bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1290: PTEN loss-mediated cell surface translocation of Annexin A2 promotes prostate cancer metastasis
Authors: Chaudhary Pankaj , Howard-Smith Christianna
Published: April 2023

Introduction: Metastasis is a major cause of morbidity in prostate cancer patients. Despite substantial efforts to understand prostate cancer metastasis, the mechanisms involved in preparing the metastatic niche for colonizing the prostate cancer cells are still unknown.

Methods/Materials: The normal prostate epithelial and cancer cells were purchased from American Type Culture Collection (Manassas, VA). Immunoblotting, immunofluorescence, flow cytometry, and plasmin generation assay were performed.

Results: Our results demonstrated that Pten deletion or mutation caused significantly high phosphorylation of Annexin A2 (AnxA2) at tyrosine-23 in human and murine prostate cancer cell lines compared to the normal prostate epithelial cell lines. However, we did not observe any change in total AnxA2 protein level after Pten deletion/mutation in human and murine prostate cancer cells compared to normal prostate epithelial cells. Our flow cytometry data, immunofluorescence, and membrane wash fraction analysis suggest that AnxA2 expression was higher at the cell surface of Pten null/mutated prostate cancer cell lines compared to the normal prostate epithelial cells. Confocal microscopy and immunoprecipitation analysis revealed that AnxA2 interacts with Pten in normal prostate epithelial cells and inhibits AnxA2 phosphorylation at tyrosine 23. Pten deletion/mutation-mediated translocation of AnxA2 is dependent on the phosphorylation of tyrosine 23, and mutation of tyrosine 23 to a non-phosphomimetic variant inhibits the cell surface translocation process. The cell surface-translocated AnxA2 promotes the plasmin generation in prostate cancer cells, and an AnxA2 inhibitory hexapeptide can neutralize this activity.

Conclusions: PTEN loss is the primary driving force for the translocation of AnxA2 to the cell surface and promotes plasmin generation.

Citation Format: Pankaj Chaudhary, Christianna Howard-Smith. PTEN loss-mediated cell surface translocation of Annexin A2 promotes prostate cancer metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1290.
Abstract 1312: Oncogenic mutant p53 proteins are selectively shuttled by cancer extracellular vesicles toeducate the tumor microenvironment
Authors: Bhatta Bibek , Luz Ishai , Krueger Christian , Teo Fanny Xueting , Lane David P. , Sabapathy Kanaga , Cooks Tomer
Published: April 2023

IntroductionMost cancers harbor mutations in the TP53 gene (encoding for the p53 tumor suppressor protein). Furthermore, gain-of-function (GOF) mutation in p53 imparts an aggressive traits to the cells when compared to the cancer cells harboring inactivating mutations or wild-type (WT)-p53. Notably, multiple studies have delineated the presence of GOF-mutant p53 protein in untransformed cells or in stromal compartments of tumor microenvironment (TME). In recent years, the involvement of extracellular vesicles (EVs) in cell-to-cell communication has emerged as a major route by which cancer cells can interact and educate immune, and non-immune cells in TME to become tumor supportive. To this end, we hypothesize that mutant p53 protein can be shuttled via EVs to TME cells thus shedding light on a novel non-cell autonomous role of mutant p53 cancers.MethodsEVs were isolated from various cancer cell lines (pancreas, lung, colon) differing by their p53 status and the effect on neighboring cancer cells and TME cells was studied in vitro and in-vivo. We also utilized the human colorectal Colo-320DM cancer cell xenograft model, which expresses the R248W p53 mutant. FFPE sections of subcutaneous tumors derived from the Colo-320DM xenografts were stained for p53 using the DO-1 antibody that specifically recognizes human p53.ResultsOur data demonstrated that mutant p53 protein can be selectively sorted into EVs; that mutant p53 in EVs can be taken up by neighboring cancer cells and macrophages that do not harbor mutant p53 protein. Evident of macrophage education was seen with the increased expression and secretion of pro-inflammatory cytokines. Notably, mutant p53 expression was also found in non-tumor cells in both human cancers, and in non-human tissues in human xenografts.SummaryCancer cells harboring GOF p53 mutants, can package mutant p53 proteins in EVs, and deliver them to neighboring cancer cells and to the TME.

Citation Format: Bibek Bhatta, Ishai Luz, Christian Krueger, Fanny Xueting Teo, David P. Lane, Kanaga Sabapathy, Tomer Cooks. Oncogenic mutant p53 proteins are selectively shuttled by cancer extracellular vesicles toeducate the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1312.
Abstract 1315: NETs act as immunosuppressive agents within lymph nodes during cancer
Authors: Brassard Ariane , Su Xin , Dhoparee-Doomah Iqraa , Leo Sabrina , Feng Lixuan , Bourdeau France , Giannias Betty , Larson Corissa , Qiu Qian , Spicer Jonathan , Ferri Lorenzo , Cools-Lartigue Jonathan
Published: April 2023

