bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4955: Kurarinone, a natural notch signaling inhibitor and modulator of breast cancer stemness
Authors: Kushwaha Prem Prakash , Shuaib Mohd , Prajapati Kumari Sunita , Singh Atul Kumar , Kulharia Mahesh , Kumar Shashank , Gupta Sanjay
Published: April 2023

Aberrant activation of Notch signaling pathway plays an important role in breast cancer development, progression, and stem cell renewal. Activation of Notch signaling is triggered by the interaction of Notch receptors with different ligands expressed on neighboring cells that bind and induce a second cleavage by a membrane-tethered metalloprotease. The remaining membrane-bound fragment is cleaved by γ-secretase (GS) releasing the Notch Intracellular Domain (NICD), which translocate to the nucleus to activate the expression of Notch target genes. Current indirect approaches to inhibit Notch signaling are based on agents that bind to GS substrates. In the search of novel Notch signaling inhibitor(s), we screened a number of plant flavonoids for their binding potential to GS substrate. Our in-silico study identified kurarinone (KU), a lavandulyl flavanone, having ability to bind to GS complex. The KU-GS complex was structurally and energetically stable during the molecular dynamics simulation period. Interestingly, dose and time dependent toxicity of KU was observed in T47D and MDA-MB-231 breast cancer cells at 1-25µM concentration. KU treatment of cells led to inhibition in cyclin D1 expression, arrest of cells in the G1 phase of the cell cycle and subsequently undergoes apoptosis. KU-mediated apoptosis was a result of altered mitochondria membrane potential and increased ROS production. In dual-luciferase Notch promoter assay, KU inhibited GS-mediated NICD production in transfected HEK293 cells. Furthermore, KU treatment decreased the mammosphere formation potential of MCF-7 cells through altered gene expression involved in self-renewal and stemness. KU treated MDA-MB-231 cells and MCF-7 derived mammosphere showed downregulation of Notch signaling through alteration in the expression of downstream genes such as Hey1, Hes1, and E-cadherin at both transcript and protein levels. Taken together, the present study identified KU as a novel natural Notch signaling inhibitor that could provide a novel and safer approach for the treatment of breast cancer through reduction in stemness and self-renewal properties.

Citation Format: Prem Prakash Kushwaha, Mohd Shuaib, Kumari Sunita Prajapati, Atul Kumar Singh, Mahesh Kulharia, Shashank Kumar, Sanjay Gupta. Kurarinone, a natural notch signaling inhibitor and modulator of breast cancer stemness. [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 4955.
Abstract 4960: Fascin inhibitor decreases gynecological cancer cell growth through cell cycle regulation
Authors: Chen Wanyi , Wang Yufeng , Ji Liangliang , Li Ming O. , Huang Xin-Yun
Published: April 2023

The actin cytoskeleton is essential for maintaining cell morphology and architecture. Actin-bundling proteins such as fascin cross-link actin filaments into bundles and play critical roles in regulating cell protrusion and motility. Fascin protein expression is low or absent in normal human epithelial cells but high in cancer cells. Elevated fascin levels are correlated with aggressive clinical progression, poor prognosis, and shorter survival outcomes. It is regarded as a cancer progression biomarker and a therapeutic target. We have developed a small molecule fascin inhibitor and shown its efficacy in blocking tumor cell migration, invasion, and metastasis, as well as prolonging the overall survival of mice bearing different types of cancers. Our recent data reveals a new mechanism of this fascin inhibitor in cell cycle regulation of gynecological cancer cells. It blocks the G2/M progression and decreases the mitotic index. Fascin inhibitor treatment also results in aneuploidy, which is a well-known factor for triggering downstream apoptosis. Our data suggests that fascin is involved in maintaining the fidelity of chromosome segregation and cell division. This will advance our understanding of the interplay between actin and microtubule cytoskeleton during cell mitosis.

Citation Format: Wanyi Chen, Yufeng Wang, Liangliang Ji, Ming O. Li, Xin-Yun Huang. Fascin inhibitor decreases gynecological cancer cell growth through cell cycle regulation. [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 4960.
Abstract 5058: Impact of dexamethasone on chemotherapy response in ovarian cancer
Authors: Venkatachalam Annapoorna , Correia Cristina , Mcgehee Cordelia D. , Finstuen-Magro Sarah , VanBlaricom Jamison , Schneider Paula A. , Peterson Kevin L. , Heinzen Ethan P. , Larson Melissa C. , Hou Xiaonon , Oberg Ann L. , Weroha Saravut John , Kaufmann Scott H.
Published: April 2023

Background: Ovarian cancer ranks fifth in cancer deaths among women. The gold standard of care for advanced disease includes cytoreductive surgery along with adjuvant or neoadjuvant platinum/taxane-based chemotherapy. Although response to this treatment is high, chemotherapy-induced nausea and vomiting due to platinum agents and allergic reactions due to both agents are common. For nearly half a century, the glucocorticoid (GC) dexamethasone (DEX) has been administered as part of supportive care. However, there have periodically been questions about the possible deleterious effects of DEX on chemotherapy response, especially in solid tumors.

