Activity In Pancreatic Cancer Research Articles

Abstract 1319: Complement activation in pancreatic ductal adenocarcinoma

Abstract The complement system is an innate immune cascade which recognizes and clears microbes and damaged host cells. Complement activation has been linked to the promotion of tumor growth and metastasis across various anatomical sites with context-dependent effects. We therefore aim to understand the role for complement activation in pancreatic cancer. We have generated a tamoxifen-inducible autochthonous mouse model of PDAC which recapitulates genetic mutations found in humans using LSL-KrasG12D/+; Tgfbr2fl/fl; Ptf1a-CreER (KTC) mice. Using these mice, we assessed for complement activation in pancreatic tumors by western blot and confocal immunofluorescence. Western blotting for neoepitopes generated upon cleavage of Complement Component 3 (C3) demonstrated significantly increased complement activation in KTC tumors when compared to normal pancreas from at least 4-7 months post-induction. Using confocal immunofluorescence, we observed deposition of complement fragments in tumor stroma and on ductal carcinoma cells identified by co-staining with cytokeratin 19. We therefore hypothesized complement blockade would reduce PDAC tumor growth and sought to test this using C3 knockout (C3 KO) mice which lack all downstream complement function due to the central role of C3. We assessed flank tumor growth using two Kras; Tp53 (KPC) mutant-derived tumor cell lines in wild type (WT) C57BL/6J and C3 KO mice yet did not find a biologically significant reduction in flank tumor growth between WT and C3 KO. To validate the knockout, we assessed C3 activation by western blot but surprisingly found increased complement activation in C3 KO tumors. RT-qPCR demonstrated local transcription of C3 mRNA in both WT and C3 KO tumors as well as transcription of C3 mRNA in KPC cells in culture. This demonstrates that KPC cells are capable of autocrine production of C3. Using immunocytochemistry and RT-qPCR we identified expression of the C3a receptor in KPC cells which we hypothesized could serve as a sensor for feedback to control local C3 production. We applied 10 nM recombinant C3a to KPCY cells in vitro and found a 20% reduction in C3 mRNA transcription 1-hour post-treatment. Pre-treatment with 500 nM C3aR Antagonist (SB290157) ablated this effect. In summary, complement activation is significantly increased in PDAC and appears to be activated in part by tumor cells themselves. Autocrine production of C3 by tumor cells is sufficient to produce complement activation in the absence of systemic C3. Additional experiments are warranted to determine the role of complement in PDAC using C3 mice crossed with KTC mice to assess tumorigenesis in the total absence of C3. Citation Format: Brett I. Bell, Sanjay Pandey, Patrik Asp, Chandan Guha. Complement activation in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1319.

Abstract 3191: Tumor intrinsic HuR promotes stroma activation in pancreatic cancer

Abstract Background: One of the factors contributing to poor prognosis of pancreatic ductal adenocarcinoma (PDAC) is thought to be the characteristically dense stroma and heterogeneous tumor microenvironment. The PDAC stroma involves multiple players, including cancer associated fibroblasts (CAFs) and pancreatic stellate cells (PSCs). These stromal cells, when activated, promote extensive fibrosis in PDAC, which is associated with poor prognosis and chemoresistance. We have previously established that the RNA-binding protein Human Antigen R (HuR, ELAVL1) facilitates PDAC progression. In a tumor microarray of matched normal and malignant patient tissues, we found high active (i.e. cytoplasmic) HuR staining in 80% of PDAC tumors, while low or absent in adjacent normal tissues (n=80). Moreover, in both the orthotopic and the genetically engineered mouse models of PDAC, active HuR staining in tumors was dramatically higher than in normal pancreas. Pancreas-specific overexpression of HuR in a transgenic mouse model did not initiate tumorigenesis, but did increase fibrosis compared to control mice. We hypothesize that tumor intrinsic HuR regulates transcripts that are needed for tumor cells to activate the surrounding stroma. Methods: MiaPaCa-2 or PANC-1 human PDAC cells, either HuR-proficient or CRISPR-mediated knocked-out, were used in an orthotopic model where cells were injected into the tail of the pancreas. Both NRG and athymic mice were used. Resulting tumors underwent immunohistochemistry, immunofluorescence, or RNA-sequencing. Secreted proteins from cells in 2D culture were analyzed using a cytokine array. Results: HuR knockout tumors showed a decrease in CAF and PSC markers (i.e. podoplanin, alpha-SMA, desmin; >55% decrease, p<0.05), as well as a decrease in collagen deposition (56% decrease, p=0.0004). Additionally, HuR expression in tumors positively correlated with collagen abundance (p= 0.0002). RNA sequencing of PDAC tumors at various times during tumor development found that gene expression of stroma-activating ligands was significantly downregulated in HuR knockout tumors (e.g. TGFb, CCL2, PDGFA; -0.8<log2FC<-3.6, 0.0009<padj<2^-57). Moreover, we found a significant decrease of these stroma-activating ligands in conditioned media obtained from HuR knockout compared to wild type (e.g. PDGFAA, PTX3, CCL2; >50% decrease, p<0.0001). Conclusions: We found inhibition of HuR led to a robust decrease in stroma activation (i.e. lack of collagen, CAFs, and PSCs). To our knowledge, this is the first report of PDAC tumor intrinsic HuR having an extrinsic effect on neighboring stromal cells. Specifically, we found loss of HuR results in a decrease of secreted stroma-activating ligands, likely to be the cause of the decreased fibrosis in HuR knockout tumors. With evidence that fibrosis is associated with poor survival rates and chemoresistance, inhibiting HuR may sensitize tumors to standard of care therapy. Citation Format: Grace A. McCarthy, Jennifer M. Finan, Aditi Jain, Roberto DiNiro, Aaron Grossberg, Jonathan R. Brody. Tumor intrinsic HuR promotes stroma activation in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3191.

