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PDF file - 1066KB, MEK inhibition alone and combination PI3K + MEK inhibition decreases serotonin secretion, while BEZ235 decreases CgA expression. (A) BON and (B) QGP-1 cells were plated in 24 well plates at a density of 1x105 cells/cm2. After 24h, cells were washed and maintained with serum-free medium. Cells were subsequently treated with either (upper panels) control, BKM120 (5 �M), PD0325901 (100 nM), or combination BKM120 + PD0325901 (5 �M/100 nM), or (lower panels) control, BEZ235 (1000 nM), PD0325901 (100 nM), or combination BEZ235 + PD0325901 (1000 nM/100 nM) for 1h. Media was collected and serotonin measured by serotonin ELISA (* p < 0.05 vs. control). (C) Whole cell lysates were collected following 24h of drug treatment with either control, BKM120 (5 �M), PD0325901 (100 nM), or combination BKM120 + PD0325901 (5 �M/100 nM), or control, BEZ235 (1000 nM), PD0325901 (100 nM), or combination BEZ235 + PD0325901 (1000 nM/100 nM) and analyzed for CgA expression. β-actin was used as a loading control.

PDF file - 649KB, Combination PI3K and MEK inhibitor treatment results in a more pronounced decrease in NCI-H727 and QGP-1 cell proliferation. (A) NCI-H727 and (B) QGP-1 cells were plated in 24 well plates at a density of 3x104 cells/well. Treatments were initiated the following morning with either control, BKM120 (5 μM), PD0325901 (100 nM), or combination BKM120 + PD0325901 (5 μM/100 nM) (upper panels), or control, BEZ235 (1000 nM), PD0325901 (100 nM), or combination BEZ235 + PD0325901 (1000 nM/100 nM) (lower panels). Media was exchanged for fresh drug media at 36h. Proliferation was assessed directly by cell counting at 72h (* p < 0.05 vs. control; � p < 0.05 vs. monotherapy).

PDF file - 812 KB, PI3K inhibition decreases cell proliferation in NCI-H727 and QGP-1 cell lines. (A) NCI-H727 and (B) QGP-1 cells were plated in 24 well plates at a density of 3x104 cells/well. Treatments were initiated the following morning with increasing doses of either BKM120 (0-5 μM) (upper panels) or BEZ235 (0-1000 nM) (lower panels). Media was exchanged for fresh drug media again at 36h. Proliferation was assessed directly by cell counting at 72h (* p < 0.05 vs. control). (C) BON cells transfected with shRNA directed against the alpha subunit of PI3K were plated in 24 well plates at a density of 3x104 cells/well and proliferation assessed at 24h intervals up to 96h.

PDF file - 741KB, PI3K inhibition enhances apoptosis in NCI-H727 and QGP-1 cell lines. (A) NCI-H727 and (B) QGP-1 cells were plated in 24 well plates at a density of 5x104 cells/cm2. Treatments were initiated the following morning with increasing doses of either BKM120 (0-5 μM) (upper panels) or BEZ235 (0-1000 nM) (lower panels) and continued for 24h. Apoptosis was measured by DNA fragmentation using the Cell Death Detection ELISAplus (* p < 0.05 vs. control).

Background: The precise involvement of the PI3K/mTOR and RAS/MEK pathways in carcinoid tumors is not well defined. Therefore, the purpose of our study was to evaluate the role these pathways play in carcinoid cell proliferation, apoptosis, and secretion and to determine the effects of combined treatment on carcinoid tumor inhibition.Methods: The human neuroendocrine cell lines BON (pancreatic carcinoid), NCI-H727 (lung carcinoid), and QGP-1 (somatostatinoma) were treated with either the pan-PI3K inhibitor, BKM120, or the dual PI3K–mTOR inhibitor, BEZ235, alone or in combination with the MEK inhibitor, PD0325901; proliferation, apoptosis, and protein expression were assessed. Peptide secretion was evaluated in BON and QGP-1 cells. The antiproliferative effect of BEZ235, alone or combined with PD0325901, was then tested in vivo.Results: Both BKM120 and BEZ235 decreased proliferation and increased apoptosis; combination with PD0325901 significantly enhanced the antineoplastic effects of either treatment alone. In contrast, neurotensin peptide secretion was markedly stimulated with BKM120 treatment, but not BEZ235. The combination of BEZ235 + PD0325901 significantly inhibited the growth of BON xenografts without systemic toxicity.Conclusions: Both BKM120 and BEZ235 effectively inhibited neuroendocrine tumor (NET) cell proliferation and stimulated apoptosis. However, inhibition of the PI3K pathway alone with BKM120 significantly stimulated neurotensin peptide secretion; this did not occur with the dual inhibition of both PI3K and mTOR using BEZ235 suggesting that this would be a more effective treatment regimen for NETs. Moreover, the combination of BEZ235 and the MEK inhibitor PD0325901 was a safe and more effective therapy in vivo compared with single agents alone. Clin Cancer Res; 20(5); 1212–22. ©2014 AACR.

PDF file - 1080KB, Combination PI3K + MEK inhibition enhances apoptosis compared to monotherapy. (A) NCI-H727 and (B) QGP-1 cells were plated in 24 well plates at a density of 5x104 cells/cm2. Treatments were initiated the following morning with either control, BKM120 (5 μM), PD0325901 (100 nM), or combination BKM120 + PD0325901 (5 μM/100 nM) (upper panels), or control, BEZ235 (1000 nM), PD0325901 (100 nM), or combination BEZ235 + PD0325901 (1000 nM/100 nM) (lower panels) and continued for 24 hours. Apoptosis was measured by DNA fragmentation using the Cell Death Detection ELISAplus (* p < 0.05 vs. control; � p < 0.05 vs. monotherapy). (C) BON cells were plated in 6 well plates at 9 x 105 cells/well. After 24h of treatment, cells were collected, stained with annexin V Alexa Fluor 488, and analyzed by flow cytometry (* p < 0.05 vs. control; � p < 0.05 vs. monotherapy).