Metastasis is responsible for 90% of all cancer-related deaths, making it the most significant challenge in cancer treatment for clinicians worldwide. Metastasis is thought to occur in a stepwise process, in which local lymph nodes (LNs) are first colonized by tumour cells before proceeding to distal organs. However, the mechanism by which LN metastasis facilitates distal metastasis is poorly understood. LNs undergo environmental changes to accommodate tumour cell growth within, notably by shifting the environment towards immunosuppression to shut down anti-tumour immune cells. This immunosuppressive environment is critical for the establishment of LN metastases, as cytotoxic CD8 T cells will otherwise neutralize incoming circulating tumour cells. Neutrophils are among the first cells recruited to tumour-draining LNs to mediate the environmental shift, and yet their method of action has not been fully elucidated. They have recently been found to secrete NETs within LNs during cancer, which are pro-inflammatory web-like formations of DNA decorated with antimicrobial peptides. While NETs have been shown to exert pro-tumour effects in the tumour microenvironment and to facilitate metastasis as a whole, their role in LNs has never been explored. Using a NETs deficient mouse model (PAD4-/-), we report that NETs deposition within the tumour-draining LNs of mice upregulates the Treg population while simultaneously downregulating anti-tumour CD8 T cell proliferation and activation. Following these findings, we performed an in vitro suppression assay and found that NETs-educated Tregs hinder the expansion of CD8 T cells more than non-NETs-educated Tregs. Finally, through an induced metastasis mouse model, we have observed that tumour-draining LN resection as well as the absence of NETs protects against the development of distal metastasis, and decreases body-wide inflammation as seen by a lowered Neutrophil-to-Lymphocyte ratio in the circulation. Taken together, these findings highlight the role of NETs deposition in LNs as a key player in disease progression and bring forward a potential target for anti-metastatic drug development.

Citation Format: Ariane Brassard, Xin Su, Iqraa Dhoparee-Doomah, Sabrina Leo, Lixuan Feng, France Bourdeau, Betty Giannias, Corissa Larson, Qian Qiu, Jonathan Spicer, Lorenzo Ferri, Jonathan Cools-Lartigue. NETs act as immunosuppressive agents within lymph nodes during cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1315.
Abstract 1323: Peristalsis based modulation of colorectal cancer cells leads to changes in stemness and invasiveness in an organ on chip model
Authors: Shah Curran A. , Strelez Carly , Amzaleg Yonatan , Gunnarsson Einar B. , Foo Jasmine , Yu Min , Mumenthaler Shannon M.
Published: April 2023

Colorectal cancer (CRC) remains one of the deadliest cancers worldwide due to poor outcomes upon metastatic spread. Given the drastic impact on mortality, a better understanding of the CRC metastatic process could yield significant improvements to overall survival. Here, we investigate the influences of biomechanical forces on early metastatic spread (i.e., tumor cell intravasation events) in an organ-on-chip (OOC) model. Our microfluidic CRC OOC model consists of two channels separated by a porous membrane: a top channel representing normal and cancerous colon epithelial tissue, and a bottom channel consisting of endothelial cells in a tube-like structure representing the vasculature. Vacuum channels flank the sides of the model to apply forces in a cyclic manner to mimic peristalsis-like motions and provide further in vivo resemblance. In our prior work, we demonstrated that peristaltic forces promoted tumor cell invasion within the CRC OOC. Specifically using on-chip confocal imaging, we revealed that cyclic strain leads to an increase in the number of tumor cells capable of intravasating from the epithelial channel into the vascular channel. Additionally, we showed dynamic sensitivity of tumor cell invasion in response to intermittent stretching conditions, leading us to hypothesize mechanical stress-induced phenotypic switching. Therefore, using single-cell RNA sequencing, we investigated gene expression changes of tumor cells in the presence or absence of peristaltic forces and found alterations in stemness, adhesion and metabolism-related genes. Using median fluorescence intensity (MFI) from flow cytometry data, we have revealed differences in stem-like properties (e.g., CD44, CD133, LGR5) of tumor cells with and without the influence of peristaltic forces both prior to and after intravasation. Additionally, effluent-based analysis of the OOC has shown cytokine and metabolism changes due to peristaltic forces. Ongoing work involves using flow cytometry to further quantify changes in adhesion and metabolism as well further validation in patient derived organoids-on-a-chip. Taken together, OOCs provide a physiologically relevant model to study early CRC intravasation events, revealing phenotypic changes in response to mechanical stimuli that influence tumor cell stemness and invasiveness.

Citation Format: Curran A. Shah, Carly Strelez, Yonatan Amzaleg, Einar B. Gunnarsson, Jasmine Foo, Min Yu, Shannon M. Mumenthaler. Peristalsis based modulation of colorectal cancer cells leads to changes in stemness and invasiveness in an organ on chip model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1323.
Abstract 1345: p21 (WAF1/CIP1) drives a secretory phenotype in senescent fibroblasts and promotes bystander cancer cell growth in the tumor microenvironment
Authors: Zhang Shengliang , Huntington Kelsey E. , Zhou Lanlan , El-Deiry Wafik S.
Published: April 2023

