PURPOSE:Breast cancer brain metastasis remains a significant clinical problem. Mucins have been implicated in metastasis; however, whether they are also involved in breast cancer brain metastasis remains unknown. We queried databases of patients with brain metastasis and found mucin 5AC (MUC5AC) to be upregulated and therefore sought to define the role of MUC5AC in breast cancer brain metastasis. EXPERIMENTAL DESIGN:In silico dataset analysis, RNA-sequence profiling of patient samples and cell lines, analysis of patient serum samples, and in vitro/in vivo knockdown experiments were performed to determine the function of MUC5AC in breast cancer brain metastasis. Coimmunoprecipitation was used to unravel the interactions that can be therapeutically targeted. RESULTS:Global in silico transcriptomic analysis showed that MUC5AC is significantly higher in patients with breast cancer brain metastasis. Analysis of archived breast cancer brain metastasis tissue further revealed significantly higher expression of MUC5AC in all breast cancer subtypes, and high MUC5AC expression predicted poor survival in HER2+ breast cancer brain metastasis. We validated these observations in breast cancer brain metastatic cell lines and tissue samples. Interestingly, elevated levels of MUC5AC were detected in the sera of patients with breast cancer brain metastasis. MUC5AC silencing in breast cancer brain metastatic cells reduced their migration and adhesion in vitro and in brain metastasis in the intracardiac injection mouse model. We found high expression of cMET and CD44v6 in breast cancer brain metastasis, which increased MUC5AC expression via hepatocyte growth factor signaling. In addition, MUC5AC interacts with cMET and CD44v6, suggesting that MUC5AC promotes breast cancer brain metastasis via the cMET/CD44v6 axis. Inhibition of the MUC5AC/cMET/CD44v6 axis with the blood-brain barrier-permeable cMET inhibitor bozitinib (PLB1001) effectively inhibits breast cancer brain metastasis. CONCLUSIONS:Our study establishes that the MUC5AC/cMET/CD44v6 axis is critical for breast cancer brain metastasis, and blocking this axis will be a novel therapeutic approach for breast cancer brain metastasis.

Advanced prostate cancer (PCa) remains a significant clinical challenge, and docetaxel plays a significant role in disease management. Despite the efficacy of docetaxel as a first-line chemotherapy, resistance often develops. We developed three clinically relevant in vitro PCa cell models and transcriptomic analysis identified that the Paf1/RNA polymerase II complex component (PAF1)-associated pluripotent-transcription factor (TF), SOX2, plays a crucial role in docetaxel resistance. The cancer stem cell (CSC) transcriptional master regulator PAF1 is significantly higher in PCa cell lines, tumor tissues, and docetaxel resistant (DR) PCa cells than in age-matched control cells. To determine the molecular underlying and functional characteristics of PAF1 in resistance mechanisms, we performed coimmunoprecipitation, embryonic stem cell network proteins, in vitro tumor-initiating ability, and 3D multicellular organoid growth using PAF1 knockdown cells. Tet-inducible PAF1 depletion reduced the drug-efflux phenotype, tumor-initiating frequencies, and three-dimensional organoid growth of the docetaxel-resistant PCa cell lines. Functional studies also showed restoration of docetaxel sensitivity in a 3D tumorsphere model upon PAF1 depletion. PAF1 depletion was also associated with decreased pluripotent TFs and other CSC markers. This study provides a novel regulatory mechanism of docetaxel resistance in PCa through PAF1.