Phase 1/2 study of elraglusib (9-ING-41), a small molecule selective glycogen synthase kinase-3 beta (GSK-3β) inhibitor, alone or with irinotecan, temozolomide/irinotecan or cyclophosphamide/topotecan in pediatric patients with refractory malignancies: Interim results.

e22015 Background: GSK-3β overexpression is associated with aggressive malignancies, treatment resistance and poor prognosis. The GSK-3β inhibitor Elraglusib induces apoptosis via NFΚB and p53 pathways and has potent anti-fibrotic and immunomodulatory activity. Adult studies of elraglusib demonstrate clinical activity in pancreatic cancer, melanoma, lymphoma and sarcoma as a single agent or in combination with cytotoxic chemotherapy. Elraglusib is active in in vivo models of neuroblastoma (NBL) and malignant glioma. This first-in-pediatrics study (NCT04239092) is evaluating the safety, pharmacokinetics (PK), and efficacy of elraglusib monotherapy and in combination with chemotherapy in patients with refractory malignancies. Methods: Elraglusib is given intravenously (IV) twice-weekly at 3 dose levels (DL) (9.3, 12.4 and 15 mg/kg) as a single agent or in combination with irinotecan, cyclophosphamide/topotecan or temozolomide/irinotecan in 21-day cycles. A cohort of pts with refractory NBL will be treated at the recommended phase 2 dose (RP2D) of elraglusib with temozolomide/irinotecan. Results: As of January 2022, 23 pts (n = 7 female, median age 14.2 years) have received at least one dose of elraglusib. Tumor types: 5 NBL, 3 diffuse midline glioma (DMG), 3 osteosarcoma (OS), 3 ependymoma (EP), 2 alveolar rhabdomyosarcoma (aRMS), 1 angiosarcoma (AS), 1 Ewing sarcoma (ES), 1 glioblastoma (GBM), 1 hepatoblastoma (HB), 1 embryonal CNS tumor NOS, 1 NUT midline carcinoma, 1 pineoblastoma (PB). Median time from diagnosis is 26 months (range: 6.7 – 156.3) and median number of lines of prior systemic therapy is 2 (range 0-14). Two DLs of single agent (6 pts) have been completed (9.3 and 12.4 mg/kg) without elraglusib-attributable severe adverse events (SAEs). Of the 15 patients on the combination arm with irinotecan or cyclophosphamide/topotecan, a single adverse event (Grade 4 hypotension/infusion reaction) was reported. Grade 1/2 elraglusib attributable-AEs include: transient visual change (n = 10), nausea (n = 7), vomiting (n = 6), fatigue (n = 2), hypotension (n = 2) and infusion reaction (n = 1). One pt with recurrent ES had a radiographic and pathologic CR after 3 cycles of elraglusib/cyclophosphamide/topotecan. 6 pts (26.1%) had SD (2 NBL, 1 aRMS, 1 EP, 1 OS, 1 GBM). 8 pts (35%) remained on study treatment ≥ 3 months (2 NBL, 2 EP, 1 OS, 1 aRMS, 1 ES, 1 PB). Median treatment duration was 40 days (range 1 - 126). 4 pts remain on therapy. Conclusions: Elraglusib is well tolerated as a single agent and with several chemotherapy regimens in this heavily pretreated pediatric population with refractory cancers. It has encouraging antitumor activity, with 1 CR in a patient with recurrent ES. Enrollment is ongoing; a RPD2 has not been reached. Clinical trial information: NCT04239092.