Cancer associated fibroblasts (CAF) are major stromal cells in the tumor microenvironment (TME) and support cancer cell growth. Chemotherapy can induce CAF senescence in which p21 (the cyclin-dependent kinase inhibitor) permanently arrests the cell cycle. Cellular senescence and its associated secretory phenotype (SASP) can promote cancer progression and lead to drug resistance. We explored molecular mechanisms of p21 in the cellular communication between cancer and senescent fibroblasts. We induced fibroblast cell senescence by treatment with etoposide. Cellular senescence was confirmed by cytokine profiling, beta-gal staining and high p21 expression. To evaluate the role of p21 in the SASP of senescent fibroblasts, we transiently knocked down p21 expression in senescent fibroblast cells using siRNA. SA-beta-gal staining showed a strong beta-gal expression around nuclei in the senescent fibroblast cells with the knockdown of p21, which is the same as it was in the cells with siRNA control. Further cytokine profiling showed two panels of SASP upon p21 knockdown in senescent cells: 1) decrease of cytokines such as CCL2 and IL8 and 2) increase of cytokines such TNF-alpha correlated to the knockdown of p21 in the senescent cells. These results suggest that p21 drives a SASP in senescent fibroblast cells, in addition to its function in cell cycle arrest. We examined the effect of the p21-driven SASP in senescent fibroblasts on bystander cancer cell growth in the TME. Our co-culture system showed knockdown of p21 in senescent fibroblasts suppressed bystander cancer cell growth. We found that knockdown of TNF partially blocked the antiproliferation effect of p21-knockdown on bystander cancer cells. Our results suggest that senescent fibroblasts support bystander cancer cell growth via a p21-driven SASP in the TME and provides a new strategy for targeting p21-driven SASP in the TME for cancer therapy.

Citation Format: Shengliang Zhang, Kelsey E. Huntington, Lanlan Zhou, Wafik S. El-Deiry. p21 (WAF1/CIP1) drives a secretory phenotype in senescent fibroblasts and promotes bystander cancer cell growth in the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1345.
Abstract 1425: TTFields reduce sensitivity in glioblastoma is associated with the functional expression of the chloride intracellular channel 1 and with voltage dependent sodium channel
Authors: Castiglione Stefania , Cannavale Gaetano , Cianci Francesca , Verduci Ivan , Biella Chiara , Mercurio Chiara , Pastorelli Emanuela , Lupi Monica , Paracchini Laura , Mannarino Laura , Krex Dietmar , Brandalise Federico , Mazzanti Michele
Published: April 2023

Glioblastoma (GBM) is the most aggressive type of brain tumor, and existing treatments are inadequate at preventing recurrence. Tumor Treating Fields (TTFields) is a novel treatment that has been proven to extend patients' lives during the previous two decades. The antitumoral impact of TTFields involve different mechanism and is an optional add-on to standard temozolomide maintenance. Although TTFields therapy slows tumor progression initially, it does not prevent tumor relapse in most patients. We subjected patient-derived GBM primary cultures to continuous TTFields stimulation to mimic the GBM standard of care and examine the molecular pathways behind GBM sensitivity to TTFields treatment. TTFields initially limit cellular growth well. However, even under constant TTFields administration, the remaining cells regain their ability to proliferate at their own rate in long-term trials. Prolonged treatment with TTFields increases the expression of many stemness markers as well as the overexpression of different genes involved in cell cycle progression. There is also a rise in membrane ionic conductance activity. After sustained exposure to TTFields, sodium, potassium, and chloride channels are functionally increased. This influences the properties of tumor cells. Increased sodium channel functional activity, for example, promotes membrane potential depolarization. Furthermore, earlier research has shown that the membrane form of the chloride intracellular channel 1 (tmCLIC1) is important in the development of GBM. In patient-derived GBM cells exposed to TTFields, tmCLIC1 is significantly concentrated, and membrane depolarization improves its function. The evaluation of GBM tissues obtained from many patients who were treated with TTFields validates the in vitro tumor relapse model.Our findings imply that the process by which GBM cells develop a decrease in sensitivity to TTFields correlates with an increase in stemness markers and an increase in ionic channel functional expression.

Citation Format: Stefania Castiglione, Gaetano Cannavale, Francesca Cianci, Ivan Verduci, Chiara Biella, Chiara Mercurio, Emanuela Pastorelli, Monica Lupi, Laura Paracchini, Laura Mannarino, Dietmar Krex, Federico Brandalise, Michele Mazzanti. TTFields reduce sensitivity in glioblastoma is associated with the functional expression of the chloride intracellular channel 1 and with voltage dependent sodium channel [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1425.
Abstract 1464: Exosomal delivery of tumor suppressive microRNA-7 as a novel approach to regulating cervical cancer cell growth
Authors: McWilliams Gabriella , Roy Molly , Cortes Daniel Bilbao , Yang Yu-Ping , Daunert Sylvia , Deo Sapna
Published: April 2023

