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We described trends and disparities in utilization of systemic chemotherapy in metastatic hepato-pancreato-biliary (HPB) cancers.We queried the National Cancer Database for metastatic HPB cancers [hepatocellular carcinoma (HCC), biliary tract cancers (BTC), pancreatic adenocarcinoma (PDAC)]. We used multivariable analysis to examine the factors associated with utilization of systemic chemotherapy. We utilized marginal structural logistic models to estimate the effect of health insurance, facility type, or facility volume on utilization of systemic chemotherapy.We identified 162,283 patients with metastatic HPB cancers: 23,923 (14.7%) had HCC, 26,766 (16.5%) had BTC, and 111,594 (68.8%) had PDAC. A total of 37.2% patients with HCC, 55.6% with BTC, and 56.4% with PDAC received chemotherapy. Age ≥70 years and Charlson-Deyo score ≥2 were associated with lower likelihood of receiving chemotherapy across all cancers. Patients with private health insurance had higher receipt of chemotherapy. Receiving treatment at academic facilities had no effect on the receipt of chemotherapy. Treatment of patients with HCC or PDAC at high-volume facilities resulted in higher receipt of chemotherapy.A significant proportion of patients with metastatic HPB cancers do not receive systemic chemotherapy. Several disparities in administration of chemotherapy for metastatic HPB cancers exist.

To examine the average treatment effect of hepato-pancreato-biliary (HPB) cancer volume on survival outcomes of patients with non-resected pancreatic adenocarcinoma (PDAC).We queried the National Cancer Database (2004-2018) for patients with HPB malignancies (PDAC, pancreatic neuroendocrine neoplasms, hepatocellular carcinoma, biliary tract cancers). We determined the 25th, 50th, and 75th percentiles based on the total annual HPB volume. We then identified patients with non-resected PDAC. We utilized inverse probability (IP)-weighted Cox regression to estimate the effect of facility volume on overall survival (OS).We identified 710,988 patients with HPB malignancies. The 25th, 50th, and 75th percentiles of total annual HPB volume were 32, 71, and 177 cases/year, respectively. We included a total of 196,150 patients with non-resected PDAC. Patients treated at ≥25th, ≥50th, and ≥75th percentile facilities had improved median OS compared to those treated at facilities below these thresholds (5.8 vs. 4.2months, 6.5 vs. 4.5months, 7.5 vs. 4.8months, respectively; p 0.001 for all). Treatment at facilities ≥25th, ≥50th, and ≥75th percentile resulted in lower hazards of death than treatment at lower-percentile facilities (HR: 0.87, 95% CI: 0.84-0.90; HR: 0.87, 95% CI: 0.83-0.91; HR: 0.85, 95% CI: 0.79-0.91, respectively).Our data suggest that consolidation of care of patients with PDAC to high-volume centers may be beneficial even in the nonoperative setting.

Metabolites of tryptophan degradation are known to alter mood. Their effects have only been superficially examined in the context of pancreatic cancer. Herein, we study the role of indoleamine 2,3-dioxygenase 1 (IDO1), an enzyme important in the conversion of tryptophan to kynurenine, in a murine model of pancreatic cancer–associated depression. Behavioral tests (open field, forced swim, tail suspension, and elevated plus maze) and biochemical assays (LC-MS metabolomics) were used to characterize a depressive-phenotype in tumor-bearing mice (relative to non–tumor-bearing mice). In addition, we determine whether pharmacologic blockade of IDO1 affects mood in tumor-bearing mice. Immunocompetent mice bearing orthotopic pancreatic tumors exhibit depressive-like behavior relative to non–tumor-bearing mice. Pancreatic tumors strongly express IDO1. Consequently, serum kynurenine levels in tumor-bearing mice are elevated relative to non–tumor-bearing mice. Tumor-bearing mice treated with epacadostat, an IDO1 inhibitor, exhibited improved mood relative to mice receiving vehicle. There was a 95% reduction in serum kynurenine levels in mice receiving epacadostat relative to mice treated with vehicle. As confirmatory evidence of on-target activity, tumors of mice treated with epacadostat exhibited a compensatory increase in IDO1 protein levels. Escitalopram, an approved antidepressant, was ineffective at improving mood in tumor-bearing mice as measured by behavioral assays and did not affect kynurenine levels. Neither epacadostat, nor escitalopram, affected overall survival relative to vehicle. Mice with pancreatic cancer exhibit depressive-like behavior. Epacadostat was effective as an antidepressant for pancreatic cancer–associated depression in mice. These data offer a rationale to consider IDO1 inhibition as a therapeutic strategy to mitigate depressive symptoms in patients with pancreatic cancer.