Emerging evidence indicates that antibiotic-induced dysbiosis can play an etiological role in the pathogenesis of neuropsychiatric disorders. However, most of this evidence comes from rodent models. The objective of this study was to evaluate if antibiotic-induced gut dysbiosis can elicit changes in gut metabolites and behavior indicative of gut-brain axis disruption in common marmosets (Callithrix jacchus) - a nonhuman primate model often used to study sociability and stress. We were able to successfully induce dysbiosis in marmosets using a custom antibiotic cocktail (vancomycin, enrofloxacin and neomycin) administered orally for 28 days. This gut dysbiosis altered gut metabolite profiles, behavior, and stress reactivity. Increase in gut Fusobacterium spp. post-antibiotic administration was a novel dysbiotic response and has not been observed in any rodent or human studies to date. There were significant changes in concentrations of several gut metabolites which are either neurotransmitters (e.g., GABA and serotonin) or have been found to be moderators of gut-brain axis communication in rodent models (e.g., short-chain fatty acids and bile acids). There was an increase in affiliative behavior and sociability in antibiotic-administered marmosets, which might be a coping mechanism in response to gut dysbiosis-induced stress. Increase in urinary cortisol levels after multiple stressors provides more definitive proof that this model of dysbiosis may cause disrupted communication between gut and brain in common marmosets. This study is a first attempt to establish common marmosets as a novel model to study the impact of severe gut dysbiosis on gut-brain axis cross-talk and behavior.

Abstract Background: While T-cell-based immunotherapy emerged as a promising tool for cancer management, clinical trials and translational studies revealed that cancer cells develop resistance by modulating tumor microenvironment and cell-intrinsic mechanisms. Pancreatic cancer (PC), consisting of dense stroma, immunosuppressive environment, and aberrant mucin expression, has a dismal survival rate and responds poorly to immunotherapies. MUC4, a member of the mucin family, has been reported to block lymphokine-activated killer cells and induce apoptosis of cytotoxic T-cells, suggesting its possible role in immune modulation. However, how the T-cell secretome influences MUC4 expression in PC and its role in modulating the T-cell response is poorly investigated. Method: The scRNA-seq data analysis and primary T-cell conditioned media (CM) were utilized to investigate the MUC4 and T-cell crosstalk. T-cell CM-treated murine PC cell line (KCT-3266) was analyzed by RNA-seq analysis followed by MUC4 silencing studies to investigate its role in evading T-cell response. The subcutaneous murine model was utilized to investigate the influence of MUC4 KO on immune cells and cytotoxic cell infiltration. The RNA isolated from tumor tissues from KrasG12D/+, Trp53R127H/+, Pdx-1-Cre (KPC), and Muc4 knockout (KPMC) murine models were used for PanCancer immune profiling. Results: The scRNA-seq analysis suggests that intratumoral T-cells positively correlate with MUC4 expression in PC patients. The activated T-cell CM consisting of predominantly IL-2, IFN-γ, and TNF-α cytokines, significantly induce the expression of MUC4 transcriptionally and translationally in both human (SW1990 and COLO357) and murine (KCT-3248 and KCT3266) PC cell lines. RNA-seq analysis from T-cell CM-treated cancer cells revealed that T-cell secretome induced the pathways related to cancer cell death. The CRISPR knockout of MUC4 in PC cells showed increased expression of cleaved caspase-3 after T-cell CM treatment, indicating that MUC4 plays a protective role against the T-cell CM-mediated killing. Subcutaneous implantation of MUC4 proficient and deficient PC cells on the flanks of C57BL/6 immunocompetent mice demonstrated a significantly higher infiltration of CD3 positive and cytotoxic T-cell infiltration in the MUC4 deficient tumors, resulting in significantly lower tumor weight. The PanCancer immune profiling also showed that depletion of Muc4 increases the T-cells and cytotoxic T-cell signaling scores in the KPMC tumor tissues compared to KPC. Conclusion: T-cell secretome induces MUC4 expression in pancreatic cancer cells, and the increased MUC4 expression reduces T-cell infiltration and protects PC cells against T-cell-mediated killing. Citation Format: Xiaoqi Li, Imran Khan, Rachel Kehrberg, Zahraa Wajih Alsafwani, Rakesh Bhatia, Sushil Kumar, Surinder K. Batra. Pancreatic cancer and T-cell crosstalk-induced MUC4 expression attenuates T-cell-mediated response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1594.

