Botensilimab, an Fc-enhanced anti-CTLA-4 antibody, is effective against tumors poorly responsive to conventional immunotherapy.

Conventional immune checkpoint inhibitors (ICI) targeting CTLA-4 elicit durable survival, but primarily in patients with immune-inflamed tumors. Although the mechanisms underlying response to anti-CTLA-4 remain poorly understood, Fc-gamma receptor (FcγR) IIIA co-engagement appears critical for activity, potentially explaining the modest clinical benefits of approved anti-CTLA-4 antibodies. We demonstrate that anti-CTLA-4 engineered for enhanced FcγR affinity leverages FcγR-dependent mechanisms to potentiate T cell responsiveness, reduce intratumoral Tregs, and enhance antigen presenting cell activation. Fc-enhanced anti-CTLA-4 promoted superior efficacy in mouse models and remodeled innate and adaptive immunity versus conventional anti-CTLA-4. These findings extend to patients treated with botensilimab, an Fc-enhanced anti-CTLA-4 antibody, with clinical activity across multiple poorly immunogenic and ICI treatment-refractory cancers. Efficacy was independent of tumor neoantigen burden or FcγRIIIA genotype. However, FcγRIIA and FcγRIIIA expression emerged as potential response biomarkers. These data highlight the therapeutic potential of Fc-enhanced anti-CTLA-4 antibodies in cancers unresponsive to conventional ICI therapy.

Aronia Berry Extract Modulates MYD88/NF-kB/P-Glycoprotein Axis to Overcome Gemcitabine Resistance in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease with poor survival rates, primarily due to the limited effectiveness of gemcitabine (Gem)-based chemotherapy, as well as the acquisition of chemotherapeutic resistance. Aronia berry extracts (ABEs), abundant in phenolic constituents, have been recently recognized for their anticancer properties as well as their encouraging potential to help overcome chemoresistance in various cancers. In the present study, we explored ABE's potential to overcome Gem resistance in PDAC and identify specific growth regulatory pathways responsible for its anticancer activity. Through a series of in vitro experiments in gemcitabine-resistant (Gem-R) cells, we elucidated the synergistic interactions between Gem and ABE treatments. Using advanced transcriptomic analysis and network pharmacology, we revealed key molecular pathways linked to chemoresistance and potential therapeutic targets of ABE in Gem-R PDAC cells. Subsequently, the findings from cell culture studies were validated in patient-derived 3D tumor organoids (PDOs). The combination treatment of ABE and Gem demonstrated significant synergism and anticancer effects on cell viability, proliferation, migration, and invasion in Gem-R cells. Transcriptomic analysis revealed a correlation between the NF-Κb signaling pathway and Gem-R (p < 0.05), exhibiting a marked upregulation of MYD88. Additionally, MYD88 exhibited a significant correlation with the overall survival rates in patients with PDAC patients in the TCGA cohort (HR = 1.58, p < 0.05). The MYD88/NF-Κb pathway contributes to chemoresistance by potentially upregulating efflux transporters like P-glycoprotein (P-gp). Our findings revealed that the combined treatment with ABE suppressed the NF-Κb pathway by targeting MYD88 and reducing P-gp expression to overcome Gem resistance. Lastly, the combination therapy proved highly effective in PDOs in reducing both their number and size (p < 0.05). Our study offers previously unrecognized insights into the ability of ABE to overcome Gem resistance in PDAC cells through its targeting of the MYD88/NF-κb/P-gp axis, hence providing a safe and cost-effective adjunctive therapeutic strategy to improve treatment outcomes in PDAC.

Tumor and Peritoneum-Associated Macrophage Gene Signature as a Novel Molecular Biomarker in Gastric Cancer.

