Faculty Bibliography

Background Metastatic pancreatic adenocarcinoma (mPDAC) remains notoriously treatment-refractory, particularly to immunotherapy; however, recent promise has been demonstrated with chemoimmunotherapy combinations.1,2 REVOLUTION is an adaptive platform trial, designed to further these advancements by assessing the safety and antitumor activity of parallel, novel chemoimmunotherapy combinations in patients with untreated mPDAC. Coupled with deep immune biomarker profiling, this approach will enable rapid insights from each combination, generating data to be leveraged for future cohorts. REVOLUTION also builds upon the collaborative framework between academic, nonprofit and industry partners, laid by the PRINCE trial.1 Methods REVOLUTION is an open-label, non-randomized, exploratory platform trial. Each cohort utilizes a Simon twostage design: Stage 1 enrolling n=15 patients, expansion to Stage 2 (an additional n=15 patients) based on the totality of safety, efficacy and biomarker analyses. Key inclusion criteria histologically or cytologically confirmed, treatment-naive, recurrent or de novo mPDAC, measurable by RECIST 1.1. Primary endpoints: safety, as assessed by the incidence and severity of adverse events. Secondary endpoints: ORR (per RECIST 1.1), DCR, DOR, PFS, and OS. Exploratory endpoints: pharmacodynamics and association of tumor, blood, and stool biomarkers with clinical activity. Three cohorts are underway, all using a backbone of standard- of-care gemcitabine/nab-paclitaxel (gem/nP). Cohort A: nivolumab + ipilimumab + gem/nP. We hypothesize chemotherapy will induce antigen release, ipilimumab will enhance T cell activation, proliferation and tumor infiltration, and nivolumab will overcome immunosuppression while re-invigorating therapeutically relevant T cells. Cohort B: high-dose hydroxychloroquine (HCQ), an autophagy inhibitor, + ipilimumab + gem/nP. The same mechanisms of action for chemotherapy and ipilimumab as Cohort A are hypothesized, with HCQ augmenting T cell priming and cytotoxicity by upregulating MHC-1.3 Cohort C: NG-350A, an intravenously administered adenovirus that selectively replicates in tumor cells and expresses a fully human agonistic CD40 monoclonal antibody, + ipilimumab + gem/nP. The same mechanisms of action for chemotherapy and ipilimumab as Cohorts A and B are hypothesized, with NG-350A re-programming the tumor microenvironment, activating antigen-presenting cells, and facilitating immune priming.4 In accordance with recent findings, all current cohorts are also testing a novel dosing schedule of ipilimumab (2 doses at 1 mg/kg, Q6W).5 Results Cohorts A and B are fully enrolled for Stage 1 and accumulating data to support an expansion decision. Cohort C is in development

RATIONALE/BACKGROUND:Patient-derived organoids (PDOs) are a promising technology to support precision medicine initiatives for patients with pancreatic ductal adenocarcinoma (PDAC). PDOs may improve clinical next-generation sequencing (NGS) and enable rapid ex vivo chemotherapeutic screening (pharmacotyping). METHODS:PDOs were derived from tissues obtained during surgical resection and endoscopic biopsies and studied with NGS and pharmacotyping. PDO-specific pharmacotype is assessed prospectively as a predictive biomarker of clinical therapeutic response by leveraging data from a randomized-controlled clinical trial. RESULTS:Clinical sequencing pipelines often fail to detect PDAC-associated somatic mutations in surgical specimens that demonstrate a good pathological response to previously administered chemotherapy. Sequencing the PDOs derived from these surgical specimens, after biomass expansion, improves the detection of somatic mutations and enables quantification of copy number variants. The detection of clinically relevant mutations and structural variants is improved following PDO biomass expansion. On clinical trial, PDOs were derived from biopsies of treatment naïve patients prior to treatment with FOLFIRINOX (FFX). Ex vivo PDO pharmacotyping with FFX components predicted clinical therapeutic response in these patients with borderline resectable or locally advanced PDAC treated in a neoadjuvant or induction paradigm. PDO pharmacotypes suggesting sensitivity to FFX components were associated with longitudinal declines of tumor marker, CA-19-9 and favorable RECIST imaging response. CONCLUSION/CONCLUSIONS:PDOs establishment from tissues obtained from patients previously receiving cytotoxic chemotherapies can be accomplished in a clinically-certified laboratory. Sequencing PDOs following biomass expansion improves clinical sequencing quality. High in-vitro sensitivity to standard-of-care chemotherapeutics predicts good clinical response to systemic chemotherapy in PDAC.

PURPOSE/UNASSIGNED:We sought to determine the feasibility of delivering a Supportive Oncology Care at Home intervention among patients with pancreatic cancer. METHODS/UNASSIGNED:We prospectively enrolled patients with pancreatic cancer from a parent trial of neoadjuvant fluorouracil, leucovorin, oxaliplatin, and irinotecan (FOLFIRINOX). The intervention entailed (1) remote monitoring of patient-reported symptoms, vital signs, and body weight; (2) a hospital-at-home care model; and (3) structured communication with the oncology team. We defined the intervention as feasible if ≥ 60% of patients enrolled in the study and ≥ 60% completed the daily assessments within the first 2-weeks of enrollment. We determined rates of treatment delays, urgent clinic visits, emergency department visits, and hospitalizations among those who did (n = 20) and did not (n = 24) receive Supportive Oncology Care at Home from the parent trial. RESULTS/UNASSIGNED:62.5%) compared with those not receiving the intervention from the same parent trial. CONCLUSION/UNASSIGNED:Findings demonstrate the feasibility and acceptability of a Supportive Oncology Care at Home intervention. Future work will investigate the efficacy of this intervention for decreasing health care use and improving patient outcomes.