Methods: Platinum-sensitive, glucocorticoid receptor (GRα) expressing ovarian cancer cells were exposed to DEX prior to chemotherapy and clonogenic survival was assessed. The effects of DEX on cell cycle and apoptosis were assessed by flow cytometry. Changes in the tumor cell transcriptome after acute DEX treatment were determined by RNA sequencing. Mice bearing orthotopic ovarian cancer patient-derived xenografts (PDXs) were treated with chemotherapy ± DEX to assess changes in chemotherapy response in vivo.

Findings: DEX pre-treatment decreased platinum cytotoxicity in ovarian cancer cell lines in vitro and diminished platinum response in some PDXs analyzed for chemotherapy response ex vivo. RNA-seq analysis revealed changes in transcripts encoding proteins involved in platinum detoxification and TNFα pro-survival signaling. In ovarian cancer PDXs, DEX decreased platinum response in one model and had limited impact on both platinum and platinum/taxane chemotherapy in three distinct studies.

Interpretation: DEX appears to modulate chemotherapy response in some settings in vitro and ex-vivo. The impact of DEX on chemotherapy response in vivo appears to be limited. However, further investigation of this question in vivo in non-immunosuppressed murine models appears warranted.

Citation Format: Annapoorna Venkatachalam, Cristina Correia, Cordelia D. Mcgehee, Sarah Finstuen-Magro, Jamison VanBlaricom, Paula A. Schneider, Kevin L. Peterson, Ethan P. Heinzen, Melissa C. Larson, Xiaonon Hou, Ann L. Oberg, Saravut John Weroha, Scott H. Kaufmann. Impact of dexamethasone on chemotherapy response in ovarian 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 5058.
Abstract 5112: Immune biomarkers of clinical response after NSAID treatment in combination with vaccination targeting EGFR-COX2-CDC25B in the APC(Min/+) mouse model
Authors: Cecil Denise L. , Liu Ying , Corulli Lauren , Disis Mary L.
Published: April 2023

Studies from our group have shown that administration of NSAIDs in mouse models of Familial Adenomatous Polyposis (FAP) results in a decrease in the development in small intestinal polyps with a significant increase in CD8 tumor infiltrating lymphocytes (TIL) and downregulation of PD-L1 on tumors. Furthermore, we have demonstrated that NSAIDs could be used in combination with vaccination to even further reduce the development of intestinal polyps. We questioned what tissue specific biomarkers could define the efficacy of response with the combination treatment. APC(Min/+) male and female mice (4-6 weeks old) were immunized with a multi-antigen (EGFR, COX2, CDC25B) peptide vaccine with CFA/IFA as an adjuvant. Control mice received immune adjuvant alone. Regular chow or chow containing 400 ppm naproxen was provided intermittently (3 weeks on, 3 weeks off for 3 cycles) or continuously (18 weeks on) at the initiation of the vaccine regimen. Animals were euthanized at 22 weeks and tumors quantified. TIL were evaluated by flow cytometry. Since intermittent naproxen treatment was just as effective at inhibiting tumor development as continuous naproxen treatment, with or without vaccine, we did not differentiate between NSAID regimens. We observed an increase in CD8+ (83%; p<0.0001), Tbet+CD4+ (94%; p<0.001), effector memory (EM) CD4+ (76%; p<0.0001) and EM CD8+ (94%; p<0.0001) TIL in the tumors that did develop in the mice treated with the combination NSAID and vaccination as compared to control. There was increased CD8+ (Pearson correlation coefficient: −0.506, p<0001), Tbet+CD4+ (Pearson correlation coefficient: -0.6222, p<0.001), EM CD4+ (Pearson correlation coefficient: 0.-6074, p<0.001) and EM CD8+ (Pearson correlation coefficient: -0.7155, p<0.001) in mice with greater tumor inhibition than those with less protection. Receiver operator curves revealed that CD8+ (AUC: 0.7623; p=0.002), Tbet+CD4+ (AUC 0.7273; p=0.008), EM CD4+ (AUC 0.6782; p=0.04) and EM CD8 (AUC 0.7897; p<0.001) TIL can differentiate mice receiving the combination of NSAID and vaccination as compared to either monotherapy alone. These data define potential biomarkers of response that can guide human clinical trials of NSAIDs +/- vaccination in patients with FAP.

Citation Format: Denise L. Cecil, Ying Liu, Lauren Corulli, Mary L. Disis. Immune biomarkers of clinical response after NSAID treatment in combination with vaccination targeting EGFR-COX2-CDC25B in the APC(Min/+) mouse 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 5112.
Abstract 5135: Standardized off-the-shelf engineered NK cell therapy with improved ADCC properties to treat malignancies
Authors: Lin Yanan , Zhang Shengbang , Ran Jie , Song Can , Zhu Shengcang , Shi Hui , Guo Ping , Tan Shengjiang , Gu Yuchun , Wu Lida
Published: April 2023