A spectrum of immune states resulting from tumor resident macrophages and T-lymphocytes in the solid tumor microenvironment correlates with patient outcomes. We hypothesized that in gastric cancer (GC), macrophages in a polarized immunosuppressive transcriptional state would be prognostic of poor survival. We derived transcriptomic signatures for M2 (M2TS, MRC1; MS4A4A; CD36; CCL13; CCL18; CCL23; SLC38A6; FGL2; FN1; MAF) and M1 (M1TS, CCR7; IL2RA; CXCL11; CCL19; CXCL10; PLA1A; PTX3) macrophages, and cytolytic T-lymphocytes (CTLTS, GZMA; GZMB; GZMH; GZMM; PRF1). Primary GC in a TCGA stomach cancer dataset was evaluated for signature expressions, and a log-rank test determined overall survival (OS) and the disease-free interval (DFI). In 341 TCGA GC entries, high M2TS expression was associated with histological types and later stages. Low M2TS expression was associated with significantly better 5-year OS and DFI. We validated M2TS in prospectively collected peritoneal fluid of a GC patient cohort (n = 28). Single-cell RNA sequencing was used for signature expression in CD68+CD163+ cells and the log-rank test compared OS. GC patients with high M2TS in CD68+CD163+ cells in their peritoneal fluid had significantly worse OS than those with low expression. Multivariate analyses confirmed M2TS was significantly and independently associated with survival. As an independent predictor of poor survival, M2TS may be prognostic in primary tumors and peritoneal fluid of GC patients.

Inhibiting stromal Class I HDACs curbs pancreatic cancer progression.

Oncogenic lesions in pancreatic ductal adenocarcinoma (PDAC) hijack the epigenetic machinery in stromal components to establish a desmoplastic and therapeutic resistant tumor microenvironment (TME). Here we identify Class I histone deacetylases (HDACs) as key epigenetic factors facilitating the induction of pro-desmoplastic and pro-tumorigenic transcriptional programs in pancreatic stromal fibroblasts. Mechanistically, HDAC-mediated changes in chromatin architecture enable the activation of pro-desmoplastic programs directed by serum response factor (SRF) and forkhead box M1 (FOXM1). HDACs also coordinate fibroblast pro-inflammatory programs inducing leukemia inhibitory factor (LIF) expression, supporting paracrine pro-tumorigenic crosstalk. HDAC depletion in cancer-associated fibroblasts (CAFs) and treatment with the HDAC inhibitor entinostat (Ent) in PDAC mouse models reduce stromal activation and curb tumor progression. Notably, HDAC inhibition (HDACi) enriches a lipogenic fibroblast subpopulation, a potential precursor for myofibroblasts in the PDAC stroma. Overall, our study reveals the stromal targeting potential of HDACi, highlighting the utility of this epigenetic modulating approach in PDAC therapeutics.

Advanced glycation end product (AGE) targeting antibody SIWA318H is efficacious in preclinical models for pancreatic cancer.

SIWA318H is a novel monoclonal antibody that selectively targets an advanced glycation end product biomarker found in damaged/dysfunctional cells exhibiting (a) aerobic glycolysis, and (b) oxidative stress. Cells with this biomarker are dysfunctional and are associated with stresses and/or damages relating to aging, cancer and other disease processes. In this study, we evaluated the biological effects and antitumor activity of SIWA318H in preclinical models for pancreatic cancer. SIWA318H binds to pancreatic cancer cells and cancer-associated fibroblasts, as well as tumor xenografts derived from pancreatic cancer patients. Furthermore, SIWA318H induced significant antibody-dependent cell-mediated cytotoxicity (ADCC) against pancreatic cancer cells. In a humanized CD34+ NSG mouse xenograft model for pancreatic cancer, tumors in mice treated with SIWA318H grew significantly slower compared to those in control mice (p < 0.001). After 3 weeks of treatment with SIWA318H, the tumor growth was suppressed by 68.8% and 61.5% for the high and low dose regimens, respectively, when compared to the isotype antibody control (ANOVA p < 0.002). Moreover, a significant increase in complete remission (CR) rate was observed in mice receiving the high dose (60%, p < 0.04) or low dose (77.8%, p < 0.02) of SIWA318H treatment compared with control mice (6.7%). Immunohistochemical analyses of the tumor tissues showed a significant decrease in senescent cells in the tumor microenvironment of SIWA318H treated mice compared to that of control treated mice (p < 0.05). These results provide compelling evidence that SIWA318H is a promising novel therapeutic against pancreatic cancer.

Single-cell mapping identifies MSI+ cells as a common origin for diverse subtypes of pancreatic cancer.

