Manoj Garg

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive malignancies globally. PDAC remains a leading cause of cancer-related mortality due to a high rate of metastasis, late diagnosis, and higher therapeutic resistance. Therefore, the limited effectiveness of existing therapeutic regimens highlights the need for continued development of new anticancer agents. Recently, pyridine-linked benzamide compounds have been explored and reported to have promising anticancer activity. Henceforth, we synthesized 3-(Arylsulfonamido)-N-(pyridin-2/3-yl)benzamide derivatives for evaluating their anticancer potential against PDAC cells. The structures of the compounds (5a-d) were evaluated by 1H NMR, 13C NMR, and MALDI-TOF mass spectrometry. During initial screening in SW1990 cells, the 5a derivative demonstrated marked and significant anticancer activity among all the synthesized derivatives. Hence, the derivative 5a was selected for evaluation of its anticancer potential against a panel of PDAC cell lines. The treatment of SW1990, Panc10.05, Capan-2, CFPAC-1, and MIA PaCa-2 cells with the 5a derivative showed significant inhibition in cell viability and clonogenicity. The treatment of 5a derivative induced G2/M arrest with marked accumulation of SW1990 and Panc10.05 cells in the sub-G1 phase during cell cycle analysis. Further, Annexin V/PI staining displayed that 5a treatment significantly induced apoptotic cell death in SW1990 and Panc10.05 cells. Mechanistic studies revealed a marked downregulation of cyclin D1 and CDK-6 protein expression, associated with the regulation of cell cycle arrest. Interestingly, the 5a derivative showed either a low or a non-significant effect on the growth, clonogenicity, and apoptosis of the hTERT-HPNE, normal pancreatic cells. Furthermore, molecular docking of the 5a derivative showed binding affinity with the AKT1 (PDB ID: 4EJN), mTOR (PDB ID: 4JT6), and STAT3 (PDB ID: 6NJS) proteins. The molecular analysis, conducted through a Western blot experiment in PDAC cells, revealed that the 5a derivative suppressed the phosphorylation of AKT, mTOR, and STAT3 in SW1990 and Capan-2 cells. Taken together, the results of molecular docking, molecular dynamics, and biological assays demonstrated that 5a exhibited promising anticancer activity by regulating the AKT/mTOR /STAT3 axis in PDAC with therapeutic potential.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive and often fatal cancer with limited treatment options. Small nucleolar RNA host gene 10 (SNHG10) has emerged as a key regulator in the progression and metastasis of human cancers. However, the potential of SNHG10 in PDAC tumorigenesis, gemcitabine resistance, and the underlying molecular mechanisms remains poorly understood. Our data analysis revealed significant upregulation of the SNHG10 transcript in 179 PDAC cases compared with 171 normal pancreatic specimens, with a positive association with clinical stages of PDAC. Further, we confirmed the significant overexpression of the SNHG10 transcript in several PDAC cell lines compared to normal pancreatic cells. Our results revealed that the downregulation of SNHG10 significantly decreased the cellular proliferation, clonogenic ability, cell migration, and the epithelial to mesenchymal transition, leading to the induction of cell cycle arrest and apoptosis of PDAC cells. Mechanistically, the downregulation of SNHG10 significantly inhibited the expression of vimentin, N-cadherin, survivin, CDK4, CDK6, cyclin B1, cyclin D1, aurora kinase A, and aurora kinase B, with an increased expression of E-cadherin and p21. The bioinformatics analysis, RNA Immunoprecipitation, and qRT-PCR results showed the physical interaction among SNHG10, miR-150-5p, and VEGF-A, which are the integral parts of the ternary complex in PDAC cell lines. Interestingly, silencing of SNHG10 led to the significant induction of miR-150-5p, which repressed the expression of VEGF-A in PDAC cells. Moreover, the miR-150-5p rescued the VEGF-A expression in PDAC cells even during the silencing of SNHG10. Interestingly, the downregulation of SNHG10 enhanced gemcitabine sensitivity in gemcitabine-resistant PDAC cells. The depletion of SNHG10 in the PDAC xenograft model significantly reduced tumor growth, volume, and weight. Importantly, downregulation of SNHG10 suppressed the phosphorylation of EGFR, AKT, ERK1/2, mTOR, and c-MET signaling pathways in both in vitro and xenograft models of PDAC. Our study unveils the oncogenic potential of SNHG10 in tumorigenesis through modulation of the EGFR/AKT/ERK/mTOR, and miR-150-5p/VEGF-A axis, as well as gemcitabine resistance of PDAC as a prospective therapeutic strategy.