In the treatment of solid tumors, chimeric antigen receptor(CAR)-engineered NK (CAR-NK) cells have distinct advantages over CAR-T cells, such as a lack graft-versus-host disease in allogenic setting to make “off the shelf” medicine; much safer because they are less likely to cause cytokine storms; and NK cells have their own activated receptors that recognize tumor surface antigens and thus have a natural ability to kill a wide range of tumors. In addition, NK cells are the main performers of antibody-dependent cell-mediated cytotoxicity (ADCC), which binds to the Fc-terminus of antibodies through their surface Fc receptor CD16A, thereby killing antibody-targeted tumor cells. Enhancing the ADCC action of NK cells themselves enhances the therapeutic efficacy of monoclonal antibodies targeting tumor surface antigens and is therefore of great importance in the treatment of tumors, especially solid tumors. We screened two optimal structures among nine different Fc chimeric receptors, and NK cells overexpressing these two receptors killed K562 and Daudi six times more intensely than normal NK cells when combined with rituximab, while the killing ability was comparable to that of normal NK without rituximab, indicating that NK cells expressing chimeric Fc receptors have stronger ADCC effects. Moreover, these NK cells can kill target cells in multiple rounds. In vivo experiments in mice demonstrated that NK cells expressing chimeric Fc receptors in combination with EGFR monoclonal antibodies had a stronger inhibitory effect on tumor growth than monoclonal antibodies or NK cells alone. Here we provide a novel broad “off the shelf” NK cells that can significantly enhance the killing ability of hematological and solid tumors in combination with different monoclonal antibodies.

Citation Format: Yanan Lin, Shengbang Zhang, Jie Ran, Can Song, Shengcang Zhu, Hui Shi, Ping Guo, Shengjiang Tan, Yuchun Gu, Lida Wu. Standardized off-the-shelf engineered NK cell therapy with improved ADCC properties to treat malignancies. [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 5135.
Abstract 5138: Systemic agonist anti-CD40 treatment delivers precise melanoma immunotherapy within established autoimmune landscapes
Authors: Valencia Julio C. , Sanford Michael , Young Howard A.
Published: April 2023

The overlapping signatures of type 1 interferons (IFN) and IFN-gamma (IFNg) has proven challenging in human autoimmunity and cancer immunotherapy, particularly for resistance to anti-programmed death 1 (anti-PD1) monotherapy. By contrast, agonist abs targeting of CD40 (anti-CD40) antibodies (abs) has clinically elicit anti-tumor immunity with limited autoimmune complications. Here, we explore the relationships between anti-PD1 or agonist CD40 responses in melanoma tumors from type 1 IFN alpha receptor (IFNAR)-sufficient and type 1 IFNAR-deficient lupus-prone mice that overexpress IFNg. To address this, RNA-sequencing (RNA-seq) data on tumors treated with either anti-PD1 or agonist anti-CD40 was evaluated for tumor objective responses compared to untreated controls. Cellular and canonical pathway identification from RNA-seq data were interrogated from the IPA library for significantly differential expressed genes (DEG). Variance patterns of the anti-PD1 and anti-CD40 monotherapies identified known resistance and response pathways highly concordant with IFN co-expression patterns in the RNA-sequencing data, respectively. IPA biomarker analysis for uniquely regulated genes showed that anti-PD1 induced only 59 DEG; while anti-CD40 induced 494 DEG including Il-27, and PR/SET Domain 1 (PRDM1), a transcription factor regulated by IL-27. Specifically, signatures of MDSC development (irf8, il10); and adaptive resistance (pdcd1, pdcd1lg2, and TNFRSF5/CD40)) were negatively associated with response to anti-P1 monotherapy. By contrast, signatures of IFN activation (stat1, Ifng, and Ifngr1), chemotaxis (Cxcl10, Il12b and Il-27), attractants (Sema3g and Sema4a), and antigen presentation MHC II (h2-Aa, h2-Ab1, and ciita) were associated positively with response to agonist anti-CD40 monotherapy. Importantly, anti-CD40 alone delivered anti-tumor immune responses in type 1 IFN receptor-deficient lupus-prone mice that overexpress IFNg (Ifnar-/-ARE-/-) mice suggesting synergy with therapies blocking type 1 IFN signaling. Thus, these findings indicate that the rational use of CD40 agonists abs provide a better therapeutic platform to deliver precise anti-melanoma responses in complex autoimmune landscapes.

Citation Format: Julio C. Valencia, Michael Sanford, Howard A. Young. Systemic agonist anti-CD40 treatment delivers precise melanoma immunotherapy within established autoimmune landscapes. [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 5138.
Abstract 5148: Interrogating immuno-oncological interactions in the tumor microenvironment
Authors: Patterson Ben , Chen g-Yi , Fernandez Nicolas , Colbert Leiam , He Jiang
Published: April 2023

Single-cell RNA sequencing is a powerful and innovative tool for assessing individual cancer prognoses and outcomes; however, the process of generating this data typically destroys the spatial complexity of the tumor microenvironment. Understanding the spatial complexities within cancer provides crucial information for evaluating individual tumor development and progression. Vizgen’s MERSCOPETM Platform, built on Multiplexed Error-Robust Fluorescence in situ Hybridization (MERFISH) technology, enables the direct profiling of the spatial organization of intact tissue with subcellular resolution. Here, we present a Pan-Cancer approach to demonstrate the MERSCOPE’s ability to characterize breast, colon, prostate, ovarian, lung, and skin cancers from human clinical samples. Specifically, using a 500-gene panel to assess the canonical signaling pathways of cancer, cancer type-specific genes, select immune genes, proto-oncogenes, and tumor-suppressor genes, we show that the MERSCOPE was able to spatially profile gene expression across multiple tumor types. We utilized the Vizgen Cell Boundary Stain kit to generate a molecular and cellular atlas of individual patient tumors by clustering cells based on gene expression and mapping their spatial organization. To gather insight into immuno-oncological interactions and their effect on gene expression across these tumor types, we performed a neighborhood analysis to determine how proximity to immune cells altered gene expression in tumor cells, and vice versa. We found significant gene expression changes in MHC activation and downstream pathways in immune cells located nearest to tumor cells; in tumor cells nearest to immune cells, we identified increased expression of genes related to immune response mediation and inflammation. Furthermore, we compared the annotated immune cell populations across the various cancer types to observe differences in gene expression. Here, we used publicly available single-cell RNA-sequencing data to impute gene expression for these immune cell populations. This analysis demonstrated the transcriptional variation possible in identical immune cell types that are acting in different tissue contexts. Notably, we were able to map and catalogue different patterns of gene expression relating to inflammation, immune signaling, and immune cell exhaustion. These findings demonstrate how the MERSCOPE platform can provide deep insights into the complex heterogeneity observed between different tumor types and tissues.