Identifying the cells from which cancers arise is critical for understanding the molecular underpinnings of tumor evolution. To determine whether stem/progenitor cells can serve as cells of origin, we created a Msi2-CreERT2 knock-in mouse. When crossed to CAG-LSL-MycT58A mice, Msi2-CreERT2 mice developed multiple pancreatic cancer subtypes: ductal, acinar, adenosquamous, and rare anaplastic tumors. Combining single-cell genomics with computational analysis of developmental states and lineage trajectories, we demonstrate that MYC preferentially triggers transformation of the most immature MSI2+ pancreas cells into multi-lineage pre-cancer cells. These pre-cancer cells subsequently diverge to establish pancreatic cancer subtypes by activating distinct transcriptional programs and large-scale genomic changes, and enforced expression of specific signals like Ras can redirect subtype specification. This study shows that multiple pancreatic cancer subtypes can arise from a common pool of MSI2+ cells and provides a powerful model to understand and control the programs that shape divergent fates in pancreatic cancer.

Inhibiting Stromal Class I HDACs Curbs Pancreatic Cancer Progression.

Oncogenic lesions in pancreatic ductal adenocarcinoma (PDAC) hijack the epigenetic machinery in stromal components to establish a desmoplastic and therapeutic resistant tumor microenvironment (TME). Here we identify Class I histone deacetylases (HDACs) as key epigenetic factors facilitating the induction of pro-desmoplastic and pro-tumorigenic transcriptional programs in pancreatic stromal fibroblasts. Mechanistically, HDAC-mediated changes in chromatin architecture enable the activation of pro-desmoplastic programs directed by serum response factor (SRF) and forkhead box M1 (FOXM1). HDACs also coordinate fibroblast pro-inflammatory programs inducing leukemia inhibitory factor (LIF) expression, supporting paracrine pro-tumorigenic crosstalk. HDAC depletion in cancer-associated fibroblasts (CAFs) and treatment with the HDAC inhibitor entinostat (Ent) in PDAC mouse models reduce stromal activation and curb tumor progression. Notably, HDAC inhibition (HDACi) enriches a lipogenic fibroblast subpopulation, a potential precursor for myofibroblasts in the PDAC stroma. Overall, our study reveals the stromal targeting potential of HDACi, highlighting the utility of this epigenetic modulating approach in PDAC therapeutics.

A super-enhancer-regulated RNA-binding protein cascade drives pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy in need of new therapeutic options. Using unbiased analyses of super-enhancers (SEs) as sentinels of core genes involved in cell-specific function, here we uncover a druggable SE-mediated RNA-binding protein (RBP) cascade that supports PDAC growth through enhanced mRNA translation. This cascade is driven by a SE associated with the RBP heterogeneous nuclear ribonucleoprotein F, which stabilizes protein arginine methyltransferase 1 (PRMT1) to, in turn, control the translational mediator ubiquitin-associated protein 2-like. All three of these genes and the regulatory SE are essential for PDAC growth and coordinately regulated by the Myc oncogene. In line with this, modulation of the RBP network by PRMT1 inhibition reveals a unique vulnerability in Myc-high PDAC patient organoids and markedly reduces tumor growth in male mice. Our study highlights a functional link between epigenetic regulation and mRNA translation and identifies components that comprise unexpected therapeutic targets for PDAC.

Andrographis Reverses Gemcitabine Resistance through Regulation of ERBB3 and Calcium Signaling Pathway in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies, primarily due to intrinsic or acquired resistance to chemotherapy, such as Gemcitabine (Gem). Naturally occurring botanicals, including Andrographis (Andro), can help enhance the anti-tumorigenic therapeutic efficacy of conventional chemotherapy through time-tested safety and cost-effectiveness. Accordingly, we hypothesized that Andro might reverse Gem resistance in PDAC. The critical regulatory pathways associated with Gem resistance in PDAC were identified by analyzing publicly available transcriptomic profiling and PDAC tissue specimens. A series of systematic in vitro experiments were performed using Gem-resistant (Gem-R) PDAC cells and patient-derived 3D-organoids to evaluate the Andro-mediated reversal of Gem resistance in PDAC. Transcriptomic profiling identified the calcium signaling pathway as a critical regulator of Gem-resistance (Fold enrichment: 2.8, p = 0.002). Within this pathway, high ERBB3 expression was significantly associated with poor prognosis in PDAC patients. The combination of Andro and Gem exhibited superior anti-cancer potential in Gem-R PDAC cells through potentiating cellular apoptosis. The combined treatment down-regulated ERBB3 and decreased intracellular calcium concentration in Gem-R PDAC cells. Finally, these findings were successfully interrogated in patient-derived 3D-organoids. In conclusion, we demonstrate novel evidence for Andro-mediated reversal of chemoresistance to Gem in PDAC cells through the regulation of ERBB3 and calcium signaling.