The HDL particle composition determines its antitumor activity in pancreatic cancer.

Despite enormous efforts to improve therapeutic options, pancreatic cancer remains a fatal disease and is expected to become the second leading cause of cancer-related deaths in the next decade. Previous research identified lipid metabolic pathways to be highly enriched in pancreatic ductal adenocarcinoma (PDAC) cells. Thereby, cholesterol uptake and synthesis promotes growth advantage to and chemotherapy resistance for PDAC tumor cells. Here, we demonstrate that high-density lipoprotein (HDL)-mediated efficient cholesterol removal from cancer cells results in PDAC cell growth reduction and induction of apoptosis in vitro. This effect is driven by an HDL particle composition-dependent interaction with SR-B1 and ABCA1 on cancer cells. AAV-mediated overexpression of APOA1 and rHDL injections decreased PDAC tumor development in vivo. Interestingly, plasma samples from pancreatic-cancer patients displayed a significantly reduced APOA1-to-SAA1 ratio and a reduced cholesterol efflux capacity compared with healthy donors. We conclude that efficient, HDL-mediated cholesterol depletion represents an interesting strategy to interfere with the aggressive growth characteristics of PDAC.

Sotorasib Demonstrates Clinically Meaningful Activity in Pancreatic Cancer

Sotorasib Demonstrates Clinically Meaningful Activity in Pancreatic Cancer

Genetics and Therapeutic Responses to Tumor-Infiltrating Lymphocyte Therapy of Pancreatic Cancer Patient-Derived Xenograft Models.

Pancreatic cancer is the seventh leading cause of cancer-related deaths worldwide. Checkpoint immunotherapy has not yet shown encouraging results in pancreatic cancer possibly because of a poor immunogenicity and/or an immune suppressive microenvironment. The aim of this study was to develop patient-derived xenograft (PDX) models, compare their genetics to the original biopsies, and assess if autologous tumor-infiltrating lymphocytes (TILs) would have antitumoral activity in pancreatic cancer. We subcutaneously transplanted tumors from 29 patients into NOG mice to generate PDX models. We established TIL cultures and injected them into PDX mice. We analyzed histology and genetics of biopsies and PDX tumors. Tumor growths were confirmed in 11 of 29 transplantations. The PDX tumors histologically resembled their original biopsies, but because stromal cells in the PDX model tumors were from mouse, their gene expression differed from the original biopsies. Immune checkpoint ligands other than programmed death ligand-1 (PD-L1) were expressed in pancreatic cancers, but PD-L1 was rarely expressed. When it was expressed, it correlated with tumor take in PDX models. One of the 3 tumors that expressed PD-L1 was an adenosquamous cancer, and another had a mismatch repair deficiency. TILs were expanded from 6 tumors and were injected into NOG or human interleukin-2 transgenic-NOG mice carrying PDX tumors. Regression of tumors could be verified in human interleukin-2 transgenic-NOG mice in 3 of the 6 PDX models treated with autologous TILs, including the adenosquamous PDX model. PDX models of pancreatic cancer can be used to learn more about tumor characteristics and biomarkers and to evaluate responses to adoptive cell therapy and combination therapies. The major benefit of the model is that modifications of T cells can be tested in an autologous humanized mouse model to gain preclinical data to support the initiation of a clinical trial.

Therapeutic Targeting of Autophagy in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease characterized by early metastasis, late detection, and poor prognosis. Progress towards effective therapy has been slow despite significant efforts. Novel treatment approaches are desperately needed and autophagy, an evolutionary conserved process through which proteins and organelles are recycled for use as alternative energy sources, may represent one such target. Although incompletely understood, there is growing evidence suggesting that autophagy may play a role in PDAC carcinogenesis, metastasis, and survival. Early clinical trials involving autophagy inhibiting agents, either alone or in combination with chemotherapy, have been disappointing. Recently, evidence has demonstrated synergy between the MAPK pathway and autophagy inhibitors in PDAC, suggesting a promising therapeutic intervention. In addition, novel agents, such as ONC212, have preclinical activity in pancreatic cancer, in part through autophagy inhibition. We discuss autophagy in PDAC tumorigenesis, metabolism, modulation of the immune response, and preclinical and clinical data with selected autophagy modulators as therapeutics.

Promoter Hypermethylation Promotes the Binding of Transcription Factor NFATc1, Triggering Oncogenic Gene Activation in Pancreatic Cancer.