PDF file - 1864KB, mTOR inhibition decreases proliferation but does not increase apoptosis or enhance effect in combination with a MEK inhibitor. (A) BON, (B) NCI-H727 and (C) QGP-1 cells were plated in 24 well plates at a density of 3x104 cells/well. Treatments were initiated the following morning with either control, rapamycin (20 nM), PIK-75 (1 μM), PD98059 (10 μM), rapaymycin + PD98059 (20 nM/10 μM), or PIK-75 + PD98059 (1 μM /10 μM). Proliferation (upper panels) was assessed directly by cell counting at 72h (* p < 0.05 vs. control; � p < 0.05 vs. monotherapy). Apoptosis (middle panels) was measured by DNA fragmentation using the Cell Death Detection ELISAplus (* p < 0.05 vs. control; � p < 0.05 vs. monotherapy). For Western blot analysis (lower panels), cells were plated in 6 well plates at a density of 5 x104 cells/cm2. Whole cell lysates were collected following 24h of drug treatment. β-actin was used as a loading control.

Background Treatment recommendations for patients with neuroendocrine tumors (NETs) include the use of octreotide long-acting release (LAR) for long-term therapy and immediate-release (IR) as rescue therapy to control the breakthrough symptoms of carcinoid syndrome (CS). High doses of LAR are commonly used in clinical practice. This study aimed to evaluate the real-world utilization of LAR and preceding IR use at the prescription and patient levels. Methods We used an administrative claims database (2009-2018) containing privately insured enrollees. We calculated the normalized LAR dose from pharmacy claims and the initial mean IR daily dose at the prescription level. At the patient level, we conducted a retrospective cohort study that included patients continuously enrolled with ≥1 pharmacy claim of LAR and evaluated the frequency and the clinical reason for dose escalation of LAR. The definition of the above-label maximum dose of LAR was ≥30 mg/4 weeks. Results Nineteen percent of LAR prescriptions had an above-label maximum dose. Only 7% of LAR prescriptions had preceding IR use. There were 386 patients with NETs or CS vs. 570 with an unknown diagnosis. Comparing patients with NETs or CS to those with an unknown diagnosis, 22.3% vs. 11.0 % experienced dose escalations and 29.0% vs. 26.6% had IR use before dose escalation, respectively. LAR dose escalation occurred in 50.9% vs. 39.2% for symptom control, 12.3% vs. 7.1% for tumor progression control, and 16.6% vs. 6.0% for both reasons in NETs/CS and unknown groups, respectively. Conclusion Octreotide LAR dosing above the label-maximum dose is common and IR rescue dosing appears to be underutilized.

Rationale and Objectives Standardized Uptake Value (SUV) is an important semiquantitative measurement used in the clinical and research domains to assess radiopharmaceutical concentration in tumors versus normal organs, but is susceptible to many factors beyond the tumor biological environment. So, the aim of this study is to identify the optimum internal reference among organs with physiological uptake in 68Ga-DOTATATE PET/CT (DOTA PET/CT) scans. Materials and Methods This HIPAA-compliant, IRB-approved study with waiver of consent included retrospective imaging review of 180 consecutive patients with neuroendocrine tumors presenting for DOTA PET/CT image acquisition: Ga-68 DOTATATE dose was reported as (0.054 mCi/Kg) scans between September 2018 and May 2019. Mean value of body weight normalized SUV (SUVbw) and lean body mass normalized SUV (SUL) of liver and spleen were measured. Information about the patients and scan characteristics were collected. The paired Grambsch test was used to compare variance among the measured SUVs. Spearman's rank correlation coefficient was used to assess correlation between SUVs and potential patient- and scan-specific confounding factors. Results Variance of SUL was significantly lower than variance of SUVbw in both liver and spleen (p-value Conclusion SUL is a more reproducible, less variable, and therefore more reliable quantitative measure in DOTA PET/CT scans, compared SUVbw. Among the available organs with physiological uptake, liver SUL is the optimum internal reference given the liver's larger size and uniform SUL values resulting in lower variability and better reproducibility.

Octreotide acetate (octreotide) is the most prescribed and most studied somatostatin congener, or analog, for gastroenteropancreatic neuroendocrine tumors (GEP‐NETs) and carcinoid syndrome, the latter of which may be characterized by debilitating diarrhea and flushing. Approved in the U.S. more than 30 years ago, octreotide is widely used to control the symptoms of carcinoid syndrome and has been shown to demonstrate antiproliferative activity. The two formulations available in the U.S. include a subcutaneous immediate‐release (IR) injection introduced in 1989 and a long‐acting repeatable (LAR) intramuscular injection approved in 1999. Lanreotide depot (lanreotide), a more recent somatostatin congener, has been available in the U.S. since 2014. Despite widespread use of octreotide LAR, several key challenges exist with the current depot‐based treatment paradigm. Studies indicate that LAR formulations are associated with continued unmet patient needs, owing in part to a loss of bioactivity over time that may necessitate progressive supplemental treatment with IR octreotide to adequately control symptoms. Clinicians should understand the key differences in the pharmacokinetic profiles of the LAR and IR formulations that may contribute to bioactivity loss and somatostatin receptor desensitization. In addition, there is a need to re‐evaluate the role of IR octreotide in combination with depot therapy to provide consistent bioavailability and better control of carcinoid syndrome symptoms. The purpose of this review is to explore all these issues and to re‐establish a rationale for the IR formulation, particularly with respect to novel use cases and its use during the COVID‐19 pandemic. Implications for Practice There is a need to re‐evaluate the role of immediate‐release octreotide in combination with depot therapy to provide consistent bioavailability and better control of carcinoid syndrome symptoms., This review explores key challenges with depot‐based treatment of gastroenteropancreatic neuroendocrine tumors with octreotide and the key differences between the LAR and IR formulations.