Citation Format: Ben Patterson, g-Yi Chen, Nicolas Fernandez, Leiam Colbert, Jiang He. Interrogating immuno-oncological interactions 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 5148.
Abstract 5190: Systematic immune-profiling of immune-deficient mouse models: A rational to select ideal host for tumor implantation
Authors: Luche Hervé , Hadjem Lillia , Mello Marielle , Canavese Priscilla , Angelis Fabien , Joachim Anais , Bouilly Sylvie , Guinut Frederic , Fiore Frederic , Malissen Bernard , Zarubica Ana , Corcuff Erwan
Published: April 2023

Preclinical tumor oncology research relies historically on the analysis of mouse or human tumor cell lines implanted onto severely immune-deficient mouse models carrying nude, SCID or Rag mutations on different genetical background (C57Bl/6N, BalbC, NMRI, CB17 or NOD). These lines harbor defects in several leukocyte lineages among which Tc, Bc and NKc might be affected. Humanized mice are emerging models that have been transplanted with human cells or tissues (and/or equipped with human transgenes). Currently, the most advanced strains are the nonobese diabetic, severe combined immunodeficiency (NOD-SCID) mouse with complete disruptions in the interleukin-2 (IL-2) common γ-chain (IL2R γ null) receptor (NSG) and BALB/c Rag2−/− IL2R γ null SirpaNOD mice (BRGS).1 Juvenile chimera NSG or BRGS are reconstituted with hematopoietic progenitor cell (HPCs) from human cord blood yielding robust engraftment of a human immune system (HIS).. Recent work showing the binding of human IgG4 to mouse FcgR receptor reducing clinical efficacy of therapeutic antibodies highlights the need to characterize extensively the residual immune system present in these models prior humanization in order to precisely evaluate immunotherapies. In other to fulfill this task, we genetically modified the mouse genome introducing mutations involved in Tc, Bc and NKc development. We then immuno-phenotyped over 20 lines of mouse mutants produced from the same animal house to minimize the impact of microbiota on innate immune cell populations. High content cytometry analysis of BM, spleen, thymus and peripheral blood was performed in a standardized manner. The resource obtained of these immunotypes was used to highlight similarities and differences among these lines across several genetical background. This work has shown instrumental in selecting the most appropriate model to use for humanization followed before tumor implantation.

Citation Format: Hervé Luche, Lillia Hadjem, Marielle Mello, Priscilla Canavese, Fabien Angelis, Anais Joachim, Sylvie Bouilly, Frederic Guinut, Frederic Fiore, Bernard Malissen, Ana Zarubica, Erwan Corcuff. Systematic immune-profiling of immune-deficient mouse models: A rational to select ideal host for tumor implantation. [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 5190.
Abstract 5326: Comparison of CAR-T cell-mediated cytotoxicity assays with suspension tumor cells using high-throughput plate-based image cytometry method
Authors: Chan Leo Li-Ying , Sun Yu-Jun , Chen Yi-Chun , Hua Wei-Kai , Wu Sariena Chiung-Yuan
Published: April 2023

In the recent decade, chimeric antigen receptor (CAR)-T cell therapy has revolutionized strategies for cancer treatments due to its highly effective clinical efficacy and response for B cell malignancies. Currently, there are six CAR-T cell products available in the market including Kymriah, Yescarta, Tecartus, Breyanzi, Abecma, and Carvykt. The success of CAR-T cell therapy has stimulated the increase in the research and development of various CAR constructs to target different tumor types. Therefore, a robust and efficient in vitro potency assay is needed to quickly identify potential CAR gene design from a library of construct candidates. Traditionally, in vitro CAR-T cell-mediated cytotoxicity is assessed using release assays such as 51Cr (radioactivity), calcein (fluorescence), and LDH (enzymatic). However, release assays indirectly measure cell death via molecules released in supernatant and typically limited to only endpoint assays. In addition, handling and disposing of 51Cr hazardous materials is less preferred. Luciferase reporter assay, although highly sensitive, has similar drawbacks as the release assays. Finally, flow cytometry method can directly measure cell death and viability, but can be time-consuming and requires a large number of CAR-T cells when the effector-to-target (E:T) ratio is high. Furthermore, additional steps are required for adherent cells that require trypsinization. Image cytometry methodologies have been utilized for various CAR-T cell-mediated cytotoxicity assay using different fluorescent labeling methods, mainly due to their ease-of-use, ability to capture cell images for verification, and higher throughput performance. In this work, we employed the Celigo high-throughput plate-based Image Cytometer to evaluate and compare two CAR-T cell-mediated cytotoxicity assays using GFP-expressing or fluorescent dye-labeled myeloma and plasmacytoma cells. Performing time- and E:T ratio-dependent CAR-T cell-mediated cytotoxicity assays, the GFP-based method demonstrated higher sensitivity in detecting the level of cytotoxicity when compared to the CMFDA/DAPI viability method. We have established the criteria and considerations for the selection of cytotoxicity assays that are fit-for-purpose to ensure the results produced are meaningful for the specific testing conditions.