Nutrient-deprived conditions in the tumor microenvironment (TME) restrain cancer cell viability due to increased free radicals and reduced energy production. In pancreatic cancer cells a cytosolic metabolic enzyme, wild-type isocitrate dehydrogenase 1 (wtIDH1), enables adaptation to these conditions. Under nutrient starvation, wtIDH1 oxidizes isocitrate to generate α-ketoglutarate (αKG) for anaplerosis and NADPH to support antioxidant defense. In this study, we show that allosteric inhibitors of mutant IDH1 (mIDH1) are potent wtIDH1 inhibitors under conditions present in the TME. We demonstrate that low magnesium levels facilitate allosteric inhibition of wtIDH1, which is lethal to cancer cells when nutrients are limited. Furthermore, the Food & Drug Administration (FDA)-approved mIDH1 inhibitor ivosidenib (AG-120) dramatically inhibited tumor growth in preclinical models of pancreatic cancer, highlighting this approach as a potential therapeutic strategy against wild-type IDH1 cancers.

Consensus guidelines recommend for perioperative chemotherapy and surgery for patients with clinical stage (cT) T2 or greater gastric adenocarcinoma. We compared adherence to guidelines in these patients stratified by race.Non-Hispanic White and Black patients with resected ≥cT2 gastric adenocarcinoma were identified within the National Cancer Database (2008-2017). We compared administration of preoperative chemotherapy by race, adjusting for clinicodemographic variables. We performed marginal standardization of logistic regression to calculate adjusted probabilities of administration of preoperative chemotherapy in patients under the age of 80 years with insurance.A total of 13,850 patients were identified (White = 12,161; Black = 1,689). Black race was associated with lower likelihood of receiving preoperative chemotherapy than White race (odds ratio = 0.46, 95% confidence interval: 0.39-0.54). Other factors associated with lower likelihood of preoperative chemotherapy included age ≥70 years, female sex, treatment at community facilities, non-private or no health insurance, and cT4 disease. Factors associated with higher likelihood of preoperative chemotherapy included treatment at high-volume facilities, longer distance to facility, higher education and income levels, cT3 disease, and cN+ disease. In patients80 years with insurance, marginal standardization models demonstrated that Black race was associated with a lower adjusted probability of receiving preoperative chemotherapy regardless of age, insurance payor, facility type/volume, distance to facility, cT stage, cN stage, sex, and education/income levels.Black race was associated with underutilization of preoperative chemotherapy for cT2 or greater gastric cancer, in discordance to published guidelines. The etiology of these disparities is multifactorial, and correcting the root causes represents a critical area for improvement.

Irreversible electroporation (IRE) is used as a locoregional treatment modality for patients with locally advanced pancreatic cancer (LAPC), but is non-curative and is associated with postoperative morbidity and mortality. We performed a systematic review and meta-analysis comparing survival outcomes of multimodal therapy with or without IRE.Separate searches were performed for multimodal therapy + IRE and multimodal therapy alone given the lack of comparative literature using PubMed, SCOPUS, and Cochrane Library in 3/2021. We determined overall survival (OS) and progression-free survival (PFS) from diagnosis and time of IRE. Treatment-related morbidity and mortality was determined.Of 585 published articles, 48 met inclusion criteria for IRE (n = 27) and without IRE (n = 21) with data for 1420 (IRE) and 1348 (without IRE) patients. The 6/12/24 months OS with IRE was 99%/84%/28%. The 6/12/24 months OS without IRE was 99%/80%/12%. At 12 months from IRE, OS was 55% and PFS was 12%. The mean major complication and 90-day mortality rates for IRE were 17.95% and 2.65%.Multimodal therapy alone is associated with similar OS to multimodal therapy + IRE in patients with LAPC. Most patients progress and nearly half die within 1 year of the IRE procedure. Given the lack of quality prospective data, IRE should remain experimental and be used with caution in LAPC.