Background Telotristat ethyl is approved to treat carcinoid syndrome diarrhea in combination with somatostatin analogs. In TELESTAR and TELECAST phase III studies, patients with carcinoid syndrome received telotristat ethyl 250 or 500 mg 3 times per day (tid) or placebo tid in addition to somatostatin analogs. The aim of this prespecified analysis was to examine the time to reductions in bowel movements (BMs) in the TELESTAR and TELECAST studies using survival analysis methods. Methods First occurrence of sustained response was defined as the time to the first day of 2 consecutive weeks with a mean BM frequency improvement of ≥ 30% from baseline during the 12-week double-blind treatment periods. Time to first ≥ 30% worsening in BM frequency was also measured. Treatments were compared with the log-rank test; Cox regression models provided point and confidence interval estimates of the hazard ratios for each trial. Results In TELESTAR and TELECAST, majority of patients (69%) on telotristat ethyl experienced a sustained ≥ 30% improvement in BM frequency. The median time to sustained reduction of at least 30% in BM frequency was significantly faster (fewer days to onset) for telotristat ethyl compared with placebo in both TELESTAR (250 mg, HR = 2.3 [95% CI, 1.3–4.1, P = 0.004]; 500 mg, HR = 2.2 [95% CI, 1.2–3.9, P = 0.009]) and TELECAST (250 mg, HR = 3.9 [95% CI, 1.6–11.1, P = 0.003]; 500 mg, HR = 4.2 [95% CI, 1.7–11.7, P = 0.002]). In TELECAST, 42% of patients on placebo experienced sustained worsening in BM frequency compared with 20% on telotristat ethyl; no significant difference was observed in TELESTAR. Conclusion The time of onset of sustained BM frequency improvement mean and range are important when considering use of telotristat ethyl in patients with carcinoid syndrome diarrhea. Telotristat ethyl may also reduce sustained worsening in BM frequency. Trial Registration ClinicalTrials.gov Identifiers: NCT01677910, NCT02063659

Current blood-based biomarkers for neuroendocrine neoplasms (NENs) lack both sensitivity and specificity. Human circulating progastrin (hPG80) is a novel biomarker that can be easily measured in plasma by ELISA. This study is the first to examine hPG80 in NENs. Plasma hPG80 was quantified from 95 stage IV NEN patients, using DxPG80 technology (ECS Progastrin, Switzerland) and compared with hPG80 concentrations in two cohorts of healthy donor controls aged 50–80 (n = 252) and 18–25 (n = 137). Median hPG80 in NENs patients was 5.54 pM compared to 1.5 pM for the 50–80 controls and 0.29 pM the 18–25 cohort (p < 0.0001). Subgroup analysis revealed median hPG80 levels significantly higher than for either control cohort in neuroendocrine carcinoma (NEC; n = 25) and neuroendocrine tumors (NET; n = 70) including the small-cell lung cancer (SCLC) sub-cohort (n = 13). Diagnostic accuracy, estimated by AUCs, was high for NENs, as well as both sub-groups (NEC/NET) when compared to the younger and older control groups. Plasma hPG80 in NENs may be a diagnostic blood biomarker for both low- and high-grade NENs; further study is warranted. A prospective multi-center trial is ongoing in NET to evaluate hPG80 as a means of monitoring disease (NCT04750954).

Purpose: Immune checkpoint inhibitor (ICI) maintenance therapy is standard of care for frontline management of extensive stage small cell lung cancer (ES SCLC). However, overall survival benefit of addition of ICI maintenance to frontline ES SCLC treatment is modest and further improvement is needed. We hypothesized that addition of poly (ADP-ribose) polymerase inhibition to ICI maintenance therapy for patients with platinum sensitive ES SCLC could improve antitumor efficacy of ICI. Methods: A single-arm, investigator-initiated phase II trial (NCT03958045) enrolled patients with platinum sensitive ES SCLC who received frontline maintenance nivolumab 480 mg IV every 4 weeks, and rucaparib, 600 mg PO twice a day after completion of 4-6 cycles of platinum doublet. The primary outcome was median progression free survival. Secondary endpoints included assessment of objective response and adverse effects (AEs) per CTCAE 5.0. Correlative studies included pretreatment and during-treatment immune assays and circulating tumor DNA TP53 mutation status. Results: A total of 42 patients were consented and 33 met eligibility criteria and were treated. All patients received 4-6 cycles of frontline platinum doublet and had at least a partial response by RECIST at the time of enrollment. In the 33 participants, the most common grade 3 and 4 AEs (at least possibly related) were hypokalemia (3%), hyponatremia (3%), elevated alanine aminotransferase (3%), neutropenia (3%) and leukocytopenia (3%). No grade 5 AE was noted. The median PFS (mPFS) was 3 months from time of enrollment on frontline maintenance (post platinum doublet). The mPFS was 11 months from cycle 1 of platinum doublet. Overall, 89.8% patients were alive at 12 months and 54.4 % patients were alive at 24 months from the start of platinum doublet. Currently 2 patients are on active treatment and other two have completed study treatment and are on observation with stable disease. Conclusions: Maintenance rucaparib combined with immune checkpoint inhibition was tolerable and showed promising activity after completion of frontline chemotherapy in platinum sensitive extensive stage small cell lung cancer patients. Citation Format: Aman Chauhan, Jill Kolesar, Donglin Yan, Zhonglin Hao, Ronald McGarry, John Villano, Ralph Zinner, Ashish Maskey, Jordan Miller, Timothy Mullett, Aman Khurana, Xitong Zhou, Garima Gupta, Daniel Flora, Colleen Darnell, Richard O'Neil, Charles Kunos, Mark B. Evers, Lowell Anthony, Susanne Arnold. Phase II of frontline maintenance rucaparib in combination with nivolumab in ES SCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT221.