Effect of acoustic cluster therapy (ACT®) combined with chemotherapy in a patient-derived xenograft mouse model of pancreatic cancer.

Pancreatic ductal adenocarcinomas respond poorly to chemotherapy, in part due to the dense tumor stroma that hinders drug delivery. Ultrasound (US) in combination with microbubbles has previously shown promise as a means to improve drug delivery, and the therapeutic efficacy of ultrasound-mediated drug delivery is currently being evaluated in multiple clinical trials. However, most of these utilize echogenic contrast agents engineered for imaging, which might not be optimal compared to specialized formulations tailored for drug delivery. In this study, we evaluated the in vivo efficacy of phase-shifting microbubble-microdroplet clusters that, upon insonation, form bubbles in the size range of 20-30 µm. We developed a patient-derived xenograft model of pancreatic cancer implanted in mice that largely retained the stromal content of the originating tumor and compared tumor growth in mice given chemotherapeutics (nab-paclitaxel plus gemcitabine or liposomal irinotecan) with mice given the same chemotherapeutics in addition to ultrasound and acoustic cluster therapy. We found that acoustic cluster therapy significantly improved the effect of both chemotherapeutic regimens and resulted in 7.2 times higher odds of complete remission of the tumor compared to the chemotherapeutics alone.

Berberine Overcomes Gemcitabine-Associated Chemoresistance through Regulation of Rap1/PI3K-Akt Signaling in Pancreatic Ductal Adenocarcinoma.

Gemcitabine (Gem)-based chemotherapy is one of the first-line treatments for pancreatic ductal adenocarcinoma (PDAC). However, its clinical effect is limited due to development of chemoresistance. Various naturally occurring compounds, including Berberine (BBR), provide an anti-cancer efficacy with time-tested safety, individually and in combination with chemotherapeutic drugs. Accordingly, we hypothesized that BBR might enhance the chemosensitivity to Gem in PDAC. In this study, cell culture studies using MIA PaCa-2 and BxPC-3 cells, followed by analysis in patient-derived organoids were performed to evaluate the anti-cancer effects of BBR in PDAC. Considering that cancer is a significant manifestation of increased chronic inflammatory stress, systems biology approaches are prudent for the identification of molecular pathways and networks responsible for phytochemical-induced anti-cancer activity, we used these approaches for BBR-mediated chemosensitization to Gem. Firstly, Gem-resistant (Gem-R) PDAC cells were established, and the combination of BBR and Gem revealed superior anti-cancer efficacy in Gem-R cells. Furthermore, the combination treatment induced cell cycle arrest and apoptosis in Gem-R PDAC cells. Transcriptomic profiling investigated the Rap1 and PI3K-Akt signaling pathway as a key regulator of Gem-resistance and was a key mediator for BBR-mediated chemosensitization in PDAC cells. All cell culture-based findings were successfully validated in patient-derived organoids. In conclusion, we demonstrate that BBR-mediated reversal of chemoresistance to Gem manifests through Rap1/PI3K-Akt signaling in PDAC.

Eicosanoids in the pancreatic tumor microenvironment - a multicellular, multifaceted progression.

Background and Aims: Eicosanoids, oxidized fatty acids that serve as cell-signaling molecules, have been broadly implicated in tumorigenesis. Here, we aimed to identify eicosanoids associated with pancreatic tumorigenesis and the cell types responsible for their synthesis. Methods: We profiled normal pancreas and pancreatic ductal adenocarcinoma (PDAC) in mouse models and patient samples using mass spectrometry. We interrogated RNA sequencing datasets for eicosanoid synthase or receptor expression. Findings were confirmed by immunostaining. Results: In murine models, we identified elevated levels of PGD2, prostacyclin, and thromboxanes in neoplasia while PGE2, 12-HHTre, HETEs, and HDoHEs are elevated specifically in tumors. Analysis of scRNA-seq datasets suggests that PGE2 and prostacyclins are derived from fibroblasts, PGD2 and thromboxanes from myeloid cells, and PGD2 and 5-HETE from tuft cells. In patient samples, we identified a transition from PGD2 to PGE2-producing enzymes in the epithelium during the transition to PDAC, fibroblast/tumor expression of PTGIS, and myeloid/tumor cell expression of TBXAS1. Conclusions: Our analyses identify key changes in eicosanoid species during pancreatic tumorigenesis and the cell types that contribute to their synthesis. Thromboxane and prostacyclin expression is conserved between animal models and human disease and may represent new druggable targets.