There is an unmet need for more safe, effective, targeted therapy in cervical cancer which this study seeks to meet through the use of exosomes and miRNA. Although cervical cancer is the fourth most prevalent female cancer worldwide, it currently has sparce options for treatment. Early-stage tumors are often surgically removed, while later stage cancers are treated with traditional radiation and chemotherapy methods, which have limited efficacy. Thus, to bridge this gap, we propose the use of exosomes, which are naturally occurring cell-derived vesicles, in combination with microRNA-7 (miR-7), a known tumor-suppressive microRNA, as a potential treatment for cervical cancer. We first transfected adipocyte-derived stem cells (ADSC) with miR-7. ADSC produce exosomes that are immune-compatible, have a long half-life, and have an especially low toxicity, making them a versatile nanocarrier. We collected the conditioned media produced by the ADSC, from which we isolated the miR-7-containing exosomes. The collected exosomes were confirmed to contain the appropriate miRNA by qPCR and applied to HeLa cells, and cellular response and gene expression changes were analyzed via qPCR. Functional assays including proliferation and migration were also performed to analyze the effect of exosome treatment on cell function. Controls for this study included a parallel ADSC transfection with a non-coding, scramble miRNA, as well as HeLa cells treated with exosomes from non-transfected ADSC. Our studies demonstrate that transfection of ADSC with miR-7 produces exosomes with significantly increased levels of this regulatory miRNA, and that these exosomes can specifically deliver their cargo and effectively downregulate the target gene Epidermal Growth Factor Receptor (EGFR), which is involved in tumor progression in cervical cancer. Additionally, we show that these miR-7-containing exosomes decrease proliferation and migration of HeLa cells in our in vitro functional analysis. Ultimately, this technology provides an alternative treatment option that will improve clinical outcomes of women who are affected by cervical cancer.

Citation Format: Gabriella McWilliams, Molly Roy, Daniel Bilbao Cortes, Yu-Ping Yang, Sylvia Daunert, Sapna Deo. Exosomal delivery of tumor suppressive microRNA-7 as a novel approach to regulating cervical cancer cell growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1464.
Abstract 16: The role of innate and adaptive immunity in the sex bias of LKB1-mutant lung adenocarcinoma
Authors: Fan Yijian , Monterroza Lenore , Jin Rui , Liu Xiuju , Selvaraj Periasamy , Tirouvanziam Rabindra , Zhou Wei
Published: April 2023

The tumor suppressor gene, LKB1/STK11, is one of the most frequently mutated genes in lung adenocarcinoma (LUAD). A multidimensional analysis of TCGA-LUAD patient data indicates that LKB1 mutation is one of the top sex-biased mutations, being more frequent in males than in females. A similar pattern was observed in other large LUAD cohorts. However, the underlying mechanism was unknown, partially due to a lack of an appropriate mouse model to represent this clinical phenomenon. We previously demonstrated a similar sex bias in a Lenti-Cre-induced KRAS/LKB1-mutant genetically engineered mouse model (GEMM) and a syngeneic tail-vein injection model. Here, we demonstrated that intravenous injection of the same cell line in male and female NOD scid gamma (NSG) mice does not exhibit a sex-bias pattern of lung metastasis formation (P=0.9746). However, sex bias was observed when we used CB17-SCID mice. This data suggest that innate immunity is partly responsible for the sex bias of LKB1-mutant lung cancer formation. We also characterized immune cells from the lung with LKB1-mutant tumors. Monocytes/macrophages decreased from 23% to 19%, and neutrophils increased from 15% to 19%. Lymphoid cells remained the same at 58%, and T-cells account for 34% of all CD45+ cells. Interestingly, T-cells doubled to account for 81% of all CD45+ cells from the lung in female mice resistant to the formation of lung metastasis. Our data indicated that both the innate and the adaptive immunity are responsible for eliminating LKB1-mutant lung adenocarcinoma in female mice.

Citation Format: Yijian Fan, Lenore Monterroza, Rui Jin, Xiuju Liu, Periasamy Selvaraj, Rabindra Tirouvanziam, Wei Zhou. The role of innate and adaptive immunity in the sex bias of LKB1-mutant lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 16.
Abstract 1653: Heme sequestration modulates the tumor immune microenvironment in KRAS mutation/LKB1 inactivation non-small cell lung cancer
Authors: Castro Maria del Carmen Chacon , Berisha Eranda , Salamat Narges , Zhang Li
Published: April 2023

Non-small cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancers. KRAS mutations and LKB1 inactivation commonly occur in NSCLC and are associated with poor prognosis. Several studies enunciate the role of the immune system in lung cancer progression through cancer cells' ability to escape immune surveillance. Therefore, understanding NSCLC progression requires the examination of the molecular mechanisms behind this process. The importance of oxidative phosphorylation (OXPHOS) and mitochondrial metabolism in lung cancer has been continuously emphasized; OXPHOS and mitochondrial biogenesis markers predict low survival in NSCLC patients. Additionally, heme is an essential molecule in the OXPHOS pathway, and previous studies have shown that heme sequestration using heme sequestering protein 2 (HeSP2) arrests tumor progression. HeSP2 can potentially normalize the tumor immune microenvironment (TIME) via multiple mechanisms including the normalization of vessels which may promote immune cell infiltration, the reduction of hypoxia which may promote antitumor immune function, and the reduction of heme levels which potentially decreases the production of immunosuppressive enzymes requiring heme as a cofactor. In this study, control and HeSP2-treated tumors generated from a genetically engineered mouse model LSL-KrasG12D LKB1-/- (LSL-KRASG12D; LKBloxP/loxP; LSL-Luciferase (KLLuc) infected with AdenoCre virus and sacrificed at 10 months were used for immunohistochemical analysis. We examined immune markers under HeSP2 treatment to determine the immune system response to heme targeting and its effects on tumor progression. Results demonstrated that HeSP2 can modulate the tumor immune microenvironment increasing the levels of immune markers, which suggests that heme targeting can potentiate immunotherapy efficacy in lung cancer treatment.