Background: Radiation is a potent inducer of DNA double-strand breaks, and ribonucleotide reductase (RNR) is the rate-limiting enzyme in the synthesis and repair of DNA, making RNR-targeted therapy a rationale therapeutic strategy for radiosensitization. ETCTN 10388 (NCT04234568) evaluated safety and efficacy of the combination of lutetium 177 DOTATATE, a beta-emitting radionuclide in combination with triapine, a ribonucleotide reductase (RNR) inhibitor. Method: This study was a multicenter phase 1 dose escalation trial [using the Bayesian optimal interval design (BOIN)] of triapine in combination with fixed dose lutetium Lu 177 DOTATATE for well-differentiated somatostatin receptor-positive gastroenteropancreatic neuroendocrine tumor (GEP-NETs) after the failure of at least one line of prior systemic cancer treatment with an expansion cohort at the recommended phase 2 dose (RP2D). Oral triapine (100mg, 150mg, 200mg) was administered once daily on days 1-14 and Lu-177 DOTATATE [200 mCi] intravenously on day 1 of every 56-day cycle. A total of 4 cycles were administered. Response and adverse effects were assessed per RECIST and CTCAE 5.0, respectively. Exploratory correlative studies included tumor somatic and germline mutation testing, RNA sequencing, pharmacokinetics, deoxynucleosides and circulating cell free DNA analysis. Primary endpoints were safety and RP2D. Results: Overall, 31 patients were enrolled between 6 sites, 15 in the dose escalation phase and 16 in the dose expansion phase. Adverse events (AE) were assessed in all 31 patients per CTCAE 5.0. One DLT in dose level 1, seven DLTs in dose level 2, and one grade 5 DLT in dose level 3 were observed. The RP2D of the combination is triapine 150 mg QD (dose level 2) on days 1-14 in combination with Lu-177 DOTATATE on day 1 of every 56-day cycle. Detailed safety and adverse event data will be presented at the meeting. There were 28 patients evaluable for efficacy, of which 6 (21%) achieved a partial response. At 12 months, 6 patients had progressed, while 22 (86%) remained progression free. Median PFS has not been reached. PK data were available for 12 patients enrolled in the dose escalation cohort. The geometric mean (SD) AUC0-inf was 1159 (1.22) µg/L•h for the 100mg dose level and 1862 (1.76) µg/L•h for the 150 mg dose level, suggesting that exposure increased with dose, and inter-patient variability was as expected for an oral agent. Conclusion: The combination of triapine and Lu-177 DOTATATE was safe with preliminary efficacy signals, which will be further evaluated in ETCTN 10558, a randomized phase 2 study that is comparing the effectiveness of triapine and Lu-177 DOTATATE to Lu-177 DOTATATE alone. Citation Format: Aman Chauhan, Susanne Arnold, Jill Kolesar, William Carson, Heidi Weiss, Rani Jayswal, Donglin Yan, Riham El Khouli, Aman Khurana, Jan Beumer, Heloisa Soares, Mary Mulcahy, Thorvardur Halfdanarson, Daneng Li, Heather Jacene, Percy Ivy, Elise Kohn, John Wright, Larry Rubinstein, Zeta Chow, Piotr Rychahou, Mark B. Evers, Charles Kunos, Lowell Anthony, Bhavana Konda. ETCTN 10388: a first in human phase I trial of triapine and lutetium Lu 177 DOTATATE in well-differentiated somatostatin receptor-positive gastroenteropancreatic neuroendocrine tumors (GEP-NETs) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT194.

[Introduction] Telotristat ethyl is indicated for use in combination with somatostatin analogs (SSAs) to treat carcinoid syndrome (CS) diarrhea uncontrolled by SSAs alone in adults, but long-term safety and efficacy data beyond 48 weeks are needed., [Objectives] The aims of the study were to evaluate the long-term safety and tolerability of telotristat ethyl and its effect on quality of life (QOL) in patients with CS., [Methods] In this phase 3, nonrandomized, multicenter, open-label, long-term extension study (TELEPATH), patients who participated in phase 2 or 3 trials of telotristat ethyl continued treatment at their present dose level (250 or 500 mg thrice daily) for 84 weeks. Safety and tolerability, the primary endpoint, were assessed by monitoring adverse events (AEs), serious AEs, AEs of special interest (AESIs; including liver-related AEs, depression, and gastrointestinal AEs), and deaths. The secondary objective was to evaluate changes in patients’ QOL using validated cancer questionnaires and a subjective global assessment of CS symptoms, [Results] In 124 patients exposed to telotristat ethyl for a mean of 102.6 ± 53.2 weeks, the type and frequency of AEs were consistent with those reported in previous trials. The occurrence of AESIs was not related to dosage or duration of therapy. Most AEs were mild to moderate in severity, and no deaths were related to telotristat ethyl. QOL scores remained stable, and the majority of patients reported adequate symptom relief throughout the study., [Conclusions] Safety results of TELEPATH support the long-term use of telotristat ethyl in patients with CS diarrhea. Telotristat ethyl was well-tolerated and associated with sustained improvement in QOL scores (NCT02026063)., This study was supported and conducted by Lexicon Pharmaceuticals, Inc.

Current blood-based biomarkers for neuroendocrine neoplasms (NENs) lack both sensitivity and specificity. Human circulating progastrin (hPG

Background Carcinoid syndrome is associated with a reduced quality of life that can be attributed to symptoms such as diarrhea and fatigue as well as social and financial issues. This study was conducted to psychometrically assess meaningful change in bowel movement frequency among carcinoid syndrome patients using data from the TELESTAR clinical study. Methods An anchor-based approach for deriving meaningful change thresholds consisted of mapping change from baseline bowel movement frequency to other patient-reported assessments of change. These included the European Organization for Research and Treatment of Cancer (EORTC) Quality of Life Questionnaire - Core Questionnaire (QLQ-C30) Diarrhea Symptom responders, the EORTC Gastrointestinal NET questionnaire (GI.NET21) GI Symptom responders, and reported adequate relief at Week 12 (≥ 10-point score decrease from Day 1 to Week 12). Parameters included within-group mean change from baseline to Week 12, t-tests of the change (Wilcoxon rank sum for adequate relief), and effect size. Results There were 135 carcinoid syndrome patients with a mean baseline frequency of 5.7 bowel movements a day. A distribution-based method yielded meaningful change estimates of 0.62 bowel movements a day for overall frequency and 0.83 bowel movements a day at Week 12. Anchor-based analysis indicated a large effect size among patients who reported adequate relief at Week 12 (− 1.58; n = 18; P = 0.014), the QLQ-C30 Diarrhea domain responders (− 1.24; n = 40; P n = 25; P = 0.005). Exit interview data for meaningful change yielded effect size estimates of − 1.57 for overall change during the Double-blind Treatment Period and − 1.97 for change between Baseline and Week 12. Conclusions Meaningful change derivation is critical to interpret patient outcomes for evaluating treatment efficacy. In this study, carcinoid syndrome patients experienced clinically meaningful reductions in bowel movement frequency of ≥30% over 12 weeks with telotristat ethyl treatment. Trial registration NCT01677910.

