, Yige Wu, Alla Karpova, Siqi Chen, Chia-Kuei Mo, Yize Li, Feng Chen, Li Ding. Profiling the tumor microenvironment within clear cell renal cell carcinoma by single-nuclei RNA-Seq [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3873.
Abstract 3875: De novo spatial proteomic profiling of immune synapses using machine learning-guided microscoop
Authors: Liu You-Pi , Chong Weng Man , Huang Harry , Chen Yi-De , Chung Chia-Wen , Chang Hsiao-Jen , Chang Chih-Wei , Liao Jung-Chi
Published: June 2022

The spatial proteome of the immune synapse (IS) between a target cell and a lymphocyte is fundamentally important to understand the mechanism of cell-mediated immunity for both immuno-oncology and therapeutic applications. In this research, we used Microscoop, a fully-automatic microscope system integrated with a machine learning-based algorithm, to best determine ISs for proteomic mapping. We used Raji B cells as antigen-presenting cells (APCs) and induced the formation of ISs by incubating with Jurkat T cells. Multiple IS images were applied to train our algorithm using convolution neural network-based deep learning. A sequential process including fluorescence imaging, deep learning-enabled pattern generation, and photochemical labeling was implemented to achieve spatial-specific biotinylation of the IS proteins. Moreover, Microscoop is capable to repeat the process automatically on thousands of fields of view to label sufficient immune-synaptic proteins with biotin-tag for further enrichment allowing protein identification with mass spectrometry. We have successfully labeled and isolated proteins from spatially reorganized interfaces between T cells and APCs. Following MS-based proteomic analysis, several hundreds of proteins were identified, including the proteins known to be specifically associated with T-cell receptor (TCR) activation such as LCK, one of the major factors involved in TCR signaling at ISs. More interestingly, we identified several proteins novel for ISs, including proteins involved in phosphatidylinositol signaling. Our data showcases the capability of subcellular de novo spatial proteomics of the Microscoop technology, revealing novel factors responsible for initiating the immune response of a lymphocyte and shedding light on immune checkpoint signaling and tumor immunotherapy.

Citation Format: You-Pi Liu, Weng Man Chong, Harry Huang, Yi-De Chen, Chia-Wen Chung, Hsiao-Jen Chang, Chih-Wei Chang, Jung-Chi Liao. De novo spatial proteomic profiling of immune synapses using machine learning-guided microscoop [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3875.
Abstract 3879: High dimensional analysis of tumor microenvironment and heterogeneity in FIGO IB endometrial carcinomas using imaging mass cytometry
Authors: Lien Hilde Eide , Berg Hege Fredriksen , Hoivik Erling Andre , Trovik Jone , Haldorsen Ingfrid Salvesen , Akslen Lars Andreas , Krakstad Camilla
Published: June 2022

Patients with localized endometrial carcinomas (FIGO IA and IB) have a 5-year relative survival rate of > 90%. Still, some patients with an expected good prognosis experience recurrence of disease which dramatically reduces survival rates. Currently, we lack the knowledge to predict which patients will get recurrence. We used Imaging Mass Cytometry to examine 27 proteins in tissue microarrays of 36 primary FIGO IB endometrial carcinomas, of which 17 later recurred. High dimensional images were segmented to extract single-cell information for each tumor. Distinct epithelial-, stromal- and immune cell types of the tumor microenvironment were identified and phenotyped, using protein expression intensities and unsupervised cell clustering. Neighboring cells of CD8+ cells were extracted and phenotyped. In non-recurrent tumors, more cytotoxic T cells were identified adjacent to two epithelial cell populations characterized by high vimentin and progesterone receptor expression (p < 0.01 and p = 0.02). Tumors that were enriched for vimentin showed significantly better recurrence-free survival than tumors with low vimentin or high alpha-smooth muscle actin (a-SMA) signal (p = 0.026). This study reveals distinct characteristics in the microenvironment of low stage tumors that may aid in identifying high-risk patients. Further, the study provides new insight on the pathogenic processes involved in tumor progression of endometrial cancer.

Citation Format: Hilde Eide Lien, Hege Fredriksen Berg, Erling Andre Hoivik, Jone Trovik, Ingfrid Salvesen Haldorsen, Lars Andreas Akslen, Camilla Krakstad. High dimensional analysis of tumor microenvironment and heterogeneity in FIGO IB endometrial carcinomas using imaging mass cytometry [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3879.
Abstract 3881: Immune profiling of advanced prostate cancer harboring homologous recombination deficiency
Authors: Soleimani Maryam , Kung Sonia , Saxena Neetu , Fazli Ladan , Nappi Lucia
Published: June 2022

Background: Most patients with metastatic prostate cancer (mPCa) do not derive benefit from immune checkpoint inhibitors (ICI). Homologous recombination deficiency (HRD) affects up to 6-8% of patients with localized PCa and up to 30% of patients with metastatic castrate-resistant prostate cancer (mCRPC) and is predictive of response to PARP inhibitors. The potential activity of ICI in this subset of patients is unknown and the tumor microenvironment (TME) associated with HRD is poorly understood. We used PCa tumors of patients with known HRD alterations as a model to study clinically relevant, therapeutically targetable drivers of tumor immunology.