Pharmacologic Targeting of TFIIH Suppresses KRAS-Mutant Pancreatic Ductal Adenocarcinoma and Synergizes with TRAIL.

Pancreatic ductal adenocarcinoma (PDAC) typically presents as metastatic disease at diagnosis and remains refractory to treatment. Next-generation sequencing efforts have described the genomic landscape, classified molecular subtypes, and confirmed frequent alterations in major driver genes, with coexistent alterations in KRAS and TP53 correlating with the highest metastatic burden and poorest outcomes. However, translating this information to guide therapy remains a challenge. By integrating genomic analysis with an arrayed RNAi druggable genome screen and drug profiling of a KRAS/TP53 mutant PDAC cell line derived from a patient-derived xenograft (PDCL), we identified numerous targetable vulnerabilities that reveal both known and novel functional aspects of pancreatic cancer biology. A dependence on the general transcription and DNA repair factor TFIIH complex, particularly the XPB subunit and the CAK complex (CDK7/CyclinH/MAT1), was identified and further validated utilizing a panel of genomically subtyped KRAS mutant PDCLs. TFIIH function was inhibited with a covalent inhibitor of CDK7/12/13 (THZ1), a CDK7/CDK9 kinase inhibitor (SNS-032), and a covalent inhibitor of XPB (triptolide), which led to disruption of the protein stability of the RNA polymerase II subunit RPB1. Loss of RPB1 following TFIIH inhibition led to downregulation of key transcriptional effectors of KRAS-mutant signaling and negative regulators of apoptosis, including MCL1, XIAP, and CFLAR, initiating caspase-8 dependent apoptosis. All three drugs exhibited synergy in combination with a multivalent TRAIL, effectively reinforcing mitochondrial-mediated apoptosis. These findings present a novel combination therapy, with direct translational implications for current clinical trials on metastatic pancreatic cancer patients. Significance: This study utilizes functional genetic and pharmacological profiling of KRAS-mutant pancreatic adenocarcinoma to identify therapeutic strategies and finds that TFIIH inhibition synergizes with TRAIL to induce apoptosis in KRAS-driven pancreatic cancer.

An Exosome-based Transcriptomic Signature for Noninvasive, Early Detection of Patients With Pancreatic Ductal Adenocarcinoma: A Multicenter Cohort Study.

BACKGROUND & AIMS: Pancreatic ductal adenocarcinoma (PDAC) incidence is rising worldwide, and most patients present with an unresectable disease at initial diagnosis. Measurement of carbohydrate antigen 19-9 (CA19-9) levels lacks adequate sensitivity and specificity for early detection; hence, there is an unmet need to develop alternate molecular diagnostic biomarkers for PDAC. Emerging evidence suggests that tumor-derived exosomal cargo, particularly micro RNAs (miRNAs), offer an attractive platform for the development of cancer-specific biomarkers. Herein, genomewide profiling in blood specimens was performed to develop an exosome-based transcriptomic signature for noninvasive and early detection of PDAC. METHODS: Small RNA sequencing was undertaken in a cohort of 44 patients with an early-stage PDAC and 57 nondisease controls. Using machine-learning algorithms, a panel of cell-free (cf) and exosomal (exo) miRNAs were prioritized that discriminated patients with PDAC from control subjects. Subsequently, the performance of the biomarkers was trained and validated in independent cohorts (n = 191) using quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays. RESULTS: The sequencing analysis initially identified a panel of 30 overexpressed miRNAs in PDAC. Subsequently using qRT-PCR assays, the panel was reduced to 13 markers (5 cf- and 8 exo-miRNAs), which successfully identified patients with all stages of PDAC (area under the curve [AUC] = 0.98 training cohort; AUC = 0.93 validation cohort); but more importantly, was equally robust for the identification of early-stage PDAC (stages I and II; AUC = 0.93). Furthermore, this transcriptomic signature successfully identified CA19-9 negative cases (<37 U/mL; AUC = 0.96), when analyzed in combination with CA19-9 levels, significantly improved the overall diagnostic accuracy (AUC = 0.99 vs AUC = 0.86 for CA19-9 alone). CONCLUSIONS: In this study, an exosome-based liquid biopsy signature for the noninvasive and robust detection of patients with PDAC was developed.