Background Weight changes during neoadjuvant chemotherapy (NAC) for pancreatic cancer (PDAC) are not well studied. We hypothesized that weight loss may predict poor outcomes. Methods Weight change from NAC initiation to pancreatectomy was grouped: gain (≥5%), stable, and loss (≥5%). Pathologic, postoperative, and survival outcomes were compared. Results 95 patients were included: 31.6% lost weight, 58.9% maintained weight, and 9.5% gained weight. There were no differences in chemotherapeutic regimens. Median recurrence-free survival (RFS) and overall survival (OS) were similar between patients with stable weight and those who lost weight (RFS: 9.6vs14.0months; OS: 25.8vs26.7months). Among those who gained weight, RFS (29.5months) and OS (38.4months) were greater relative to the other weight categories. On multivariable regression, weight gain was associated with improved RFS compared to loss (HR = 0.16). Conclusion Most patients maintain or lose weight during NAC, and weight loss does not predict poor outcomes. Weight gain may predict improved RFS.

Supplementary Figure from IDO1 Is a Therapeutic Target for Pancreatic Cancer–Associated Depression

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Pancreatic ductal adenocarcinoma (PDA) is one of the most lethal cancers, in part, due to resistance to both conventional and targeted therapeutics. TRAIL directly induces apoptosis through engagement of cell surface Death Receptors (DR4 and DR5), and has been explored as a molecular target for cancer treatment. Clinical trials with recombinant TRAIL and DR-targeting agents, however, have failed to show overall positive outcomes. Herein, we identify a novel TRAIL resistance mechanism governed by Hu antigen R (HuR, ELAV1), a stress-response protein abundant and functional in PDA cells. Exogenous HuR overexpression in TRAIL-sensitive PDA cell lines increases TRAIL resistance whereas silencing HuR in TRAIL-resistant PDA cells, by siRNA oligo-transfection, decreases TRAIL resistance. PDA cell exposure to soluble TRAIL induces HuR translocation from the nucleus to the cytoplasm. Furthermore, it is demonstrated that HuR interacts with the 3′-untranslated region (UTR) of DR4 mRNA. Pre-treatment of PDA cells with MS-444 (Novartis), an established small molecule inhibitor of HuR, substantially increased DR4 and DR5 cell surface levels and enhanced TRAIL sensitivity, further validating HuR's role in affecting TRAIL apoptotic resistance. NanoString analyses on the transcriptome of TRAIL-exposed PDA cells identified global HuR-mediated increases in antiapoptotic processes. Taken together, these data extend HuR's role as a key regulator of TRAIL-induced apoptosis.Implications: Discovery of an important new HuR-mediated TRAIL resistance mechanism suggests that tumor-targeted HuR inhibition increases sensitivity to TRAIL-based therapeutics and supports their re-evaluation as an effective treatment for PDA patients. Mol Cancer Res; 14(7); 599–611. ©2016 AACR.

Figure S1: HuR inhibition was successfully achieved by CRISPR/cas9 system. Figure S2: HuR inhibition sensitizes cells to drugs. Figure S3: PDA xenografts with HuR inhibition failed to form tumors. Table S1: Lists the three guide RNAs used to generate CRISPR knockout of HuR in MIA PaCa-2 and Hs 766T cell lines. Table S2: Lists the primers used to detect the mutation created by all three guide RNAs in CRISPR knockout of HuR MIA PaCa-2 and Hs 766T cell lines. Table S3: Lists the clones along with their genotypes obtained for all three guide RNAs.

PDF file - 28K, S1. Cell growth in a high USP11 expression cell line (PL5) following siRNA treatment targeting USP11.