Simple SummaryHigh promoter methylation is not necessarily associated with shutting down gene activity but does sometimes correlate with increased transcription instead. In a genome-wide analysis, we studied gene regulation in pancreatic cancer. We identified a substantial number of genes with both promoter hypermethylation and high transcription levels. Subsequently, we screened for transcription factors that exhibit specific binding to such hypermethylated sequences. NFATc1 was one of several transcription factors that bound specifically methylated DNA motifs and triggered transcription. A particularly affected gene was ALDH1H3, whose expression has strong oncogenic implications. Activation of ALDH1H3 was due to a direct regulative process involving NFATc1 binding to its hypermethylated promoter. The results provide insights into the activation of gene transcription that is promoted by DNA methylation.Studies have indicated that some genes involved in carcinogenesis are highly methylated in their promoter regions but nevertheless strongly transcribed. It has been proposed that transcription factors could bind specifically to methylated promoters and trigger transcription. We looked at this rather comprehensively for pancreatic ductal adenocarcinoma (PDAC) and studied some cases in more detail. Some 2% of regulated genes in PDAC exhibited higher transcription coupled to promoter hypermethylation in comparison to healthy tissue. Screening 661 transcription factors, several were found to bind specifically to methylated promoters, in particular molecules of the NFAT family. One of them—NFATc1—was substantially more strongly expressed in PDAC than control tissue and exhibited a strong oncogenic role. Functional studies combined with computational analyses allowed determining affected genes. A prominent one was gene ALDH1A3, which accelerates PDAC metastasis and correlates with a bad prognosis. Further studies confirmed the direct up-regulation of ALDH1A3 transcription by NFATc1 promoter binding in a methylation-dependent process, providing insights into the oncogenic role of transcription activation in PDAC that is promoted by DNA methylation.

Identification of Novel Molecular Therapeutic Targets and Their Potential Prognostic Biomarkers Among Kinesin Superfamily of Proteins in Pancreatic Ductal Adenocarcinoma.

BackgroundKinesin superfamily of proteins (KIFs) has been broadly reported to play an indispensable role in the biological process. Recently, emerging evidence reveals its oncogenic role in various cancers. However, the prognostic, oncological, and immunological values of KIFs have not been comprehensively explored in pancreatic ductal adenocarcinoma (PDAC) patients. We aimed to illustrate the relationship between KIFs and pancreatic ductal adenocarcinoma by using bioinformatical analysis.MethodsWe use GEPIA, Oncomine datasets, cBioPortal, LOGpc, TIMER, and STRING bioinformatics tools and web servers to investigate the aberrant expression, prognostic values, and oncogenic role of KIFs. The two-gene prognostic model and the correlation between KIFs and KRAS and TP53 mutation were performed using an R-based computational framework.ResultsOur results demonstrated that KIFC1/2C/4A/11/14/15/18A/18B/20B/23 (we name it prognosis-related KIFs) were upregulated and associated with unfavorable clinical outcome in pancreatic cancer patients. KIF21B overexpression is associated with better clinical outcome. The KIFC1/2C/4A/11/14/15/18A/18B/20B/23 profiles were significantly increased compared to grade 1 and grade 2/3. Besides, KIFC1/2C/4A/11/14/15/18A/18B/20B/23 was significantly associated with the mutation status of KRAS and TP53.Notably, most prognosis-related KIFs have strong correlations with tumor growth and myeloid-derived suppressor cells infiltration (MDSCs). A prognostic signature based on KIF20B and KIF21B showed a reliable predictive performance. Receiver operating characteristic (ROC) curve was employed to assess the predictive power of two-gene signature. Consequently, the gene set enrichment analysis (GSEA) showed that KIF20B and KIF21B’s overexpression was associated with the immunological and oncogenic pathway activation in pancreatic cancer. Finally, real-time quantitative PCR (RT-qPCR) was utilized to investigate the expression pattern of KIF20B and KIF21B in pancreatic cancer cell lines and normal pancreatic cell.ConclusionsKnowledge of the expression level of the KIFs may provide novel therapeutic molecular targets and potential prognostic biomarkers to pancreatic cancer patients.

Immunoregulation related gene of CSMD2 and RNA activity as potential predicter of 2-year survival in pancreatic cancer: A DNA and RNA sequencing analysis.