Citation Format: Maria del Carmen Chacon Castro, Eranda Berisha, Narges Salamat, Li Zhang. Heme sequestration modulates the tumor immune microenvironment in KRAS mutation/LKB1 inactivation non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1653.
Abstract 1676: PRMT5 regulates the chemoresistnace of ovarian cancer cells
Authors: Do Yen Thi , Kim Jin Young , Lee Seungmee , Vo Tam Thuy Lu , Pham Thi Tuyet Mai , Ha Eunyoung , Cho Chi-Heum , Shin So-Jin , Seo Ji Hae
Published: April 2023

Ovarian cancer resistance to chemotherapy accounts for the most therapeutic challenge, leading to the high mortality rate of gynecological cancer worldwide. Meanwhile, cancer stem cells (CSCs) are identified as key features in regulating ovarian chemoresistant cancer. In this study, we first found that PRMT5 was upregulated in paclitaxel (PTX)-resistant ovarian cancer cells compared with non-resistant cancer cells. Then, we determined the oncogenic role of PRMT5 in chemoresistant ovarian cancer by using its potent and selective inhibitor, GSK3326595. Disruption of PRMT5 activity by GSK3326595 or depletion of PRMT5 in PTX-resistant ovarian cancer cells restored the sensitivity to chemotherapeutic drug via triggering cell apoptosis. Then, we also validated that inhibition of PRMT5 decreased cell migration ability through the transition of motile mesenchymal phenotypes into epithelial characteristics. Moreover, PRMT5 inhibitor significantly decreased the number of sphere formation and attenuated the stem cell-like properties in PTX-resistant ovarian cancer cells. Therefore, our study suggests that PRMT5 is a powerful target to overcome resistance to conventional chemotherapy for the treatment strategy in ovarian cancer.

Citation Format: Yen Thi Do, Jin Young Kim, Seungmee Lee, Tam Thuy Lu Vo, Thi Tuyet Mai Pham, Eunyoung Ha, Chi-Heum Cho, So-Jin Shin, Ji Hae Seo. PRMT5 regulates the chemoresistnace of ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1676.
Abstract 1682: Bridging gaps in treatment resistance 2D models with the incorporation of 3D models
Authors: Gadde Manasa , Schwabe Elizabeth , Mulam Kshithija , Hill Lindsay , Bulow Christopher
Published: April 2023

Introduction: 3D cellular models have become of increasing interest in anti-cancer drug discovery as a bridge between clinical models and traditional in vitro methods. An important component of this research is the comparison of drug-to-cell interactions and morphological characteristics between cells in mono-layer culture and the cells in 3D spheroids.

Materials and Methods:The ResCu system is a novel long term, physiologically relevant culture system that avoids the need for passaging. It propagates tumoroids in a microenvironment that resembleshuman tissue. Cells with KRAS G12C mutation were cultured in the ResCu system for three months, during which they were constantly exposed to G12C inhibitors (G12Ci) for the evolution of resistant cell populations. Cells were also cultured in traditional 2D cultures and exposed to G12Ci. Transcriptomics was performed to determine resistant networks and identify different resistant subclones in each group.

Results: We applied the ResCu system to model clinical resistance. We captured 95% of clinically identified on-target and pathway bypass resistance changes, and in the 2D system, only a fraction of mechanisms were discovered. Additionally, we noticed certain cell populations could tolerate higher doses of the G12Ci after resistance evolution in ResCu compared to 2D. A few had similar responses suggesting that the origin of the cell line plays a role in the microenvironment's impact on cellular response.

Conclusion: By comparatively analyzing resistance evolution data for KRAS G12Ci in traditional 2D and ResCu, we were able to collect physiologically relevant data that can more confidently be applied to clinical studies than mono-layer cell culture data alone.

Citation Format: Manasa Gadde, Elizabeth Schwabe, Kshithija Mulam, Lindsay Hill, Christopher Bulow. Bridging gaps in treatment resistance 2D models with the incorporation of 3D models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1682.
Abstract 1736: Role of brain fatty acid binding protein in glioblastoma microtube formation
Authors: Choi Won-Shik , Mak Caitlin , Githaka John M. , Glubrecht Darryl , Jeon Pureunsol , Godbout Roseline
Published: April 2023

Glioblastoma (GBM) is a highly infiltrative brain cancer with a dismal prognosis. Conventional treatment using surgery, radiation therapy, and chemotherapy (usually temozolomide) invariably fails. GBM microtubes, which are ultra-long membrane-enclosed protrusions, are associated with therapy resistance and are believed to function by forming multicellular communication networks. Growth-associated protein GAP43 is an important protein for GBM microtube formation.