Citation Format: Leo Li-Ying Chan, Yu-Jun Sun, Yi-Chun Chen, Wei-Kai Hua, Sariena Chiung-Yuan Wu. Comparison of CAR-T cell-mediated cytotoxicity assays with suspension tumor cells using high-throughput plate-based image cytometry method. [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 5326.
Abstract 5536: Landscape and analysis of ERBB2 amplification and short variant mutations in large-scale Chinese patients with colorectal cancer
Authors: Chen Zhiqin , Yu Shijun , Wu Yin , Zhang Bei , Zhang Ding , Gao Yong , Quan Ming
Published: April 2023

Background: Human epidermal growth factor receptor 2 (ERBB2) amplification is a rapidly emerging therapeutic target in CRC, in spite of a relatively low incidence. While ERBB2 amplification has been considered as a critical biomarker to predict the efficacy and safety of anti-ERBB2 therapy of CRC, its point mutation status is largely ambiguous.Methods: Blood or tissues samples from 2454 Chinese patients with CRC were collected, and the genomic landscape, clinical characteristics and genetic features of ERBB2 in these samples were retrospectively investigated using next-generation sequencing (NGS) analysis. Results: Of the 2454 colorectal cancer patients, 3.46% (85) carried ERBB2 amplification, 2.24% (55) carried ERBB2 mutation, and some patients harbored both ERBB2 amplification and point mutation at the same time. Microsatellite instability high (MSI-H) was found in approximately 32.7% (18/55) of patients with ERBB2-mutated CRC, but almost negative in ERBB2-amplified CRC. The median fold change of ERBB2 amplification was 30 (Range 4 to 245). Among the 16 mutation sites detected from 55 patients with ERBB2 point mutations, p.R678Q was the most common mutation site (28%), followed by p.V842I (24%) and p.S310F/Y (12%), which appeared to be slightly different from previous reports. ERBB2 mutation was more frequently found in CRC patients aged < 50 years. 50.9% of CRC patients with wild-type ERBB2 carried KRAS mutation, which was significantly higher than those with mutated ERBB2 (25.6%). A higher incidence of BRAF mutation was observed in patients with mutated-ERBB2 (8.5% vs. 2.3%). In addition, CRC patients with ERBB2 CNV showed a higher prevalence of TP53 mutation comparing with those with ERBB2 SNV (92.3% vs. 58.3%), while KRAS mutation exhibited a contrary trend (14.1% vs. 45.8%)Conclusions: The overall variation rate of ERBB2 is 5.4% in Chinese patients with CRC, which is accompanied by significantly different molecular pathological characteristics compared to patients with wild-type ERBB2, and different mutation types (CNV or SNV) display different molecular pathological characteristics.

Citation Format: Zhiqin Chen, Shijun Yu, Yin Wu, Bei Zhang, Ding Zhang, Yong Gao, Ming Quan. Landscape and analysis of ERBB2 amplification and short variant mutations in large-scale Chinese patients with colorectal 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 5536.
Abstract 554: The role of Mycobacterium w as a novel treatment for osteosarcoma
Authors: Stefan Kirsten , Dheeraj Arpit , Tailor Dhanir , Kumar Sushil , Li Wenqi , Modi Rajiv , Joshipura Manjul , Khamar Bakulesh , Davis Lara E. , Malhotra Sanjay V.
Published: April 2023

Osteosarcoma (OS) is the most common malignant primary tumor of bone and occurs in adolescents and young adults as well as in the elderly. Despite a number of large clinical trial efforts, metastatic sarcoma remains a lethal disease, and there have been no meaningful advances in therapy or improvements in patient outcomes for decades. Mycobacterium w (Mw, also known as Mycobacterium indicus pranii) is a non-pathogenic strain of mycobacterium which has been used as an immunomodulator against leprosy, tuberculosis and several cancers and is approved for treatment of NSCLC along with chemotherapy in India. Mw promotes a Th1 immune response and its administration in tumor bearing animals is associated with an increase in total number of tumor infiltrating immune cells (TIIC) with decrease in FOXP3 and PD-1 expressing TIIC in pancreatic cancer and melanoma. Previous studies in our lab have shown that Mw promotes mature dendritic cell and activated CD8+ T cell accumulation within the tumor microenvironment in a mouse model of pancreatic cancer. We compared the effects Mw with standard of care doxorubicin on OS tumor progression in a subcutaneous K7M2 syngeneic mouse model. Mw treatment was more effective at tumor inhibition than doxorubicin. Immunohistochemistry analyses of OS tumors revealed an increase in macrophages following Mw treatment. We aim to further analyze the tumor immune microenvironment using a panel of 28 antibodies to determine alterations due to Mw treatment. With these results we hope to show that Mw is a promising candidate for further development as a therapy for OS.