SignificanceThe highly desmoplastic and immunosuppressive microenvironment of pancreatic tumors is a major determinant of the aggressive nature and therapeutic resistance of pancreatic cancer. Therefore, improving our understanding of the mechanisms that regulate the composition and function of the pancreatic tumor microenvironment is critical for the design of intervention strategies for this devastating malignancy. This study identifies a modality for the reprogramming of tumor-associated macrophages involving collagen scavenging followed by a metabolic switch toward a profibrotic paracrine phenotype. These findings establish a molecular framework for the elucidation of regulatory processes that could be harnessed to mitigate the stroma-dependent protumorigenic effects in pancreatic cancer.

Activating mutations in KRAS (KRAS*) are present in nearly all pancreatic ductal adenocarcinoma (PDAC) cases and critical for tumor maintenance. By using an inducible KRAS* PDAC mouse model, we identified a deubiquitinase USP21-driven resistance mechanism to anti-KRAS* therapy. USP21 promotes KRAS*-independent tumor growth via its regulation of MARK3-induced macropinocytosis, which serves to maintain intracellular amino acid levels for anabolic growth. The USP21-mediated KRAS* bypass, coupled with the frequent amplification of USP21 in human PDAC tumors, encourages the assessment of USP21 as a novel drug target as well as a potential parameter that may affect responsiveness to emergent anti-KRAS* therapy.

Oxygen is critical for a multitude of metabolic processes that are essential for human life. Biological processes can be identified by treating cells with ¹⁸O₂ or other isotopically labelled gases and systematically identifying biomolecules incorporating labeled atoms. Here we labelled cell lines of distinct tissue origins with ¹⁸O₂ to identify the polar oxy-metabolome, defined as polar metabolites labelled with ¹⁸O under different physiological O₂ tensions. The most highly ¹⁸O-labelled feature was 4-hydroxymandelate (4-HMA). We demonstrate that 4-HMA is produced by hydroxyphenylpyruvate dioxygenase-like (HPDL), a protein of previously unknown function in human cells. We identify 4-HMA as an intermediate involved in the biosynthesis of the coenzyme Q10 (CoQ10) headgroup in human cells. The connection of HPDL to CoQ10 biosynthesis provides crucial insights into the mechanisms underlying recently described neurological diseases related to HPDL deficiencies^1-4 and cancers with HPDL overexpression⁵.

α-ketoglutarate (KG), also referred to as 2-oxoglutarate, is a key intermediate of cellular metabolism with pleiotropic functions. Cell-permeable esterified analogs are widely used to study how KG fuels bioenergetic and amino acid metabolism and DNA, RNA, and protein hydroxylation reactions, as cellular membranes are thought to be impermeable to KG. Here we show that esterified KG analogs rapidly hydrolyze in aqueous media, yielding KG that, in contrast to prevailing assumptions, imports into many cell lines. Esterified KG analogs exhibit spurious KG-independent effects on cellular metabolism, including extracellular acidification, arising from rapid hydrolysis and de-protonation of α-ketoesters, and significant analog-specific inhibitory effects on glycolysis or mitochondrial respiration. We observe that imported KG decarboxylates to succinate in the cytosol and contributes minimally to mitochondrial metabolism in many cell lines cultured in normal conditions. These findings demonstrate that nuclear and cytosolic KG-dependent reactions may derive KG from functionally distinct subcellular pools and sources.

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive disease with a 5-year survival rate of <10%. The tumour microenvironment (TME) of PDAC is characterized by excessive fibrosis and deposition of extracellular matrix, termed desmoplasia. This unique TME leads to high interstitial pressure, vascular collapse and low nutrient and oxygen diffusion. Together, these factors contribute to the unique biology and therapeutic resistance of this deadly tumour. To thrive in this hostile environment, PDAC cells adapt by using non-canonical metabolic pathways and rely on metabolic scavenging pathways such as autophagy and macropinocytosis. Here, we review the metabolic pathways that PDAC use to support their growth in the setting of an austere TME. Understanding how PDAC tumours rewire their metabolism and use scavenging pathways under environmental stressors might enable the identification of novel therapeutic approaches.

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest forms of cancer. The elevated macroautophagy/autophagy in these tumors supports growth, promotes immune evasion, and increases therapeutic resistance. Therefore, targeting autophagy is a therapeutic strategy that is being pursued to treat PDAC patients. Whereas autophagy inhibition impairs mitochondrial metabolism in PDAC, the specific metabolite(s) that becomes limiting when autophagy is inhibited has not been identified. We report that loss of autophagy specifically results in intracellular cysteine depletion under nutrient-replete conditions. Mechanistically, we show that PDAC cells utilize the autophagy machinery to regulate the activity and localization of the cystine transporter SLC7A11 at the plasma membrane. Upon inhibition of autophagy, SLC7A11 is localized to lysosomes in an MTORC2-dependent manner. Our findings reveal a novel connection between autophagy and cysteine metabolism in pancreatic cancer.

Metabolic reprogramming enables cancer cell growth, proliferation, and survival. This reprogramming is driven by the combined actions of oncogenic alterations in cancer cells and host cell factors acting on cancer cells in the tumor microenvironment. Cancer cell intrinsic mechanisms activate signal transduction components that either directly enhance metabolic enzyme activity or upregulate transcription factors that in turn increase expression of metabolic regulators. Extrinsic signaling mechanisms involve host-derived factors that further promote and amplify metabolic reprogramming in cancer cells. This review describes intrinsic and extrinsic mechanisms driving cancer metabolism in the tumor microenvironment and how such mechanisms may be targeted therapeutically.