A phase II trial of the super-enhancer inhibitor Minnelide™ in advanced refractory adenosquamous carcinoma of the pancreas.

Adenosquamous carcinoma of the pancreas (ASCP) is a very rare and highly aggressive variant of pancreatic ductal adenocarcinoma, accounting for 0.5-4% of all pancreatic cancer cases in the USA. Current data indicate that epigenetic changes and MYC overexpression lead to squamous transdifferentiation of pancreatic tumor cells and development of ASCP. Minnelide™, an oral anti-super-enhancer drug that inhibits MYC expression in preclinical models of ASCP, has demonstrated safety in a phase I study. We describe the design for a phase II, open-label, single-arm trial of Minnelide in patients with advanced refractory ASCP.

Adenosquamous carcinoma of the pancreas (ASCP) is a rare and highly aggressive variant of pancreatic cancer, with limited treatment options. Changes in activation of DNA elements called super-enhancers drive the growth of ASCP. Minnelide™ is an oral drug that blocks the super-enhancer network and is safe to give to patients with advanced cancer. This trial is designed to determine whether Minnelide can shrink tumors in patients with ASCP who have already received at least one previous treatment for their cancer.  Clinical Trial Registration: NCT04896073 (ClinicalTrials.gov).

Ketogenic diet and chemotherapy combine to disrupt pancreatic cancer metabolism and growth.

Background: Ketogenic diet is a potential means of augmenting cancer therapy. Here, we explore ketone body metabolism and its interplay with chemotherapy in pancreatic cancer. Methods: Metabolism and therapeutic responses of murine pancreatic cancer were studied using KPC primary tumors and tumor chunk allografts. Mice on standard high-carbohydrate diet or ketogenic diet were treated with cytotoxic chemotherapy (nab-paclitaxel, gemcitabine, cisplatin). Metabolic activity was monitored with metabolomics and isotope tracing, including 2H- and 13C-tracers, liquid chromatography-mass spectrometry, and imaging mass spectrometry. Findings: Ketone bodies are unidirectionally oxidized to make NADH. This stands in contrast to the carbohydrate-derived carboxylic acids lactate and pyruvate, which rapidly interconvert, buffering NADH/NAD. In murine pancreatic tumors, ketogenic diet decreases glucose's concentration and tricarboxylic acid cycle contribution, enhances 3-hydroxybutyrate's concentration and tricarboxylic acid contribution, and modestly elevates NADH, but does not impact tumor growth. In contrast, the combination of ketogenic diet and cytotoxic chemotherapy substantially raises tumor NADH and synergistically suppresses tumor growth, tripling the survival benefits of chemotherapy alone. Chemotherapy and ketogenic diet also synergize in immune-deficient mice, although long-term growth suppression was only observed in mice with an intact immune system. Conclusions: Ketogenic diet sensitizes murine pancreatic cancer tumors to cytotoxic chemotherapy. Based on these data, we have initiated a randomized clinical trial of chemotherapy with standard versus ketogenic diet for patients with metastatic pancreatic cancer (NCT04631445).

Deprogramming metabolism in pancreatic cancer with a bi-functional GPR55 inhibitor and biased ß2 adrenergic agonist.