e16212 Background: The 5-year survival rate of pancreatic cancer remains as low as 3%-15%. One of the key approaches to enrich current treatment options or improve effectiveness is new biomarker probing. We conducted DNA and RNA sequencing analysis to reveal potential biomarkers related to overall survival. Methods: Whole-exome sequencing, RNA sequencing and clinical data for 209 patients with pancreatic cancer were downloaded from TCGA. Clinical factors and mutational landscape (insertion/ deletion/ single nucleotide variant) were compared between group of OS2+ (OS longer than 2 years) and OS2- (OS longer than 2 years) with T test and Chi-square Test. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were conducted with RNA sequencing data to clarify the functional differences between the two groups. Results: The rates of OS2+ for patients in stage of I/II/III/IV was 43% (9/21), 17.8% (27/152), 0% (0/4), 0% (0/5), respectively. 152 patients in stage II were included for further analysis. No difference of sex and age were found between group of OS2+ and OS2-. Tumor mutation burden was comparable between the two groups. Mutation landscape showed the two groups had the accordance of 50% in top 10 genes. Mutations of CSMD2(18.5% vs. 5.0% , P = 0.026), CMYA5(14.8% vs, 2.5% , P = 0.019) and KCNA6(14.8% vs, 3.3%, P = 0.034) were more frequent in OS2+ group. CSMD2 is thought to be involved in the control of complement cascade of the immune system, and its low expression was significantly associated with differentiation, lymphatic invasion, and tumor size in colorectal cancer. CMYA5 was predicted as novel oncogene in breast cancer with the tool of Moonlight, it may also participate tumor activity in pancreatic cancer. The role of KCNA6 in cancer cell activity is barely known yet. Evaluation of differentially expressed genes between the two groups detected difference in leukocyte differentiation and T cell activation (GO analysis) and MAPK signal pathway (KEGG panalysis), these immunoregulation and MAPK pathways may play critical roles in tumor development and progression and affect the prognosis of pancreatic cancer. Conclusions: Pancreatic cancer with 2-year survival presented significant different DNA and RNA alterations, in which CSMD2 and pathway of leukocyte differentiation and T cell activation are closely associated with immunoregulation. These might provide guidance for prognose management and development of new therapeutic targets. Further mechanistic insights and prospective validation studies are warranted.

Mild hyperthermia promotes immune checkpoint blockade-based immunotherapy against metastatic pancreatic cancer using size-adjustable nanoparticles

Immune checkpoint blockade treatment is one of the most promising immunotherapies, which exhibits promising therapeutic effects on inhibition of metastasis. However, immunotherapy has little effect on pancreatic cancer, due to its extensive fibrotic matrix and immunosuppressive tumor microenvironment. Mild hyperthermia induced by photothermal therapy (PTT) has been proven to activate the immune responses in the tumor microenvironment. Herein, we designed a combine strategy of mild hyperthermia and immune checkpoint blockade (BMS-202) treatment with size-adjustable thermo- and fibrotic matrix- sensitive liposomes (HSA-BMS@CAP-ILTSL), in which BMS-202 loaded small-sized albumin nanoparticle (HSA-BMS) was encapsulated. Mild hyperthermia reduced the tumor hypoxia, relieved the interstitial pressure and increased the recruitment of endogenous immune cells in tumors. In the meantime, small-sized HSA-BMS was released from large-sized HSA-BMS@CAP-ILTSL in response to fibroblast activation protein-α (FAP-α) and near-infrared (NIR) laser, and enhanced the immunological responses by recovering the activity of T lymphocytes, accompanied by secreting relevant cytokines (TNF-α and IFN-γ). The combined therapy (HSA-BMS@CAP-ILTSL) could not only significantly suppress the tumor growth in vivo, but also decrease the amounts of metastatic nodules in distant organs. These results suggested that size-adjustable nanoparticles had a great potential in the treatment of metastatic pancreatic cancer. Statement of significanceThe desmoplastic stroma and hypoperfusion of pancreatic cancer imposed physical barriers to effective therapies, including chemotherapy, radiotherapy, targeted therapy, and immunotherapy. We constructed size-adjustable thermo- and fibrotic matrix- sensitive liposomes (HSA-BMS@CAP-ILTSL) with size around 120 nm, where small sized albumin nanoparticle (10 nm) of immune checkpoint inhibitor (HSA-BMS) were encapsulated inside. Mild hyperthermia not only contributed to release HSA-BMS for penetration (blocking the immunosuppressive signals deep in the tumor), but enhanced tumor blood perfusion for infiltration of endogenous immune cells. In the two-pronged treatment, the pancreatic cancer immunotherapy significantly enhanced and the risk of cancer metastasis was reduced. Overall, the strategy provides a promising approach to increase drug accumulation and improve the anti-tumor immune activity in pancreatic cancer.

The natural product trienomycin A is a STAT3 pathway inhibitor that exhibits potent in vitro and in vivo efficacy against pancreatic cancer.