Although lipids are important for GBM progression, the roles of lipid chaperones such as the fatty acid-binding proteins (FABPs) are not well understood. FABPs are involved in fatty acid uptake from the microenvironment and intracellular transport of fatty acids. Brain fatty acid binding protein (FABP7) is highly expressed and is associated with a poor prognosis in GBM. FABP7 levels increase in therapy-resistant GBM cells, suggesting that FABP7 provides a cancer cell survival advantage when GBMs are treated with conventional therapy. FABP7 has a high affinity for polyunsaturated fatty acids (PUFAs), such as omega-3 docosahexaenoic acid (DHA) and omega-6 arachidonic acid (AA), both of which are abundant in brain. Previous studies have shown a correlation between FABP7 expression, increased neural stem cell properties, and increased cell migration in glioblastoma cells, especially when FABP7 is bound to AA. We have recently found that FABP7 is expressed in microtubes, leading to the hypothesis that FABP7 along with its fatty acid ligands can be transported to sites distant from the main tumor mass through these connecting tubes. We are studying the mechanism of action of FABP7 in microtubes by: (i) examining the effect of DHA and AA, as well as FABP7 inhibition on microtube formation and cell migration, and (ii) using both neurosphere cultures and animal models of glioblastoma to investigate the pathways that are activated by FABP7. Our results to date support the use of FABP7 inhibitors to sensitize therapy-resistant cells as part of the standard-of-care for the treatment of GBM patients.

Citation Format: Won-Shik Choi, Caitlin Mak, John M. Githaka, Darryl Glubrecht, Pureunsol Jeon, Roseline Godbout. Role of brain fatty acid binding protein in glioblastoma microtube formation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1736.
Abstract 1788: A novel costimulatory signaling domain for chimeric antigen receptor-engineering
Authors: Cho Hyun-Il , Kang Chung-Hyo , Lee Sang-Eun , Kim Mi-So , Lim Bo-Ra , Ha Jung-Min , Sohn Hyun-Jung , Kim Tai-Gyu
Published: April 2023

Chimeric antigen receptor (CAR)-engineered adoptive cell therapy has demonstrated a feasible, attractive “off-the-shelf” approach against various malignancies. Success of CAR-engineered cell therapy depends primarily on providing optimized costimulatory signals capable of achieving robust cell proliferation, persistence, and antitumor reactivity. We describe here the capability of a novel CD30-derived costimulatory signaling domain to generate effective CAR-redirected cells capable of eradicating hematological and solid cancers. After establishing CD19-redirecting second-generation CARs incorporating the CD30-derived intracellular signaling domain and third-generation CARs combining CD30 with either CD28 or 4-1BB, the effector functionality of CAR-transduced T cells was assessed in vitro and in vivo. Further, we evaluated the efficacy of epithelial cell adhesion molecule (EpCAM)-targeting third-generation CD30-encompassing CAR constructs for targeting solid tumors. The CD30-signaling CAR-T cells exhibited similar levels of cell proliferation, cytolytic efficacies, and cytokine secretion in vitro, and demonstrated improved tumor regression and prolonged survival in vivo with increased persistence of the CAR-T cells as those of other CARs with CD28 or 4-1BB costimulatory domain, which are currently in clinical use. Also, third-generation CARs including the CD30 costimulatory domain retained potent antitumor efficacies in vitro and in vivo independent of their position or type of partner costimulatory domain. More significantly, EpCAM-targeting CAR-T cells containing CD30-signaling domain revealed effective cytolytic activities correlating with EpCAM expression levels in the target cells. Our results demonstrate that the CD30-derived costimulatory domain could be an alternative for developing CAR-engineered therapeutics which may be applicable for various designs of CAR constructs, with an emphasis on effectiveness against solid tumors.

Citation Format: Hyun-Il Cho, Chung-Hyo Kang, Sang-Eun Lee, Mi-So Kim, Bo-Ra Lim, Jung-Min Ha, Hyun-Jung Sohn, Tai-Gyu Kim. A novel costimulatory signaling domain for chimeric antigen receptor-engineering [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1788.
Abstract 1823: Overcoming siglec-9-mediated immunosuppression by using glycomimetics that block the binding of this receptor with its natural ligands
Authors: AlJohani Lojain I. , Patankar Manish , Tang Weiping , Stevens Christopher , Teng Peng
Published: April 2023