Metabolic reprogramming contributes to oncogenesis, tumor growth, and treatment resistance in pancreatic ductal adenocarcinoma (PDAC). Here we report the effects of (R,S')-4'-methoxy-1-naphthylfenoterol (MNF), a GPR55 antagonist and biased ß2-adrenergic receptor (ß2-AR) agonist on cellular signaling implicated in proliferation and metabolism in PDAC cells. The relative contribution of GPR55 and ß2-AR in (R,S')-MNF signaling was explored further in PANC-1 cells. Moreover, the effect of (R,S')-MNF on tumor growth was determined in a PANC-1 mouse xenograft model. PANC-1 cells treated with (R,S')-MNF showed marked attenuation in GPR55 signal transduction and function combined with increased ß2-AR/Gαs/adenylyl cyclase/PKA signaling, both of which contributing to lower MEK/ERK, PI3K/AKT and YAP/TAZ signaling. (R,S')-MNF administration significantly reduced PANC-1 tumor growth and circulating L-lactate concentrations. Global metabolic profiling of (R,S')-MNF-treated tumor tissues revealed decreased glycolytic metabolism, with a shift towards normoxic processes, attenuated glutamate metabolism, and increased levels of ophthalmic acid and its precursor, 2-aminobutyric acid, indicative of elevated oxidative stress. Transcriptomics and immunoblot analyses indicated the downregulation of gene and protein expression of HIF-1α and c-Myc, key initiators of metabolic reprogramming in PDAC. (R,S')-MNF treatment decreased HIF-1α and c-Myc expression, attenuated glycolysis, shifted fatty acid metabolism towards ß-oxidation, and suppressed de novo pyrimidine biosynthesis in PANC-1 tumors. The results indicate a potential benefit of combined GPR55 antagonism and biased ß2-AR agonism in PDAC therapy associated with the deprogramming of altered cellular metabolism.

Increasing Stress to Induce Apoptosis in Pancreatic Cancer via the Unfolded Protein Response (UPR).

High rates of cell proliferation and protein synthesis in pancreatic cancer are among many factors leading to endoplasmic reticulum (ER) stress. To restore cellular homeostasis, the unfolded protein response (UPR) activates as an adaptive mechanism through either the IRE1α, PERK, or ATF6 pathways to reduce the translational load and process unfolded proteins, thus enabling tumor cells to proliferate. Under severe and prolonged ER stress, however, the UPR may promote adaptation, senescence, or apoptosis under these same pathways if homeostasis is not restored. In this review, we present evidence that high levels of ER stress and UPR activation are present in pancreatic cancer. We detail the mechanisms by which compounds activate one or many of the three arms of the UPR and effectuate downstream apoptosis and examine available data on the pre-clinical and clinical-phase ER stress inducers with the potential for anti-tumor efficacy in pancreatic cancer. Finally, we hypothesize a potential new approach to targeting pancreatic cancer by increasing levels of ER stress and UPR activation to incite apoptotic cell death.

CF33-hNIS-antiPDL1 virus primes pancreatic ductal adenocarcinoma for enhanced anti-PD-L1 therapy.

Immunotherapeutic strategies that combine oncolytic virus (OV) and immune checkpoint inhibitors have the potential to overcome treatment resistance in pancreatic ductal adenocarcinoma (PDAC), one of the least immunogenic solid tumors. Oncolytic viral chimera, CF33-hNIS-antiPDL1 genetically modified to express anti-human PD-L1 antibody and CF33-hNIS-Δ without the anti-PD-L1 gene, were used to investigate the immunogenic effects of OVs and virus-delivered anti-PD-L1 in PDAC in vitro. Western blot, flow cytometry, and immunofluorescence microscopy were used to evaluate the effects of CF33-hNIS-Δ and IFNγ on PD-L1 upregulation in AsPC-1 and BxPC-3 cells, and CF33-hNIS-antiPDL1 production of anti-PD-L1 and surface PD-L1 blockade of AsPC-1 and BxPC-3 with or without cocultured activated T cells. The cytosolic and cell surface levels of PD-L1 in PDAC cell lines varied; only BxPC-3 showed high cell surface expression. Treatment of these cells with CF33-hNIS-Δ and IFNγ significantly upregulated PD-L1 expression and translocation of PD-L1 from the cytosol onto the cell surface. Following coculture of activated T cells and BxPC-3 with CF33-hNIS-antiPDL1, the cell surface PD-L1 blockade on BxPC-3 cells by virus-delivered anti-PD-L1 antibody increased granzyme B release and prevented virus-induced decrease of perforin release from activated CD8+ T cells. Our results suggest that CF33-IOVs can prime immune checkpoint inhibition of PDAC and enhance antitumor immune killing.