Citation Format: Kirsten Stefan, Arpit Dheeraj, Dhanir Tailor, Sushil Kumar, Wenqi Li, Rajiv Modi, Manjul Joshipura, Bakulesh Khamar, Lara E. Davis, Sanjay V. Malhotra. The role of Mycobacterium w as a novel treatment for osteosarcoma [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 554.
Abstract 5571: FoxP3 absolute quantification to build an effective risk model for cancer patient survival
Authors: Ponzoni Adele , Delebecq Sandra , Boute Melodie , Ramos Corinne , Stauber Jonathan
Published: April 2023

Background: Forkhead box protein P3 (FOXP3), belonging to the forkhead/winged-helix family, is of great importance in modulating the differentiation and development of T-regulatory cells (Tregs). The numbers of FOXP3+ Tregs as well as exhaustive subsets of CD4+ and CD8+ T cells have been observed increased in solid tumor tissues of patients with colorectal and breast cancers, which helps to create an immunosuppressive environment and promotes tumor progression. However, the correlation between the presence of FOXP3+ Tregs and patients’ survival has been extensively studied in many cancer types, which remains conflicting. The approach presented here aims to absolutely quantify the level of FOXP3 in single cells and constructed effective immune risk models to predict tumor prognosis.

Method: Imaging mass cytometry (IMC) was used to quantify FOXP3 inside each single cell. The indirect quantification used a free-metal titration curve in order to match the level of intensity of the metal-tagged antibodies to the level of expression of the antigen. Deep learning (StarDist QuPath extension) was used for cell segmentation and expression quantification. FOXP3 quantification in tissue or at the single cell level was validated against ELISA assay that measure FOXP3 in tissue lysate.

Results: Linearity of the metal signals with an average %CV of 17% was observed emphasizing the linear correlation between free metal concentration and intensity. With the IMC technique, the minimum amount of FoxP3 detectable is 0.09 femtogram and its absolute quantification in the different cells in tumor tissues was confronted to disease prognosis in order to start building an effective immune risk model.

Conclusion: With the introduction of new biologics in cancer therapy, it is imperative to follow real quantitative change in the expression of biological marker that are indicator of tumor prognosis. Indeed, although the FOXP3 distribution was already seen to be heterogenous in tumors, its absolute level of expression indicative of disease progression can be directly correlated to gain better understanding of the patient response stratification to biologics.

Citation Format: Adele Ponzoni, Sandra Delebecq, Melodie Boute, Corinne Ramos, Jonathan Stauber. FoxP3 absolute quantification to build an effective risk model for cancer patient survival. [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 5571.
Abstract 5666: NAV-003, a full-length IgG1 bispecific antibody targeting a unique mesothelin epitope and CD3 to improve cytotoxicity against humoral immunosuppressed tumors
Authors: Grasso Luigi , Jiang Qun , Hassan Raffit , Nicolaides Nicholas C. , Kline J. Bradford
Published: April 2023

Mesothelin (MSLN) is a cell surface protein over-expressed in a number of cancer types. Several antibody- and cellular-based MSLN targeting agents have been tested in clinical trials where their therapeutic efficacy has been moderate at best. Previous studies using antibody and CAR-T based agents have shown the importance of particular MSLN epitopes for optimal therapeutic response, while other studies have found certain MSLN-positive tumors can produce proteins that can bind to subsets of IgG1-type antibodies and suppress their immune-effector activities. In an attempt to develop a more optimized anti-MSLN targeting agent, we engineered a humanized divalent anti-MSLN/anti-CD3ε bispecific antibody called NAV-003 that targets a MSLN epitope proximal to the tumor cell surface and an anti-CD3ε single-chain antibody capable of effectively binding, activating and directing T-cells to the surface of MSLN-bound tumor cells. NAV-003 has shown significantly improved tumor cell killing against lines producing immunosuppressive proteins in vitro and in vivo. Moreover, NAV-003 demonstrated good tolerability and efficacy against patient-derived mesothelioma xenograft models co-engrafted with human peripheral blood mononuclear cells. Together these data support the potential for NAV-003 clinical development and human proof-of-concept studies in patients with MSLN-expressing cancers.

Citation Format: Luigi Grasso, Qun Jiang, Raffit Hassan, Nicholas C. Nicolaides, J. Bradford Kline. NAV-003, a full-length IgG1 bispecific antibody targeting a unique mesothelin epitope and CD3 to improve cytotoxicity against humoral immunosuppressed tumors. [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 5666.
Abstract 5818: Cadherin complexes recruit PIWIL2 to suppress pro-tumorigenic transformation
Authors: Risner Alyssa , Nair-Menon Joyce , Gangaraju Vamsi , Kourtidis Antonis
Published: April 2023