Immune checkpoint inhibitor (ICPi) therapy has transformed the way we treat cancer. However, its immune related adverse events (irAEs) can be debilitating and life threatening. Immune therapy-induced diarrhea (ITID) is one of the most commonly encountered irAEs and can lead to expensive and prolonged hospitalizations. The current standard of care for grade 3 or 4 ITID involves ICPi discontinuation, the initiation of steroids, and infliximab for refractory disease. This treatment regimen reverses the desired anti-tumor effect of ICPis, can lead to side effects, and is cost-ineffective. We report the first case of the successful treatment of grade 3 ITID with steroids and an amino acid-based oral rehydration solution (AA-ORS), enterade. Research suggests that AA-ORS may be used to reduce diarrhea and adequately hydrate patients, in contrast to glucose-based oral rehydration solutions, which have been implicated as a contributing factor to diarrhea in cancer patients. We hypothesize that an AA-ORS may mitigate ITID via safer and more economically viable means than the current standard of care, but more controlled trials are needed to test this hypothesis.

INTRODUCTION: Neurological immune-related adverse events are a rare but potentially deadly complication after immune checkpoint inhibitor (ICI) treatment. As multiple sclerosis (MS) is an immune mediated disease it is unknown how ICI treatment may affect outcomes. METHODS: We analyzed the United States Food and Drug Administration (FDA) Adverse Event Reporting System (FAERS) database for pembrolizumab, atezolizumab, nivolumab, ipilimumab, avelumab, and durvalumab two years prior their FDA approval until December 31, 2017, to include all cases with confirmed diagnosis/relapse of MS. We also included cases reported in the literature and a patient from our institution. RESULTS: We identified 14 cases of MS with median age of presentation of 52 years. Indications for ICI included melanoma in 7 (36.36%) cases, non-small cell lung carcinoma in 2 (18.18%) cases, 1 case (9.09%) each of pleural mesothelioma, renal cell carcinoma, and colorectal cancer, and unreported in 2 (18.18%) cases. History of MS was confirmed in 8 (57.1%) cases. Median time to beginning of symptoms was 29 days with rapid disease progression; 2 patients died due to their relapse. Median time for symptom resolution was 8 weeks. Outcomes did not vary by comparing CTLA-4 and PD-1/PD-L1 inhibitors. CONCLUSIONS: Reported MS relapses after ICI are rare but the adverse events described include rapid neurologic progression and death. Larger and prospective studies are warranted to assess disability and long-term outcomes and outweigh the risks of starting immunotherapy in patients with MS.

Large cell neuroendocrine carcinomas (LCNEC) are rare, aggressive high-grade neuroendocrine neoplasms within the neuroendocrine cell lineage spectrum. This manuscript provides a detailed review of published literature on LCNEC of gynecological origin. We performed a PubMed search for material available on gynecologic LCNEC. We analyzed 104 unique cases of gynecologic LCNECs, of which 45 were cervical primary, 45 were ovarian, 13 were uterine, and 1 was vaginal. A total of 45 cases of cervical LCNEC were identified with a median age of 36 years. Median overall survival was 16 months. We identified 45 ovarian LCNEC cases in the published literature with a median age of 54 years. Median overall survival was 8 months. 13 LCNEC cases of uterine origin were identified; 12 out of 13 were of endometrial origin and the median age was 71 years. The majority of patients presented with Stage III/IV disease (stages I–IV were 31%, 8%, 38%, and 23%, respectively). Gynecologic LCNEC is an aggressive malignancy. Our current understanding of the disease biology is very limited. Efforts are required to better understand the genomic and molecular characterizations of gynecological LCNEC. These efforts will elucidate the underlying oncogenic pathways and driver mutations as potential targets.

Gallium-68 DOTATATE provides physiologic imaging and assists in disease localization for somatostatin receptor (SSTR) positive neuroendocrine tumor (NET) patients. However, questions regarding usefulness of gallium- 68 DOTATATE imaging in identifying the primary site in neuroendocrine tumors (NETS) of unknown primary, correlation of NET grade with median Standardized Uptake Value (SUV) and effects of long acting somatostatin analog on gallium-68 DOTATATE imaging quality needs to be evaluated. A single institution retrospective review of the first 200 NET patients with gallium-68 DOTATATE imaging from Dec 2016 to Dec 2017 was conducted. Questions related to NETs of unknown primary, correlation of Standardized Uptake Value (SUV) to Ki-67 (which signifies proliferation rate), the effects of long-acting systemic somatostatin analog (SSA) on SUV were part of our data analysis. From these 200 patients, 59.5% (119) were females, 40.5% (81) were males; the median age was 62 years. The following primary tumor sites were identified: small bowel-37.5%; pancreas-18.5%; bronchial-14%; colon-3.5%; rectum-2%; appendix-1.5%; adrenal-0.5%; prostate-0.5%; others-3% and unknown primary-19%. Mean hepatic SUV of the lesion with the greatest radiolabeled uptake in 96 patients was similar irrespective to exposure to long acting SSA. Patients exposed to long acting SSA had mean SUV of 31.3 vs 27.8 for SSA naïve patients. The difference was not statistically significant. Gallium-68 DOTATATE imaging seems to distinguished G3 NET from G1/G2 based on mean SUV, and also identified the primary tumor site in 17 of 38 (45%) patients with unknown primary. Systemic exposure to long acting SSA does not appear to influence mean SUV of gallium-68 DOTATATE scan.

Time to Sustained Improvement in Bowel Movement Frequency With Telotristat Ethyl : Analysis of the Phase 3 TELESTAR Study

Inhibition of mTOR signaling using the rapalog everolimus is an FDA-approved targeted therapy for patients with lung and gastroenteropancreatic neuroendocrine tumors (NET). However, patients eventually progress on treatment, highlighting the need for additional therapies. We focused on pancreatic NETs (pNET) and reasoned that treatment of these tumors upon progression on rapalog therapy, with an mTOR kinase inhibitor (mTORKi), such as CC-223, could overcome a number of resistance mechanisms in tumors and delay cardiac carcinoid disease. We performed preclinical studies using human pNET cells in vitro and injected them subcutaneously or orthotopically to determine tumor progression and cardiac function in mice treated with either rapamycin alone or switched to CC-223 upon progression. Detailed signaling and RNA sequencing analyses were performed on tumors that were sensitive or progressed on mTOR treatment. Approximately 57% of mice bearing pNET tumors that progressed on rapalog therapy showed a significant decrease in tumor volume upon a switch to CC-223. Moreover, mice treated with an mTORKi exhibited decreased cardiac dilation and thickening of heart valves than those treated with placebo or rapamycin alone. In conclusion, in the majority of pNETs that progress on rapalogs, it is possible to reduce disease progression using an mTORKi, such as CC-223. Moreover, CC-223 had an additional transient cardiac benefit on valvular fibrosis compared with placebo- or rapalog-treated mice. These results provide the preclinical rationale to further develop mTORKi clinically upon progression on rapalog therapy and to further test their long-term cardioprotective benefit in those NET patients prone to carcinoid syndrome. Mol Cancer Ther; 16(11); 2432–41. ©2017 AACR.