MUC16, a 3-5 million Da mucin overexpressed by ovarian cancer cells binds to Siglec-9, an inhibitory receptor on immune cells. Upon binding to Siglec-9, MUC16 triggers inhibitory signaling in the immune cells that allows ovarian cancer cells to evade cytolytic immune responses. Ovarian and other tumors express a plethora of cell surface as well as secreted glycoproteins with glycans that are terminated with the Siglec-9 ligand, α-2,3-linked sialic acid. Therefore, targeting the sialic acid-Siglec-9 axis has been of major interest to augment antitumor immunity. Here, we demonstrate that synthetic glycomimetic Siglec-9 ligands have the potential to enhance the immune recognition of ovarian cancer cells. We first demonstrate, using siglec-9 chimeras, that the glycomimetics developed by our group inhibit the binding of Siglec-9 to its natural ligands. Our data showed that 50-fold excess of the glycomimetics over Siglec-9 chimera prevented the Siglec-9 chimera from binding to the OVCAR-3 cell line (P.value, 0.039). Additionally, we demonstrate that glycomimetics (#153 and 157) developed by our group, enhance the recognition of ovarian cancer cell targets by human NK cells. In healthy donor 1, CD107a expression was increased (21, 17, 19 %) when 0.01, 0.025, 0.05 mM concentration of glycomimetic 153 was used, respectively. A statistically significant increase (5%) in CD107a expression was observed when the NK cells were treated with 0.05 mM concentration of glycomimetic 157. At higher concentrations of glycomimetic 153 and 157, inhibition of CD107a was repeatedly observed. These experiments, therefore, suggest that there is likely a certain concentration range of the glycomimetic that can be used to overcome NK cell suppression occurring via Siglec-9. To our knowledge, these data are among the first to demonstrate that specially designed glycomimetics can be used to overcome the suppression of NK cells against ovarian cancer cells. Future studies will employ murine and humanized models to demonstrate the therapeutic potential of the Siglec-9-targeting glycopeptides in enhancing immune responses against epithelial ovarian cancer.

Citation Format: Lojain I. AlJohani, Manish Patankar, Weiping Tang, Christopher Stevens, Peng Teng. Overcoming siglec-9-mediated immunosuppression by using glycomimetics that block the binding of this receptor with its natural ligands [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1823.
Abstract 1946: Development and application of genetic ancestry reconstruction methods to study diversity of patient-derived models in the NCI PDXNet Consortium
Authors: Sanderson Brian J. , Lott Paul , Chiu Katherine , Quino Juanita Elizabeth , Vang April Pangia , Lloyd Michael W. , Consortium PDXNet , Srivastava Anuj , Chuang Jeffrey H. , Carvajal-Carmona Luis G.
Published: April 2023

Personalized medicine holds great promise for improving cancer outcomes, yet there is a large inequity in the demographics of patients from whom genomic data and models, including patient derived xenografts (PDX), are developed and for whom treatments are optimized. In this study we develop a genetic ancestry pipeline for the Cancer Genomics Cloud, which we use to assess the diversity of models currently available in the National Cancer Institute (NCI) supported PDX Development and Trial Centers Research Network (PDXNet). We show that there is an over-representation of models derived from patients of European ancestry, which is consistent with other cancer model resources. We discuss these findings in the context of disparities in cancer incidence and outcomes among demographic groups in the US. For example, for the top cancer health disparities affecting African Americans and Latinos, there is a significant lack of ethnic/race appropriate models needed to advance pre-clinical research and personalized clinical treatment. For stomach and liver tumors, which represent disparities in these two minority populations, there are only three available models derived from patients from such backgrounds. Fortunately, ongoing NCI-funded efforts in minority focused PDXNet centers are actively addressing these gaps. We further discuss these results in the context of power analyses to highlight the immediate need for the development of models from minority populations to address cancer health equity in personalized medicine.

Citation Format: Brian J. Sanderson, Paul Lott, Katherine Chiu, Juanita Elizabeth Quino, April Pangia Vang, Michael W. Lloyd, PDXNet Consortium, Anuj Srivastava, Jeffrey H. Chuang, Luis G. Carvajal-Carmona. Development and application of genetic ancestry reconstruction methods to study diversity of patient-derived models in the NCI PDXNet Consortium [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1946.
Abstract 2040: Molecular dynamic simulation of intermediates connecting KRAS and PRMT5 insight for targeting KRAS
Authors: Silverman Isaac V. , Stein Yitzchak F. , Gerber Michael Shawn , Maitra Radhashree , Goel Sanjay
Published: April 2023

Introduction: Therapeutic approaches to target mutant KRAS is in progress, and transcription regulator PRMT5 (Protein arginine methyltransferase) has been previously reported to be a viable, surrogate target. Mutation of oncogenic KRAS is prevalent in several cancer types often rendering aggressiveness to the disease. Herein we used extensive molecular dynamics simulations to understand the intermediate complexities of the cross-talk.

Methodology: GROMACS is a Molecular Dynamics Simulation software that renders a topology regulated by Newton’s Laws that can be employed to simulate interactions between proteins. Utilizing the STRING database, multiple pathways from KRAS linked to PRMT5 were extrapolated. Confidence values were determined. Protein complexes were downloaded from RCSB archives. Protein pairs were submitted to HDOCK for docking. Proteins were separated by chain identities, using PYMOL. Proteins were solvated and paired in GROMACS, and charges were neutralized. Each complex underwent energy minimization to configure the proteins in the most energetically favorable conformation. The complex was equilibrated and simulated. XmGrace was used to graph radius gyration between both proteins and their stabilities. The data produced were used to analyze the favorable interaction of the various pathways.

Results: Three pathways were analyzed, with high, intermediate, and low confidence values of interaction. The intermediate mediators were simulated in pairs and energy scoring was performed for each interaction. The radius of gyration (Rg) values was calculated for each protein in its respective pathway. A cutoff of 10 nanoseconds was used to confirm stability. A scale was constructed to predict the order of operations of each pathway. HRAS was determined to be actively involved in the crosstalk.