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Pancreatic ductal adenocarcinoma (PDA) is the third leading cause of cancer-related deaths in the United States, whereas colorectal cancer is the third most common cancer. The RNA-binding protein HuR (ELAVL1) supports a pro-oncogenic network in gastrointestinal (GI) cancer cells through enhanced HuR expression. Using a publically available database, HuR expression levels were determined to be increased in primary PDA and colorectal cancer tumor cohorts as compared with normal pancreas and colon tissues, respectively. CRISPR/Cas9 technology was successfully used to delete the HuR gene in both PDA (MIA PaCa-2 and Hs 766T) and colorectal cancer (HCT116) cell lines. HuR deficiency has a mild phenotype, in vitro, as HuR-deficient MIA PaCa-2 (MIA.HuR-KO(−/−)) cells had increased apoptosis when compared with isogenic wild-type (MIA.HuR-WT(+/+)) cells. Using this isogenic system, mRNAs were identified that specifically bound to HuR and were required for transforming a two-dimensional culture into three dimensional (i.e., organoids). Importantly, HuR-deficient MIA PaCa-2 and Hs 766T cells were unable to engraft tumors in vivo compared with control HuR-proficient cells, demonstrating a unique xenograft lethal phenotype. Although not as a dramatic phenotype, CRISPR knockout HuR HCT116 colon cancer cells (HCT.HuR-KO(−/−)) showed significantly reduced in vivo tumor growth compared with controls (HCT.HuR-WT(+/+)). Finally, HuR deletion affects KRAS activity and controls a subset of pro-oncogenic genes.Implications: The work reported here supports the notion that targeting HuR is a promising therapeutic strategy to treat GI malignancies. Mol Cancer Res; 15(6); 696–707. ©2017 AACR.

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Supplementary Fig S7 describes western blot and RTPCR of PARG in modified shPARG cells.

Patients with metastatic pancreatic ductal adenocarcinoma (PDAC) have an average survival of less than 1 year, underscoring the importance of evaluating novel targets with matched targeted agents. We recently identified that poly (ADP) ribose glycohydrolase (PARG) is a strong candidate target due to its dependence on the pro-oncogenic mRNA stability factor HuR (ELAVL1). Here, we evaluated PARG as a target in PDAC models using both genetic silencing of PARG and established small-molecule PARG inhibitors (PARGi), PDDX-01/04. Homologous repair–deficient cells compared with homologous repair–proficient cells were more sensitive to PARGi in vitro. In vivo, silencing of PARG significantly decreased tumor growth. PARGi synergized with DNA-damaging agents (i.e., oxaliplatin and 5-fluorouracil), but not with PARPi therapy. Mechanistically, combined PARGi and oxaliplatin treatment led to persistence of detrimental PARylation, increased expression of cleaved caspase-3, and increased γH2AX foci. In summary, these data validate PARG as a relevant target in PDAC and establish current therapies that synergize with PARGi.Significance:PARG is a potential target in pancreatic cancer as a single-agent anticancer therapy or in combination with current standard of care.

Flow cytometry graphs (black and white line and dot drawing)

Table S5: Results of RNA sequencing in PDA cell lines (HS-766T and MiaPaCa2) with each modulated by CRISPR/Cas9 to delete HuR expression.

Cancer aggressiveness may result from the selective pressure of a harsh nutrient-deprived microenvironment. Here we illustrate how such conditions promote chemotherapy resistance in pancreatic ductal adenocarcinoma (PDAC). Glucose or glutamine withdrawal resulted in a 5- to 10-fold protective effect with chemotherapy treatment. PDAC xenografts were less sensitive to gemcitabine in hypoglycemic mice compared with hyperglycemic mice. Consistent with this observation, patients receiving adjuvant gemcitabine (n = 107) with elevated serum glucose levels (HgbA1C > 6.5%) exhibited improved survival. We identified enhanced antioxidant defense as a driver of chemoresistance in this setting. ROS levels were doubled in vitro by either nutrient withdrawal or gemcitabine treatment, but depriving PDAC cells of nutrients before gemcitabine treatment attenuated this effect. Mechanistic investigations based on RNAi or CRISPR approaches implicated the RNA binding protein HuR in preserving survival under nutrient withdrawal, with or without gemcitabine. Notably, RNA deep sequencing and functional analyses in HuR-deficient PDAC cell lines identified isocitrate dehydrogenase 1 (IDH1) as the sole antioxidant enzyme under HuR regulation. HuR-deficient PDAC cells lacked the ability to engraft successfully in immunocompromised mice, but IDH1 overexpression in these cells was sufficient to fully restore chemoresistance under low nutrient conditions. Overall, our findings highlight the HuR–IDH1 regulatory axis as a critical, actionable therapeutic target in pancreatic cancer. Cancer Res; 77(16); 4460–71. ©2017 AACR.

PARG rescue in HuR silenced cells restores PARPi resistance. Small-molecule inhibition of HuR prevents its cytoplasmic accumulation, abrogates PARG mRNA regulation and increases PARPi sensitivity, independent of DDR status.