Background Describe a single-center real-world experience with comprehensive genomic profiling (CGP) to identify genotype directed therapy (GDT) options for patients with malignancies refractory to standard treatment options. Methods Patients who had CGP by a CLIA-certified laboratory between November 2012 and December 2015 were included. The medical records were analyzed retrospectively after Institutional Review Board (IRB) approval. The treating oncologist made the decision to obtain the assay to provide potential therapeutic options. The objectives of this study were to determine the proportion of patients who benefited from GDT, and to identify barriers to receiving GDT. Results A total of 125 pediatric and adult patients with a histologically confirmed diagnosis of malignancy were included. Among these, 106 samples were from adult patients, and 19 samples were from pediatric patients. The median age was 54 years for adults. The majority had stage IV malignancy (53%) and were pretreated with 2–3 lines of therapy (45%). The median age was 8 years for pediatric patients. The majority had brain tumors (47%) and had received none or 1 line of therapy (58%) when the profiling was requested. A total of 111 (92%) patients had genomic alterations and were candidates for GDT either via on/off-label use or a clinical trial (phase 1 through 3). Fifteen patients (12%) received GDT based on these results including two patients who were referred for genomically matched phase 1 clinical trials. Three patients (2%) derived benefit from their GDT that ranged from 2 to 6 months of stable disease. Conclusions CGP revealed potential treatment options in the majority of patients profiled. However, multiple barriers to therapy were identified, and only a small minority of the patients derived benefit from GDT.

// Aman Chauhan 1 , Susanne M. Arnold 1 , Jill Kolesar 1 , Hala Elnakat Thomas 2 , Mark Evers 1 and Lowell Anthony 1 1 Markey Cancer Center, University of Kentucky, Lexington, KY, USA 2 University of Cincinnati, Cincinnati, OH, USA Correspondence to: Aman Chauhan, email: aman.chauhan@uky.edu Keywords: large cell neuroendocrine carcinoma; high grade neuroendocrine carcinoma; immune checkpoint inhibitors Received: December 23, 2017 Accepted: February 13, 2018 Epub: February 22, 2018 Published: March 06, 2018 ABSTRACT Introduction: Large cell neuroendocrine carcinomas (LCNEC) are a group of rare high grade neuroendocrine tumors that often behave clinically like small cell carcinoma (SCLC) and are treated as such. No major advancement in the management of these tumors has occurred in the last 30 years. Methods: We present a case series of three cases from Markey Cancer center along with a review of 13 published cases in the literature wherein immune-checkpoint inhibitors were utilized in the management of LCNEC. Results: Immune-checkpoint inhibitors might have clinical activity in LCNEC. Conclusion: Role of immune-checkpoint inhibitors should be explored in prospective LCNEC clinical trials. We summarize current evidence regarding use of immune checkpoint inhibitors in the treatment of LCNEC.

Neuroendocrine tumors (NETs) comprise a heterogeneous group of neoplasms in which tumor staging/prognosis and response to treatments depend heavily on accurate and timely identification of the anatomic primary site or NET subtype. Despite recent technological advancements and use of multiple diagnostic modalities, 10% to 14% of newly diagnosed NETs are not fully characterized based on subtype or anatomic primary site. Inability to fully characterize NETs of unknown primary may cause delays in surgical intervention and limit potential treatment options. To address this unmet need, clinical validity and utility are being demonstrated for novel approaches that improve NET subtype or anatomic primary site identification. Functional imaging using 68Ga-radiolabeled DOTATATE positron emission tomography/computed tomography has been shown to overcome some false-positive and resolution issues associated with octreotide scanning and computed tomography/magnetic resonance imaging. Using a genomic approach, molecular tumor classification based on differential gene expression has demonstrated high diagnostic accuracy in blinded validation studies of different NET types and subtypes. Given the widespread availability of these technologies, we propose an algorithm for the workup of NETs of unknown primary that integrates these approaches. Including these technologies in the standard workup will lead to better NET subtype identification and improved treatment optimization for patients.

Background: Midgut neuroendocrine tumor (NET) patients are often diagnosed at advanced stages with extensive mesenteric nodal and hepatic metastasis. The only potentially curative treatment is surgical tumor eradication. Despite an aggressive resection, macro and microscopic residual disease still may remain in the resection bed. We hypothesize that the application of 5-fluorouracil (5-FU) within the tumor bed will help eliminate microscopic residual disease. Methods: Records of 189 patients who underwent extensive cytoreductive surgeries during 2003–2012 for advanced, midgut NETs with extensive mesenteric lymphadenopathy were reviewed. Eighty-six patients (46%) who had 5-FU saturated gel foam strips secured into their mesenteric resection sites served as the study group and a matching 103 patients (54%) who did not have such an intra-operative chemotherapy served as controls. Survival from the time of diagnosis and post-operative complications between the two groups were compared. Results: Mortality rates at 30, 60 and 90 days post-operatively were 4%, 0%, 0% versus 2%, 0%, 2% for study and control groups, respectively. Major complications (Grades III & IV) at the same intervals were 0, 0, 1 versus 2, 3, 2 for study and control groups, respectively. Median survival was 236 months versus 148 months for the study and control groups, respectively 24 (P=0.15). Conclusions: Intraoperative tumor resection bed chemotherapy is a safe adjuvant without discernible toxicity. This procedure may provide survival benefits to midgut NET patients with extensive mesenteric lymphadenopathy undergoing extensive cytoreductive surgery. Further study in prospective trials must be conducted to determine definitive benefit to the NET patient.