Pancreatic cancer is an exceptionally fatal disease. However, therapeutic drugs for pancreatic cancer have presented a serious shortage over the past few decades. Signal transducer and activator of transcription-3 (STAT3) is persistently activated in many human cancers where it promotes tumour development and progression. Natural products serve as an inexhaustible source of anticancer drugs. Here, we identified the natural product trienomycin A (TA), an ansamycin antibiotic, as a potential inhibitor of the STAT3 pathway with potent activity against pancreatic cancer. Effects of trienomycin A on transcriptional activity of STAT3 were assessed by the STAT3-luciferase (STAT3-luc) reporter system. In vitro and in vivo inhibitory activity of TA against pancreatic cancer made use of molecular docking, surface plasmon resonance (SPR) assay, MTS assay, colony formation assay, transwell migration/invasion assay, flow cytometric analysis, immunofluorescence staining, quantitative real-time polymerase chain reaction (PCR), western blotting, tumour xenograft model, haematoxylin and eosin (H&E) staining and immunohistochemistry. Trienomycin A directly bound to STAT3 and inhibited STAT3 (Tyr705) phosphorylation, thus inhibiting the STAT3 pathway. Trienomycin A also inhibited colony formation, proliferation, migration and invasion of pancreatic cancer cell lines. Trienomycin A also markedly blocked pancreatic tumour growth in vivo. More importantly, trienomycin A did not show obvious toxicity at the effective dose in mice. Trienomycin A exerted anti-neoplastic activity by suppressing STAT3 activation in pancreatic cancer. This natural product could be a novel therapeutic candidate for pancreatic cancer.

Invadopodia: A potential target for pancreatic cancer therapy.

Dissemination of cancer cells is an intricate multistep process that represents the most deadly aspect of cancer. Cancer cells form F-actin-rich protrusions known as invadopodia to invade surrounding tissues, blood vessels and lymphatics. A number of studies have demonstrated the significant roles of invadopodia in cancer. Therefore, the specific cells and molecules involved in invadopodia activity can provide as therapeutic targets. In this review, we included a thorough overview of studies in invadopodia and discussed their functions in cancer metastasis. We then presented the specific cells and molecules involved in invadopodia activity in pancreatic cancer and analyzed their suitability to be effective therapeutic targets. Currently, drugs targeting invadopodia and relevant clinical trials are negligible. Here, we highlighted the significance of potential drugs and discussed future obstacles in implementing clinical trials. This review presents a new perspective on invadopodia-induced pancreatic cancer metastasis and may prosper the development of targeted therapeutics against pancreatic cancer.

Combining NanoKnife with M1 oncolytic virus enhances anticancer activity in pancreatic cancer.

NanoKnife, a nonthermal ablation technique also termed irreversible electroporation (IRE), has been adopted in locally advanced pancreatic cancer (LAPC) treatment. However, reversible electroporation (RE) caused by heterogeneous electric field magnitude leads to inadequate ablation and tumor recurrence. Alphavirus M1 has been identified as a novel natural oncolytic virus which is nonpathogenic and with high tumor selectivity. This study evaluated improvements to therapeutic efficacy through combination therapy incorporating NanoKnife and M1 virus. We showed that IRE triggered reactive oxygen species (ROS)-dependent apoptosis in pancreatic cancer cells (PCCs) mediated by phosphatidylinositol-3-kinase (PI3K)/protein kinase B (Akt) pathway suppression. When NanoKnife was combined with M1 virus, the therapeutic efficacy was synergistically enhanced. The combinatorial treatment further inhibited tumor proliferation and prolonged the survival of orthotopic pancreatic cancer (PC)-bearing immunocompetent mice. In depth, NanoKnife enhanced the oncolytic effect of M1 by promoting its infection. The combination turned immune-silent tumors into immune-inflamed tumors characterized by T cell activation. Clinicopathologic analysis of specific M1 oncolytic biomarkers indicated the potential of the combination regimen. The combinatorial therapy represents a promising therapeutic efficacy and may ultimately improve the prognosis of patients with LAPC.

Kinome Array Profiling of Patient-Derived Pancreatic Ductal Adenocarcinoma Identifies Differentially Active Protein Tyrosine Kinases.