Recent studies have shown that genomic instability, oncogene expression, and high mutation rates in more than 50% of tumors can be attributed to increased transposon activity. Transposons, also called transposable elements (TEs), are parasitic mobile DNA elements that make up 45% of the human genome and can cause DNA damage and gene mutations. The reasons for increased TE activity in somatic tumors are currently unknown. TE RNA transcripts are targeted for degradation by PIWI ribonucleases, which are members of the Argonaute superfamily of proteins, through regulation of small RNAs called piRNAs (PIWI-interacting RNAs). Although the function of the PIWI-piRNA pathway has been extensively studied in the germline, there is little understanding about the presence or functionality of the pathway in somatic tissues. We have evidence of a novel interaction of the adherens junctions of well-differentiated epithelial cells with PIWIL2 (PIWI-like RNA-Mediated Gene Silencing 2), a key catalytic component of the PIWI-piRNA complex. Through immunofluorescence staining, confocal microscopy, and co-immunoprecipitation studies, we found co-localization and association of PIWIL2, as well as of other key members of the PIWI complex, such as PIWIL4 and TDRD1, with E-cadherin and p120 catenin at adherens junctions of well-differentiated breast and colon epithelial cells, whereas this association is lost in cancer cells and patient tissues. Adherens junction disruption results in mis-localization of PIWIL2, PIWIL4, and TDRD1 from the adherens junctions and their accumulation either to the cytoplasm or to the nucleus. E-cadherin depletion particularly results in increased expression of the LINE1 transposon, which is the most abundant transposon in human cells, and in increased levels of γ-H2AX, an indicator of DNA damage. Our working hypothesis is that cadherin junctions of well-differentiated epithelial cells recruit and regulate the PIWIL2 complex to suppress transposon activity. Since both loss of junctional integrity and increased transposon activity are universal events in cancer, this study has the potential to deepen our understanding of the role of the PIWI-transposon pathway in somatic tumorigenesis.

Citation Format: Alyssa Risner, Joyce Nair-Menon, Vamsi Gangaraju, Antonis Kourtidis. Cadherin complexes recruit PIWIL2 to suppress pro-tumorigenic transformation [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 5818.
Abstract 5819: Src inhibitor suppresses rab11 mediated collective cell migration in colon cancer cells
Authors: Lu Yi-Wen , Hou Xiang-Ling , Koo Hui Min , Chao Wei-Ting
Published: April 2023

Collective cell migration has been demonstrated the importance in cancer metastasis. Instead of single cell migration, collective cell migration requires E-cadherin for the cohort cell migration. Rab11 the recycling vesicle protein plays the key role in E-cadherin dynamics and mediates collective cell migration. In this study, Src was hypothesized to regulate Rab11 mediated collective cell migration. Our results demonstrated Src inhibitor Dasatinib has less toxicity in HT-29 colon cancer cells, instead, the cell morphology was turned aggregated. The immunofluorescence image also showed E-cadherin polarity was lost after Dasatinib treatment and increased the potential of cell-cell contact inhibition. In transwell assay, Rab11 was found colocalized with E-cadherin in collective migrated cells. Dasatinib treatment can suppress collective cell migration and decrease the interaction of Rab11 and E-cadherin. This study demonstrated the important role of Src in Rab11 mediated collective cell migration through E-cadherin dynamics in colon cancer.

Citation Format: Yi-Wen Lu, Xiang-Ling Hou, Hui Min Koo, Wei-Ting Chao. Src inhibitor suppresses rab11 mediated collective cell migration in colon 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 5819.
Abstract 5902: The immune features of Helicobactor pylori-positive gastric cancer provides insights into immunotherapy
Authors: Jia Keren , Chen Yang , Chong Xiaoyi , Li Yilin , Yuan Jiajia , Li Yanyan , Zhang Xiaotian , Li Jian , Shen Lin
Published: April 2023

Introduction: Helicobactoer pylori (H. pylori) is one of the most important risk factors of gastric cancer (GC). However, the effects of Helicobacter pylori on the immune microenvironment and its potential roles in tumor immunotherapy responses is still lacking.

Methods: We included 61 patients, 46 of whom received anti-PD-1/PD-L1 based immunotherapy. Patients’ samples at baseline were collected and examined with multiplex immunothistochemistry. Four detecting panels including 16 kinds of antibodies were used to label CD8+ T cells, CD4+ T cells, B cells, macrophages, neutrophils, and their functional status. Immunotherapy-related overall survival (irOS) is the interval from the initiation of immunotherapy to death or the last following up. mmunotherapy-related progression-free survival (irPFS) is the interval from the initiation of immunotherapy to disease progression or the last following up. The Student’s t test is used to compared the average density of immune cells. The Chi-squared test was used to compare the objective response rate (ORR) between two groups.

Results: H. pylori-positive rate was 29/61. In 46 patients received immunotherapy, 23 patients were infected with H. pylori. There was a increasing trend for the density of CD8+ T cells, B cells, and neutrophils in H. pylori-positive patients compared with negative group.

We observed that the densities of total CD4+ T cells, CD4+PD-L1+, and CD4+CTLA-4+ T cells, were higher in H. pylori-positive group compared with negative group (522.58 vs. 268.77/mm^2, P=0.020, 118.94 vs. 265.98/mm^2, P=0.0091; 61.30 vs. 18.56/mm^2, P=0.035). In addition, STING positive macrophages (CD68+STING+ cells) were also enriched in H. pylori-positive group than negative group (138.45 vs. 44.15/mm^2, P=0.0012).

Next, we found H. pylori-positive patients harbored higher ORR than negative group (47.62% vs. 17.39%, P=0.032). However, there were no significant difference of irPFS and irOS between positive and negative group (median irPFS: 5.97 vs. 6.83 months, hazard ratio: 1.02, P=0.96; median irOS: 12.83 vs. 10.97 months, hazard ratio: 0.855, P=0.59).