Radiolabeled somatostatin analogues provide a means of delivering targeted radiation with a high therapeutic index to NETs that express somatostatin receptors (SSTRs). Radiolabeled somatostatin analogue Lutetium Lu 177 Dotatate (Lutathera) is a beta- and gamma-emitting radionuclide. Lutathera has been recently FDA approved for use in SSTR positive gastroenteropancreatic neuroendocrine tumors (GEPNETS) in the US based on NETTER-1 Phase III trial. Despite favorable PFS, and safety profile, the drug has limited cytoreductive capability. NETTER-1 reported 17% ORR. PRRT also doesn't seem to be very effective in treating peritoneal disease. We hypothesize that addition of an effective radiation sensitizer can help improve antitumor activity of Lutathera. Rationale: Triapine is a ribonucleotide reductase (RNR) inhibitor. Radiation is a potent inducer of DNA double-strand breaks (DSBs). Targeting signaling networks involved in DSB repair is a promising approach for enhancing cellular radio sensitivity. RNR is the rate-limiting enzyme in the synthesis and repair of DNA, and it is directly involved in the cellular response to radiation, making RNR-targeted therapy to enhance radiation treatment a rational therapeutic strategy. RNR is the only enzyme responsible for conversion of ribonucleoside diphosphate to deoxyribonucleotide diphosphate (dNDP), the key building blocks for DNA synthesis. This study will test the hypothesis that triapine is an effective radiation sensitizer which can be safely combined with peptide receptor radionuclide therapy and can improve antitumor activity of Lutetium Lu 177 Dotatate. Method: This study is an investigator initiated, NCI sponsored, multicenter phase 1 trial of triapine and Lutetium Lu 177 Dotatate in combination for well-differentiated somatostatin receptor-positive gastroenteropancreatic neuroendocrine tumor (GEP-NETs) after the failure of at least one line of prior systemic cancer treatment. A total of 29 patients will be enrolled in the dose escalation (BOIN design) and dose expansion cohorts. The study will be open through the entire ETCTN. Patients will be treated with 177 lutetium dotatate in combination with Triapene. Triapene will be administered orally from D1-14 with each dose of PRRT. Primary endpoint is to evaluate RP2D. Secondary endpoints are looking into clinical activity (ORR, PFS and OS). We are also evaluating NETEST, a novel blood based predicting as well as prognosticating test. In addition, the study will evaluate baseline somatostatin receptor density, somatic tumor mutations and germline mutations and correlate with clinical outcome. ClinicalTrials.gov Identifier: NCT04234568 Citation Format: Aman Chauhan, Charles Kunos, Riham El Khouli, Jill Kolesar, Heidi L. Weiss, Bill Carson, Mark Kidd, Jan Beumer, Susanne Arnold, Elise Kohn, Lowell Anthony. A phase I trial of Triapine and Lutetium Lu 177 Dotatate in combination for well-differentiated somatostatin receptor-positive gastroenteropancreatic neuroendocrine tumors (GEP-NETs) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 674.

Background: Current blood-based biomarkers for neuroendocrine neoplasms (NENs) lack both sensitivity and specificity. This is especially true for high-grade NENs (small and large cell neuroendocrine carcinomas). Human circulating progastrin (hPG80) is a novel biomarker for NENs and is easily measured in plasma using an ELISA test. This study is the first to explore hPG80 in NENs. Methods: Progastrin concentrations were quantified in plasma from 95 patients with stage IV NEN using DxPG80 technology (ECS Progastrin, Switzerland) and compared with hPG80 concentrations in 50-80-year-old healthy donors (n=252) as well as a subgroup comprised of only the 18-25-year-olds (n=137). Results: The median hPG80 in NENs patients was 5.54 pM as compared to 1.5 pM for patients in the 50-80-year-old control group and 0.29 pM for patients in the 18-25-year-old cohort (p Conclusions: Plasma hPG80 in NENs suggests hPG80 may be a diagnostic blood biomarker for both low- and high-grade NENs and further study is warranted. A prospective trial is ongoing in small cell lung cancer to evaluate its role in monitoring of disease (NCT03958045) and further prospective studies in low-grade neuroendocrine tumors are underway. Citation Format: Aman Chauhan, Alexandre Prieur, Jill Kolesar, Susanne Arnold, Thierry Cousin, Lea Payen, Younes Mahi, Madison Sanders, B. Mark Evers, Dominique Joubert, Lowell Anthony. hPG80 (circulating progastrin), a novel blood-based biomarker for detection of poorly differentiated neuroendocrine carcinoma and well differentiated neuroendocrine tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 735.

Patients with advanced-stage gastroenteropancreatic neuroendocrine tumors (GEP-NETs) have a poor overall prognosis despite chemotherapy and radiotherapy (e.g., peptide receptor radionuclide therapy (PRRT)). Better treatment options are needed to improve disease regression and patient survival. The purpose of this study was to examine a new treatment strategy by combining PI3K/mTOR dual inhibition and radiotherapy. First, we assessed the efficacy of two PI3K/mTOR dual inhibitors, PF-04691502 and PKI-402, to inhibit pAkt and increase apoptosis in NET cell lines (BON and QGP-1) and patient-derived tumor spheroids as single agents or combined with radiotherapy (XRT). Treatment with PF-04691502 decreased pAkt (Ser473) expression for up to 72 h compared with the control, in contrast, decreased pAkt expression was noted for less than 24 h with PKI-402. Simultaneous treatment with PF-04691502 and XRT did not induce apoptosis in NET cells, however, the addition of PF-04691502 48 h after XRT significantly increased apoptosis compared to PF-04691502 or XRT treatment alone. Our results demonstrate that schedule-dependent administration of a PI3K/mTOR inhibitor, combined with XRT, can enhance cytotoxicity by promoting the radiosensitivity of NET cells. Moreover, our findings suggest that radiotherapy, in combination with timed PI3K/mTOR inhibition, may be a promising therapeutic regimen for patients with GEP-NET.