Methods: Archival tumor tissue of patients with known germline or somatic HRD alterations enrolled in our institutional GU Biobank (n=13) was used for immunohistochemistry to evaluate the expression of the following immune markers in both tumor cells (TC) and stromal lymphocytes (SL): CD8, adenosine receptor 2a (A2aR), GAL9, IL-2, LAG3, PD-L2, and TIM-3. The same markers were analyzed in untreated (n=104), neoadjuvant hormone therapy treated (NHT) (n=16), neuroendocrine (NEPC) (n=41) and CRPC (n=22) tissue. Two-tailed t-test and Pearson R correlations were performed for analysis.

Results: Among screened HRD alterations,BRCA1/2 were the most frequently identified (BRCA1/2n=6, FANCCn=1, FANCD2=2, PALB2=1, CDK12n=1, ATRn=2). CD8+ cells were primarily localized in the stroma of HRD cohort specimens and were more abundant compared to NEPC (p≤ 0.05) and untreated tumors (p≤0.01). The HRD cohort had higher levels of A2aR, in TC and SL (p≤0 .0001 for both) and higher levels of PD-L2 in TC (p≤0.0001) compared to all other cohorts and in the SL (p≤0.05) compared to untreated, CRPC and NEPC cohorts. Lower levels of LAG3 were found in the HRD cohort TC compared to NHT cohort, and in HRD cohort SL compared to all cohorts (p≤0.0001 for both). There was no difference in TIM-3 or GAL9 expression.

Conclusions: Evaluation of stromal and tumor cell immune profiling indicates that compared to other subtypes, PCa with HRD alteration have a unique immune profile abundant in CD8+ cell, and PD-L2 and A2aR expression. Not only does this indicate an immune active TME but also identifies therapeutically actionable immunotherapeutic pathways which may render clinical benefit in this patient population.

Citation Format: Maryam Soleimani, Sonia Kung, Neetu Saxena, Ladan Fazli, Lucia Nappi. Immune profiling of advanced prostate cancer harboring homologous recombination deficiency [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3881.
Abstract 3889: Identification of ADRN-specific, MES-specific, and pan-subtype therapeutic targets in neuroblastoma
Authors: Kendsersky Nathan M. , Mabe Nathaniel W. , Farrel Alvin , Grossmann Liron D. , Odrobina Michal , Stegmaier Kimberly , Maris John M.
Published: June 2022

Background: Neuroblastoma is an extracranial solid tumor of childhood that arises from the developing sympathetic nervous system. Half of patients with high-risk disease do not survive despite multimodal therapy, and survival after relapse is unlikely. Recent studies have characterized two distinct neuroblastoma cell subtypes: the adrenergic (ADRN) subtype of sympathetic noradrenergic-like identity and the mesenchymal (MES) subtype of neural crest-like identity. ADRN-like cells are more aggressive and proliferative, while the MES-like cells are chemotherapy resistant. Neuroblastoma tumors are heterogenous and comprised of both ADRN and MES cell subtypes, which likely leads to differential responses to therapy. Thus, to eliminate cells that may lead to tumor relapse, it is critical to develop therapies against both neuroblastoma subtypes.

Methods: To identify candidate subtype-specific therapeutic targets, we used ssGSEA to classify human neuroblastoma tumors into ADRN- or MES-dominant and performed differential expression. We validated subtype-specific targets in additional datasets and models, including transdifferentiated neuroblastoma cell lines (ADRN-to-MES) after inducible overexpression of the MES transcription factor PRRX1.

Results: We show ADRN-specific expression of known preclinical and clinical neuroblastoma targets with limited or no expression in MES-dominant tumors. We propose CD276 (B7-H3) and L1CAM as pan-subtype targets with similar expression in both subtypes and stable expression after ADRN-to-MES transdifferentiation. In MES-specific neuroblastoma patient samples and cell lines, we identify the receptor tyrosine kinase AXL as a candidate MES target. MES neuroblastoma cell lines are more sensitive to small molecule AXL inhibitors (Cabozantinib, NPS-1034, and ONO-7475) compared to ADRN cell lines. We also observe higher expression of PDL1 and immunosuppressive chemokines in MES-dominant tumors and cell lines. Finally, we detect AXL and Gas6 transcripts in tumor-resident macrophages and fibroblasts, which may further promote immune evasion.

Conclusion: Here we have identified and prioritized ADRN-specific, MES-specific, and pan-subtype neuroblastoma therapeutic targets and suggest AXL-targeted therapy may eliminate both MES-dominant neuroblastoma cells and immune cell populations that contribute to an immunosuppressive microenvironment.

Citation Format: Nathan M. Kendsersky, Nathaniel W. Mabe, Alvin Farrel, Liron D. Grossmann, Michal Odrobina, Kimberly Stegmaier, John M. Maris. Identification of ADRN-specific, MES-specific, and pan-subtype therapeutic targets in neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3889.