MUC16/CA125 is one of the few oldest cancer biomarkers still used in current clinical practice. As mesothelium is an abundant source of MUC16 and a major contributor to stromal heterogeneity in PDAC, we investigated the regulation of MUC16 in tumor and stromal compartments individually. The trajectories constructed using the single-cell transcriptomes of stromal cells from KPC tumors demonstrated continuity in the trajectory path between MUC16-expressing mesothelial cells and other CAF subsets. Further, the tumor tissues of MUC16 whole-body knockout (KPCM) showed dysregulation in the markers of actomyosin assembly and fibroblast differentiation (iCAF and myCAF), indicating that MUC16 has an extra-tumoral role in controlling CAF differentiation. Although we found mesothelium-derivative stromal cells to be bystanders in normal pancreas, the proportion of these cells was higher in invasive PDAC, particularly in TP53 deficient tumors. Moreover, we also detail the regulation of MUC16, KRAS, and SOX9 by TP53 family members (TP53 and TP63) using multi-omics data from knockout models, PDAC cell lines, and human PDAC tissues.

BACKGROUND & AIMS: Pancreatic ductal adenocarcinoma (PDAC) is characterized by desmoplastic stroma surrounding most tumors. Activated stromal fi broblasts, namely cancerassociated fi broblasts (CAFs), play a major role in PDAC progression. We analyzed whether CAFs in fl uence acinar cells and impact PDAC initiation, that is, acinar-to-ductal metaplasia (ADM). ADM connection with PDAC pathophysiology is indicated, but not yet established. We hypothesized that CAF secretome might play a signi fi cant role in ADM in PDAC initiation . METHODS: Mouse and human acinar cell organoids, acinar cells cocultured with CAFs and exposed to CAFconditioned media, acinar cell explants, and CAF cocultures were examined by means of quantitative reverse transcription polymerase chain reaction, RNA sequencing, immunoblotting, and confocal microscopy. Data from liquid chromatography with tandem mass spectrometry analysis of CAF - conditioned medium and RNA sequencing data of acinar cells post - conditioned medium exposure were integrated using bioinformatics tools to identify the molecular mechanism for CAFinduced ADM. Using confocal microscopy, immunoblotting, and quantitative reverse transcription polymerase chain reaction analysis, we validated the depletion of a key signaling axis in the cell line, acinar explant coculture, and mouse cancerassociated fi broblasts (mCAFs). RESULTS: A close association of acino - ductal markers ( Ulex europaeus agglutinin 1, amylase, cytokeratin-19) and mCAFs ( a -smooth muscle actin) in LSLKras G12D/ & thorn; ; LSL-Trp53 R172H/ & thorn; ; Pdx1 Cre (KPC) and LSLKras G12D/ & thorn; ; Pdx1 Cre (KC) autochthonous progression tumor tissue was observed. Caerulein treatment - induced mCAFs increased cytokeratin-19 and decreased amylase in wild -type and KC pancreas. Likewise, acinar-mCAF cocultures revealed the induction of ductal transdifferentiation in cell line, acinarorganoid, and explant coculture formats in WT and KC mice pancreas. Proteomic and transcriptomic data integration revealed a novel laminin a 5/integrin a 4/stat3 axis responsible for CAF-mediated acinar-to-ductal cell transdifferentiation. CONCLUSIONS: Results collectively suggest the fi rst evidence for CAF-in fl uenced acino - ductal phenotypic switchover, thus highlighting the tumor microenvironment role in pancreatic carcinogenesis inception.

Bioinformatics is a broad interdisciplinary field with rapidly increasing workforce needs. Many groups have worked to develop competencies expected of professionals in the bioinformatics discipline as well as specializations within the field. In this work, we examine the current literature to discern case-studies, or other resources are available to bioinformatics learners and educators to begin developing reproducibility related-curricula and/or training.