Pancreatic cancer remains one of the most difficult malignancies to treat. Minimal improvements in patient outcomes and persistently abysmal patient survival rates underscore the great need for new treatment strategies. Currently, there is intense interest in therapeutic strategies that target tyrosine protein kinases. Here, we employed kinome arrays and bioinformatic pipelines capable of identifying differentially active protein tyrosine kinases in different patient-derived pancreatic ductal adenocarcinoma (PDAC) cell lines and wild-type pancreatic tissue to investigate the unique kinomic networks of PDAC samples and posit novel target kinases for pancreatic cancer therapy. Consistent with previously described reports, the resultant peptide-based kinome array profiles identified increased protein tyrosine kinase activity in pancreatic cancer for the following kinases: epidermal growth factor receptor (EGFR), fms related receptor tyrosine kinase 4/vascular endothelial growth factor receptor 3 (FLT4/VEGFR-3), insulin receptor (INSR), ephrin receptor A2 (EPHA2), platelet derived growth factor receptor alpha (PDGFRA), SRC proto-oncogene kinase (SRC), and tyrosine kinase non receptor 2 (TNK2). Furthermore, this study identified increased activity for protein tyrosine kinases with limited prior evidence of differential activity in pancreatic cancer. These protein tyrosine kinases include B lymphoid kinase (BLK), Fyn-related kinase (FRK), Lck/Yes-related novel kinase (LYN), FYN proto-oncogene kinase (FYN), lymphocyte cell-specific kinase (LCK), tec protein kinase (TEC), hemopoietic cell kinase (HCK), ABL proto-oncogene 2 kinase (ABL2), discoidin domain receptor 1 kinase (DDR1), and ephrin receptor A8 kinase (EPHA8). Together, these results support the utility of peptide array kinomic analyses in the generation of potential candidate kinases for future pancreatic cancer therapeutic development.

1550P Combination therapy of low molecular weight heparins, chemotherapy and immunotherapy induce antitumor activity in pancreatic cancer

1550P Combination therapy of low molecular weight heparins, chemotherapy and immunotherapy induce antitumor activity in pancreatic cancer

Exosomes-Coated miR-34a Displays Potent Antitumor Activity in Pancreatic Cancer Both in vitro and in vivo.

PurposeMiR-34a, which acts as an important tumor suppressor gene, plays an important role in pancreatic cancer. However, the therapeutic application of miR-34a is limited by the lack of an effective delivery system. In the present study, we synthesize exosomes-coated miR-34a (exomiR-34a), and the anticancer effect of exomiR-34a was evaluated in pancreatic cancer.Materials and MethodsAn ultrasound approach was used to synthesize exomiR-34a, and its transfection efficiency was examined by confocal microscopy and flow cytometry. The level of miR-34a and its targeted gene Bcl-2 was detected by real-time quantitative PCR (qRT-PCR). MTT analysis was performed to determine the effect of exomiR-34a on the growth of pancreatic cancer cells. Annexin-V/PI double staining and Western blot analysis were carried out to determine the apoptosis of the pancreatic cancer cells. The xenograft nude mice model bearing human pancreatic cancer Panc28 cells was used to determine the antitumor effect of exomiR-34a in vivo.ResultsThe exomiR-34a could cross the cell membrane efficiently, and downregulated the expression of the targeted gene Bcl-2. Treatment with exomiR-34a inhibited the growth of the pancreatic cancer cells significantly and the nanoparticles also induced apoptosis in cancer cells via affecting the expression of apoptotic-related genes. In vivo study using xenograft nude mice bearing Panc28 cancer cells revealed that exomiR-34a suppressed the growth of tumors significantly.ConclusionExomiR-34a can inhibit the growth of pancreatic cancer both in vitro and in vivo. Targeting miR-34a is a promising strategy for the treatment of pancreatic cancer. ExomiR-34a has the potential to be developed as a novel anticancer agent for the treatment of human pancreatic malignancy.

Bptf determines oncogenic addiction in aggressive B-cell lymphomas.

Chromatin remodeling factors contribute to establish aberrant gene expression programs in cancer cells and therefore represent valuable targets for therapeutic intervention. BPTF (Bromodomain PhD Transcription Factor), a core subunit of the nucleosome remodeling factor (NURF), modulates c-MYC oncogenic activity in pancreatic cancer. Here, we analyze the role of BPTF in c-MYC-driven B-cell lymphomagenesis using the Eμ-Myc transgenic mouse model of aggressive B-cell lymphoma. We find that BPTF is required for normal B-cell differentiation without evidence of haploinsufficiency. In contrast, deletion of one Bptf allele is sufficient to delay lymphomagenesis in Eμ-Myc mice. Tumors arising in a Bptf heterozygous background display decreased c-MYC levels and pathway activity, together with increased activation of the NF-κB pathway, a molecular signature characteristic of human diffuse large B-cell lymphoma (DLBCL). In human B-cell lymphoma samples, we find a strong correlation between BPTF and c-MYC mRNA and protein levels, together with an anti-correlation between BPTF and NF-κB pathway activity. Our results indicate that BPTF is a relevant therapeutic target in B-cell lymphomas and that, upon its inhibition, cells acquire distinct oncogenic dependencies.