Doxycycline induction of HuR silencing in MIA PaCa-2 cells (sh290) downregulates PARG expression in vitro and tumor growth in vivo.

HuR silencing results in downregulation of PARG expression, which prevents efficient removal of PAR polymers with or without PARPi stress. Expression and function of other PAR removing enzymes is not affected with HuR silencing.

HuR directly binds and regulates poly-ADP ribose glycohydrolase mRNA expression particularly under PARPi stress, but doesn't affect expression of other PAR polymerases or hydrolyzing enzymes.

Purpose:Heritable genetic variations can affect the inflammatory tumor microenvironment, which can ultimately affect cancer susceptibility and clinical outcomes. Recent evidence indicates that IDO2, a positive modifier in inflammatory disease models, is frequently upregulated in pancreatic ductal adenocarcinoma (PDAC). A unique feature of IDO2 in humans is the high prevalence of two inactivating single-nucleotide polymorphisms (SNP), which affords the opportunity to carry out loss-of-function studies directly in humans. In this study, we sought to address whether genetic loss of IDO2 may influence PDAC development and responsiveness to treatment.Experimental Design: Transgenic Ido2+/+ and Ido2−/− mice in which oncogenic KRAS is activated in pancreatic epithelial cells were evaluated for PDAC. Two patient data sets (N = 200) were evaluated for the two IDO2-inactivating SNPs together with histologic, RNA expression, and clinical survival data.Results:PDAC development was notably decreased in the Ido2−/− mice (30% vs. 10%, P < 0.05), with a female predominance similar to the association observed for one of the human SNPs. In patients, the biallelic occurrence of either of the two IDO2-inactivating SNPs was significantly associated with markedly improved disease-free survival in response to adjuvant radiotherapy (P < 0.01), a treatment modality that has been highly debated due to its variable efficacy.Conclusions:The results of this study provide genetic support for IDO2 as a contributing factor in PDAC development and argue that IDO2 genotype analysis has the immediate potential to influence the PDAC care decision-making process through stratification of those patients who stand to benefit from adjuvant radiotherapy.

HuR expression regulates short-term cell survival and long-term anchorage-independent growth in PDA cells, irrespective of DDR proficiency. PARPi stress induces HuR translocation in a time-dependent manner.

Supplemental Methods and Supplemental Figure Legends

Supplementary.Figures 1-8

The majority of pancreatic ductal adenocarcinomas (PDAC) rely on the mRNA stability factor HuR (ELAV-L1) to drive cancer growth and progression. Here, we show that CRISPR-Cas9–mediated silencing of the HuR locus increases the relative sensitivity of PDAC cells to PARP inhibitors (PARPi). PDAC cells treated with PARPi stimulated translocation of HuR from the nucleus to the cytoplasm, specifically promoting stabilization of a new target, poly (ADP-ribose) glycohydrolase (PARG) mRNA, by binding a unique sequence embedded in its 3′ untranslated region. HuR-dependent upregulation of PARG expression facilitated DNA repair via hydrolysis of polyADP-ribose on related repair proteins. Accordingly, strategies to inhibit HuR directly promoted DNA damage accumulation, inefficient PAR removal, and persistent PARP-1 residency on chromatin (PARP-1 trapping). Immunoprecipitation assays demonstrated that the PARP-1 protein binds and posttranslationally modifies HuR in PARPi-treated PDAC cells. In a mouse xenograft model of human PDAC, PARPi monotherapy combined with targeted silencing of HuR significantly reduced tumor growth compared with PARPi therapy alone. Our results highlight the HuR–PARG axis as an opportunity to enhance PARPi-based therapies. Cancer Res; 77(18); 5011–25. ©2017 AACR.

HuR regulates all isoforms of PARG by binding a discrete minimal binding region (MBR) within its 3'UTR in both DDR- proficient MIA PaCa-2 and DDR- deficient Hs 766t cells. Such PARG mRNA and protein expression is also increased with other non-PARPi stress such as gemcitabine and oxaliplatin treatment, which simultaneously induce HuR cytoplasmic translocation.

Supplementary Table 1 describes BRCA and KRAS status of PDX models and IC50 of PDDX-001 and olaparib in PDAC/PDX lines

Table S1. Sequencing primers for IDO2 functional polymorphisms. Table S2. IDO2 genotype functionality definitions. A three-tier scale describes possible genotype combinations of IDO2 functional polymorphisms.

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