Conclusions: We observed the enrichment of total CD4+ T cells, CD4+PD-L1+ T cells, CD4+CTLA-4+ T cells, and CD68+STING+ cells in H. pylori-positive patients, indicating the combination of anti-CTLA-4 or STING may enhance the efficacy of immunotherapy in H. pylori-positive GC.

Citation Format: Keren Jia, Yang Chen, Xiaoyi Chong, Yilin Li, Jiajia Yuan, Yanyan Li, Xiaotian Zhang, Jian Li, Lin Shen. The immune features of Helicobactor pylori-positive gastric cancer provides insights into immunotherapy. [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 5902.
Abstract 5912: Bitter taste receptors link between microbiome dysbiosis and programmed cell death in oral squamous cell carcinoma
Authors: Hwang Byeong-Oh , Park Se-Young , Song Na-Young
Published: April 2023

Oral squamous cell carcinoma (OSCC) is the most common type of head and neck cancers. It has been reported that the composition of the oral microbiome was shifted in OSCC patients compared to healthy subjects. In particular, Pseudomonas aeruginosa was enriched in the oral cavity of OSCC patients. Bitter taste receptors (taste 2 receptors, TAS2Rs), known as G protein-coupled receptors (GPCRs), can recognize quorum sensing molecules, such as N-(3-oxododecanoyl)-L-homoserine lactone (oxo-C12) secreted from P. aeruginosa. Based on The Cancer Genome Atlas analysis, downregulation of TAS2Rs was correlated with poor prognosis in various cancers. This prompted us to investigate the role of TAS2Rs in oral dysbiosis and cancers. Here, we found that TAS2Rs mediated both apoptosis and pyroptosis, distinct types of programmed cell death, in response to oxo-C12, in OSCC cell lines. Of note, it was blocked by knockdown of TAS2R4, 14, or 38. Unexpectedly, TAS2Rs-mediated apoptosis and pyroptosis were GPCR-independent. Given our observation that the blockade of NLRP3 inflammasome attenuated oxo-C12-induced OSCC cell death, TAS2Rs seem to recruit NLRP3 inflammasome rather than G protein signaling pathway in order to mediate the programmed cell death in OSCC. Once NLRP3 is activated, caspase-8 triggered both apoptosis and pyroptosis in response to oxo-C12, supporting the role of TAS2Rs-NLRP3-caspase-8 axis. Interestingly, siRNA knockdown of MyD88 diminished oxo-C12-induced programmed cell death, implying the possible involvement of MyD88 in the TAS2Rs-NLRP3-caspase-8 axis. Taken together, bitter taste receptors promote the programmed cell death of OSCC in response to oral dysbiosis through activation of NLRP3 inflammasome, which further supports TAS2Rs as prognostic markers in OSCC. Thus, these results will shed light on better understanding of interplay between microbiome and cancer, proposing TAS2Rs as novel therapeutic targets in oral dysbiosis-induced OSCC development.

Citation Format: Byeong-Oh Hwang, Se-Young Park, Na-Young Song. Bitter taste receptors link between microbiome dysbiosis and programmed cell death in oral squamous cell carcinoma [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 5912.
Abstract 5917: Development of a medium-to-high throughput organoid-bacteria co-culture platform for the assessment of host pathogen interaction
Authors: Valle-Encinas Eider , Nair Roshni , Pisa Katerina , Doorn Mayke , Pourfarzad Farzin , Mateo Lani San , Desch Nicole , Gokare Prashanth , Pocalyko David , Boj Sylvia F. , Verissimo Carla
Published: April 2023

Pathogens such as colibactin-producing bacteria strains of the Enterobacteriaceae family are highly prevalent in the gut microbiota of colorectal cancer (CRC) patients. HUB patient derived Organoids, (HUB-OrganoidsTM or PDOs) are self-organized epithelial cell structures with near-physiological features, extensively used to model aspects of cancer initiation and progression. Studies conducted by Pleguezuelos-Manzanos et al. (2020) showed that repetitive cycles of colibactin-producing bacteria into the organoid lumen via microinjection, led to two co-occurring mutational signatures appearance, a single-base substitution signature (SBS-pks) and an indel signature (ID-pks), identified in a subset of CRC patients (~2% and 7%, respectively). These results highlight the relevance of organoids for modelling host-pathogen interactions and the application of these systems as in vitro platforms for investigating long-standing cause-correlation relationship between the presence of genotoxic pathogens and CRC development. The limited scalability of sophisticated co-culture strategies with pathogens such as microinjection has represented a bottleneck in the use of organoids as throughput screening tools in the development of preventive therapies. Thus, there is an unmet need to develop alternative co-culture systems compatible with the process of compound screening. An organoid-bacteria co-culture system compatible with medium-to-high throughput screening readouts has been developed. This co-culture system is currently tailored for the modeling of colibactin genotoxic effects in the gut epithelium, but theoretically could be extended as a discovery platform to identify targetable, novel and complex interactions between host and pathogen. Key words: colibactin, organoids, CRC, screening

Citation Format: Eider Valle-Encinas, Roshni Nair, Katerina Pisa, Mayke Doorn, Farzin Pourfarzad, Lani San Mateo, Nicole Desch, Prashanth Gokare, David Pocalyko, Sylvia F. Boj, Carla Verissimo. Development of a medium-to-high throughput organoid-bacteria co-culture platform for the assessment of host pathogen interaction [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 5917.