Oral cancer, a major form of head and neck cancer (HNC), remains clinically challenging due to its aggressiveness, and therapeutic resistance. Tumor microenvironment (TME) is one of the key factors of cancer progression which strongly influences how cancer cells survive, spread, and respond to treatment. However, the molecular factors underlying its dysregulation are not fully understood. Recent data suggest that oncogenic forms of p53, particularly the mutant R273H and its amyloidogenic conformer, display gain-of-functon(GOF) phenotype. In this study, we investigated how oncogenic p53 remodel the secretome to affect the extracellular matrix (ECM) organization. Proteomic analysis of conditioned media from AW13516 (tongue carcinoma) cells show distinct ECM-related secreted proteins. Among the differentially enriched proteins, Inter-Alpha-Trypsin Inhibitor Heavy Chain H2 (ITIH2) and Clusterin (CLU) were secreted in amyloid p53 and mutant p53 expressing cells. Super-resolution microscopy revealed spatial co-localization of both ITIH2 and CLU with intracellular mutant p53 aggregates. Further, exosome isolation and analysis confirmed that both proteins are actively packaged and released through exosomes. Functional studies showed that ITIH2 silencing induced apoptosis through inhibition of the AKT/NF-κB pathway. Similarly, the CLU silencing reduced Vimentin and mTOR levels, while increasing BAX and E-cadherin levels, indicating loss of stemness, induction of apoptosis and partial restoration of epithelial features. To identify potential therapeutic vulnerabilities, molecular docking analysis revealed that methotrexate (MTX) binds strongly and specifically to both ITIH2 and CLU. Consistent with these predictions, MTX treatment combined with ITIH2 or CLU silencing produced a synergistic effect on the suppression of proliferation and migration. Overall, our findings identify ITIH2 and CLU as exosome-associated downstream effectors of oncogenic p53 that contribute to tumor microenvironment dysregulation. Thus, targeting this secretome axis, particularly, in combination with MTX may offer a promising strategy to limit oral cancer progression.

SNHG10 emerged as a key regulator in progression and metastasis of cancers. However, potential of SNHG10 in PDAC tumorigenesis, gemcitabine resistance, and underlying mechanisms remains poorly understood. We observed significant upregulation of SNHG10 in 179 PDAC cases, revealing a positive correlation with clinical stages. Our results showed a significant SNHG10 overexpression in several PDAC cells. Downregulation of SNHG10 significantly decreased the proliferation, clonogenicity, EMT, tumor growth in the xenograft model, and the induction of cell cycle arrest and apoptosis of PDAC cells. Mechanistically, SNHG10 knockdown significantly inhibited the expression of vimentin, N-cadherin, survivin, CDK4, CDK6, cyclin B1, cyclin D1, aurora kinase A, and B, with an increased expression of E-cadherin and p21. RNA Immunoprecipitation data displayed physical interaction among SNHG10, miR-150-5p, and VEGFA in PDAC cells. SNHG10 silencing led to the significant induction of miR-150-5p, which repressed VEGFA expression in PDAC cells. SNHG10 downregulation enhanced gemcitabine sensitivity in PDAC cells. SNHG10 silencing suppressed the phosphorylation of EGFR, AKT, ERK1/2, mTOR, and c-MET pathways. Silencing SNHG10 and its regulated signalling offers a novel prospective therapeutic strategy.

Breast cancer metastasis remains the leading cause of cancer-related deaths in women worldwide. Infiltration of tumor-associated macrophages (TAMs) in the tumor stroma is known to be correlated with reduced overall survival. The inhibitors of TAMs are sought after for reprogramming the tumor microenvironment. Signal transducer and activator of transcription 3 (STAT3) is well known to contribute in pro-tumoral properties of TAMs. 2-Methoxyestradiol (2ME2), a potent anticancer and antiangiogenic agent, has been in clinical trials for treatment of breast cancer. Here, we investigated the potential of 2ME2 in modulating the pro-tumoral effects of TAMs in breast cancer. THP-1-derived macrophages were polarized to macrophages with or without 2ME2. The effect of 2ME2 on macrophage surface markers and anti-inflammatory genes was determined by Western blotting, flow cytometry, immunofluorescence, qRT‒PCR. The concentration of cytokines secreted by cells was monitored by ELISA. The effect of M2 macrophages on malignant properties of breast cancer cells was determined using colony formation, wound healing, transwell, and gelatin zymography assays. An orthotopic model of breast cancer was used to determine the effect of 2ME2 on macrophage polarization and metastasis in vivo. First, our study found that polarization of monocytes to alternatively activated M2 macrophages is associated with the reorganization of the microtubule cytoskeleton. At lower concentrations, 2ME2 treatment depolymerized microtubules and reduced the expression of CD206 and CD163, suggesting that it inhibits the polarization of macrophages to M2 phenotype. However, the M1 polarization was not significantly affected at these concentrations. Importantly, 2ME2 inhibited the expression of several anti-inflammatory cytokines and growth factors, including CCL18, TGF-β, IL-10, FNT, arginase, CXCL12, MMP9, and VEGF-A, and hindered the metastasis-promoting effects of M2 macrophages. Concurrently, 2ME2 treatment reduced the expression of CD163 in tumors and inhibited lung metastasis in the orthotopic breast cancer model. Mechanistically, 2ME2 treatment reduced the phosphorylation and nuclear translocation of STAT3, an effect which was abrogated by colivelin. Our study presents novel findings on mechanism of 2ME2 from the perspective of its effects on the polarization of the TAMs via the STAT3 signaling in breast cancer. Altogether, the data supports further clinical investigation of 2ME2 and its derivatives as therapeutic agents to modulate the tumor microenvironment and immune response in breast carcinoma.