Conclusion: The crosstalk between PRMT5 and KRAS can be achieved through three different biochemical pathways with varying levels of confidence. Potentially, if one pathway is affected, an alternative route could be activated to achieve signaling toward PRMT5. These pathways introduce a more targeted approach toward discovering therapeutic advances against KRAS. More simulations are underway to compare the interactions of wildtype and mutant KRAS with their respective pathway intermediates.

Citation Format: Isaac V. Silverman, Yitzchak F. Stein, Michael Shawn Gerber, Radhashree Maitra, Sanjay Goel. Molecular dynamic simulation of intermediates connecting KRAS and PRMT5 insight for targeting KRAS [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2040.
Abstract 2204: Predicting survival outcomes in patients with acute myeloid leukemia: medium-sized versus large next-generation sequencing panels
Authors: Li Yonghong , Solis-Ruiz Jose , Racke Frederick K. , Press Richard D.
Published: April 2023

Background: Next-generation sequencing (NGS) multigene panels allow the identification of genetic variants in patients with acute myeloid leukemia (AML). This study assessed the effect of the NGS panel size on positivity rate and clinic utility in helping predict survival outcomes.

Design: This is a retrospective study of AML patients (n=76) who underwent both diagnostic and post-treatment testing with a large 220-gene NGS assay. Mutation profiles and measurable residual disease (MRD) status were assessed with the large panel or a subset of the genes on the panel (i.e., a medium-sized panel with 42 genes); MRD assessment excluded DTA mutations (i.e., mutations in preleukemic genes DNMT3A, TET2 and ASXL1). Kaplan-Meier method was used to compare overall survival between MRD positive and negative patients.

Results: The 76 patients evaluated were diagnosed with de novo AML (n=69) or secondary AML (n=7). The median age at diagnosis was 57 (interquartile range [IQR], 49-66) years; 37 (49%) were women; and the median follow-up was 273 (IQR, 174-473) days. At diagnosis, the medium-sized panel detected 276 somatic variants in 35 genes (average 3.6 variants per patient) whereas the large panel detected 351 somatic variants in 74 genes (average 4.6 variants per patient). These variants were frequently detected in methylation and epigenetic genes (25%), signaling genes (20%) and transcription factors (12%). After treatment, MRD-negativity was indicated in 27 patients by the medium-sized panel versus 23 patients by the large panel; MRD-positivity was indicated in 49 patients by the medium-sized panel versus 53 patients by the large panel. Shorter overall survival was associated with MRD-positivity as determined by both panels: medium-sized panel (hazard ratio [HR], 3.00; 95% CI, 1.01-8.93; P<.05), large panel (HR, 3.41; 95% CI, 1.00-11.57; P<.05).

Conclusion: A medium-sized NGS panel detected fewer somatic mutations than large NGS panel but both panels, when used as MRD markers, are similarly predictive of overall survival in AML patients.

Citation Format: Yonghong Li, Jose Solis-Ruiz, Frederick K. Racke, Richard D. Press. Predicting survival outcomes in patients with acute myeloid leukemia: medium-sized versus large next-generation sequencing panels [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2204.
Abstract 2235: A treatment for glioblastoma with a novel epigenetic inhibitor and identification of its target
Authors: Kota Tokunaga
Published: April 2023

Glioblastoma multiforme (GBM) is one of the deadliest human cancers. Despite extensive efforts to develop a new treatment, the prognoses of patients have not changed dramatically over the past several decades. LSD1 plays an essential role in the epigenetic regulation of gene expression in many cancer cells, including GBM, as it removes methyl groups from histone H3 lysine (K) 4me1/2 and H3K9me1/2. Since it was reported that suppression of LSD1 induced apoptosis of glioblastoma stem cells (GSCs), LSD1 has been expected as an effective therapeutic target for GBMs, particularly enriched with GSCs. We newly discovered an LSD inhibitor (S2172) that suppressed the LSD1 enzymatic activity effectively and sufficiently penetrated into brain. We found that S2172 showed a strong inhibitory effect on cell growth not only in GCS cells but also in differentiated GBM cells in vitro. In GBM cell line U251, IC50 of S2172 was lower than that of other LSD1 inhibitors (IC50 10.5 μM, 29.5 μM, 11.0 μM treated with S2172, Ory-1001, and CC-90011, respectively). We also found a robust suppression of transcription of oncogene Myc in differentiated GBM cells after S2172 treatment, which is highly expressed in most GBM cells and considered as a key element to maintain its stemness and proliferation. Consistent with the expression analysis, chromatin immunoprecipitation-sequencing (ChIP-seq) revealed that enrichment of H3K4m2 and H3K4m3 was efficiently altered by S2172 treatment along with the change of Myc expression. Finally, we treated U251 brain tumor mouse model with S2172. While S2172 had a strong effect in vitro, S2172 seemed unsuccessful in killing differentiated GBM cells in vivo. These data suggests that treatment of S2172 is less effective to differentiated GBM cells and this is consistent with previous report that LSD1 as a candidate therapeutic target in stem-like tumor propagating cells in GBM.

Citation Format: Tokunaga Kota. A treatment for glioblastoma with a novel epigenetic inhibitor and identification of its target [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2235.