Phosphatidylserine-exposing blood cells, microparticles and neutrophil extracellular traps increase procoagulant activity in patients with pancreatic cancer

Patients with pancreatic cancer (PC) are at increased risk of venous thrombosis, but the precise mechanisms of hypercoagulable state in PC remain unclear. We aimed to identify how phosphatidylserine positive (PS+) blood cells (BCs), PS+ microparticles (MPs) and neutrophil extracellular traps (NETs) regulate procoagulant activity (PCA) in PC, and to assess the relationship between PCA and PC staging. A total of 83 PC patients with different stages of disease were compared to 30 healthy controls, with confocal microscopy and flow cytometry used to assess MP and cellular PS exposure. MP and cell PCA was determined using both fibrin production assays and procoagulant enzyme complex analyses, and coagulation time was further measured. Patients with stage I PC and healthy controls exhibited significantly lower frequencies of PS+ MPs and BCs relative to those with more advanced disease, which may partly due to the increased levels of inflammation cytokines in advanced disease. Functional coagulation assays indicated that PS+ MPs and BCs derived from patients with stage II/III/IV PC directly contribute to elevated FXa, thrombin, and fibrin formation, and to more rapid coagulation relative to healthy control samples. In inhibition assays, lactadherin, which antagonizes PS, led to a roughly 80% inhibition of PCA. We further used isolated NETs to stimulate endothelial cells, revealing that this led to morphological changes including retraction from cell-cell junctions and a more pro-coagulative phenotype, with DNase I and activated protein C treatment reversing these changes. In patients with stage III PC, curative resection surgery significantly reduced PCA, whereas non-curative surgery did not have a marked impact based on studies of pre- and post-operative samples. These results highlight the pathogenic activity of PS+ cells, MPs, and NETs in promoting a prothrombotic environment within individuals suffering from advanced PC. Targeting PS and NETs in these patients may thus be a viable means of preventing pathological thrombosis.

Abstract A31: Investigating the effect of myeloid Arg1 deletion on tumor growth and CD8+ T-cell infiltration and activation in pancreatic cancer

Abstract PDA is characterized by an extensive fibroinflammatory stroma. This fibroinflammatory stroma is mainly composed of fibroblasts and tumor-infiltrating immune cells. The most abundant infiltrating immune cells are myeloid cells. Myeloid cells including tumor-associated macrophages (TAMs), myeloid-derived suppressor cells, and granulocytes are required for PDA tumor growth and maintenance. Myeloid cells have the ability to suppress antitumor T-cell responses in PDA, as depletion of myeloid cells restores CD8+ T-cell immunity. Our previous characterization of myeloid cells infiltrating the neoplastic pancreas has revealed that myeloid cells express high levels of Arginase 1 (Arg1), an enzyme that depletes the amino acid L-arginine from the microenvironment and a signature marker of immunosuppressive macrophages and TAMs. In turn, L-arginine is required for CD8+ T-cell activation. An increase in Arginase levels has been reported also in other cancers, including lung, gastrointestinal, and bladder cancer. Thus, Arg1 expression in myeloid cells might be a key mediator of immune suppression, although this possibility has not been investigated directly in pancreatic cancer. Based on these observations, we test the hypothesis that myeloid cell polarization in the tumor microenvironment mediates immune suppression in PDA through expression of Arginase 1. The objective of this study is to provide novel insights into the role of Arg1 in pancreatic cancer, and the overall goal is to identify new therapeutic targets for combination therapy in pancreatic cancer. We used a genetically engineered mouse model (LysM-Cre;Arg1f/f) to delete the Arg1 gene, specifically from the myeloid cell compartment (LysM+ cells), including macrophages and neutrophils. We orthotopically implanted primary mouse pancreatic cancer cell lines into C57BL/6 LysM-Cre;Arg1f/f and wild-type (WT) mice. We evaluated tumor growth and CD8+ T-cell infiltration and activation between LysM-Cre;Arg1f/f and WT control mice. We used MRI imaging to determine tumor volume, and we used immunofluorescence and mass cytometry analysis to investigate changes in immune cell infiltration. We confirmed Arg1 depletion in LysM-Cre;Arg1f/f mice by Western blotting, co-immunofluorescence, and mass cytometry. We observed a decreasing trend in tumor growth and volume in LysM-Cre;Arg1f/f mice, an increase in iNOS expression (a marker of inflammatory macrophages), and an increase in CD8+ T-cell number and activity compared to the WT. These results support the notion that Arg1 might be a moderator of immune suppression in pancreatic cancer. Citation Format: Rosa E. Menjivar, Christopher Halbrook, Ashley Velez, Fatima Lima, Carlos Espinoza, Stefanie Galban, Yaqing Zhang, Costas Lyssiotis, Marina Pasca di Magliano. Investigating the effect of myeloid Arg1 deletion on tumor growth and CD8+ T-cell infiltration and activation in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr A31.