Background: Peptide receptor radionuclide therapy (PRRT) is a novel radiotherapy approved for gastroenteropancreatic neuroendocrine tumors (GEP-NETs); however, PRRT has a relatively low treatment response rate of 20-30%. We hypothesize that addition of 3-aminopyridine-2-carboxaldehyde thiosemicarbazone (3-AP), a ribonucleotide reductase (RnR) inhibitor that inhibits the rate-limiting enzyme of DNA synthesis and repair, can improve radiotherapy outcomes. The purpose of this study is to determine the therapeutic rationale of RnR-targeted therapy and evaluate 3-AP as a radiosensitizer in treating GEP-NET in vitro and in vivo. Methods: We utilized two human GEP-NET cell lines: BON, a pancreatic carcinoid tumor, and QGP-1, a somatostatinoma. (i) Cells were treated with 3-AP for 96h and cell proliferation assays were performed using a sulforhodamine B (SRB) kit. Next, cells were treated with 3-AP for 24, 48, 72, 96h and cell cycle analyses were performed with Beckman Coulter cell cycle kit and quantified by flow cytometry. Expression of cell cycle arrest markers (i.e., pCdk-1, cyclin D1) was assessed by immunoblotting. (ii) To determine radiosensitivity in vitro, cells were treated with 3-AP for 16h, followed by X-ray radiotherapy (XRT), and incubation in drug-free medium. Cell survival was assessed by colony formation assay and quantified by SRB kit at 14d post-XRT. Apoptosis was measured at 72 and 96h post-XRT, and expression of apoptotic proteins (i.e., cleaved PARP and cleaved CASPASE 3) was examined by immunoblotting. (iii) To determine radiosensitivity in vivo, we randomized 20 nude mice bearing QGP-1 xenografts into four groups: control; 3-AP (10mg/Kg); XRT (2 Gy); and combination (XRT immediately followed by 3-AP); each mouse received 5 consecutive treatments. Results: (i) A dose of 2500nM 3-AP arrested cells before entering G2/M phase, significantly reducing percentage of G2/M cells from 8.4% to 0.5% at 72h. Concurrently, we observed increased expression of checkpoint proteins (pRb, pCdk1, pCdk2, pCdc2) and decreased expression of cell cycle inhibitory proteins (cyclin D1, p21, p27 Kip). (ii) Exposure of QGP-1 and BON cells to 3-AP prior to XRT resulted in a synergistic decrease (i.e., 85% and 90%, respectively) in clonogenicity. Furthermore, the combination of 3-AP and XRT increased apoptosis 5-fold and cleaved PARP expression in BON cells. (iii) A significant decrease in tumor size was noted in mice with XRT alone compared to control (146mm2 vs. 280 mm2, respectively); combination treatment (3-AP + XRT) demonstrated a further 48% decrease in tumor size compared to XRT alone. There was no significant decrease in tumor size with 3-AP treatment alone. Conclusion: 3-AP treatment prior to XRT therapy resulted in significant increase in apoptosis in vitro and decrease in tumor size in vivo, suggesting synergistic cytotoxicity and radiosensitivity. Selective RNR inhibition by 3-AP provides a potent therapeutic strategy as a radiosensitizer and offers a novel approach in treating advanced GEP-NETs. Citation Format: Zeta Chow, Jeremy Johnson, Aman Chauhan, Tadahide Isumi, Lowell Anthony, Courtney M. Townsend, B. Mark Evers, Piotr Rychahou. 3-Aminopyridine-2-carboxaldehyde thiosemicarbazone (3-AP): A promising radiochemotherapy for gastroenteropancreatic neuroendocrine tumors [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-014.

Background: Neuroendocrine tumors (NETs) are a heterogeneous group of neoplasms arising from multiple organs. Peptide Receptor Radionuclide Therapy (PRRT) is a novel therapy approved for treating NETs with a treatment response rate of 20-30%. To improve therapeutic response of PRRT, 3-AP (Triapine), which is a ribonucleotide reductase (RR) inhibitor with antineoplastic and radiation sensitizing properties, can be used in combination with PRRT. The purpose of our study is to examine effects of Triapine treatment on cellular proliferation, apoptosis and activation of DNA repair pathway in NET cells in vitro. Methods: For these studies, we utilized two human NET cell lines: BON, a pancreatic carcinoid tumor, and QGP-1, a somatostatinoma. (i) To determine effective dose range of Triapine (500 - 2500nM), proliferation was measured at 48, 72, and 96 h with sulforhodamine B (SRB) assay. (ii) To evaluate the effects of radiosensitivity, cultures were treated with increasing doses (250 - 5,000nM), received 2 Gy radiation (XRT) 1h later, exposed to Triapine for 24 hours, rinsed, and fed Triapine-free media; colonies were counted and quantified with SRB assays 14 days later and survival curves were generated. (iii) To further characterize cellular responses, cells were treated with Triapine (250 - 5,000 nM), irradiated 1h later, and harvested 24 h later to examine expression levels of Cyclin D1, cleaved PARP, p-DNA-PK (S2056), p-ATM (S1981) and p-ATR (S428) by Western blot. Results: (i) SRB proliferation assay demonstrated the effective dose range, determined by half maximal inhibitory concentration (IC-50), was 1,3 µM in BON cells and 1.73 µM in QGP-1 cells at 96 h. (ii) Exposure to Triapine before irradiation resulted in a significant increase in the radiosensitivity of each of the tumor cell lines with dose enhancement factors at a surviving fraction of 0.10 of 1.8 and 1.6 for BON and QGP-1, respectively. (iii) Triapine treatment alone activated DNA repair pathway in a dose-dependent fashion as determined by pDNA-PK (S2056) expression. Increasing p-ATM (S1981) expression was observed in a dose-dependent fashion in QGP-1 cells, but not in BON cells. In addition, p-ATR (S428) expression was not significantly different in either cell line. (iv) Triapine treatment alone or in combination with 2 and 4 Gy XRT resulted in dose-dependent decrease in cyclin D1 expression in both cell lines, however, no significant increase in cleaved PARP expression at 24 and 48h. Conclusion: Our results demonstrated rapid and dose dependent decrease in cyclin D1 expression after Triapine treatment, dose-dependent activation of DNA-PK, the key component of the non-homologous end joining pathway and significant increase in the radiosensitivity of each of the tumor cell lines. In conclusion, our findings provide a rationale for inclusion of Triapine as a radiosensitizer in combination with PRRT in treating NETs. Citation Format: Zeta Chow, Aman Chauhan, Lowell Anthony, Courtney Townsend, B. Mark Evers, Piotr Rychahou. Effect of 3AP on neuroendocrine tumor cell proliferation, apoptosis and activation of DNA repair pathway [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6269.

Despite significant advances in the understanding and managing gastroenteropancreatic neuroendocrine tumors (GEPNETs), the therapeutic success of treatments remains relatively poor. Recent FDA approval of Peptide Receptor Radionuclide Therapy (PRRT) has paved the way for radiation-based treatment for GEPNETs. PRRT delivers high doses of radiation to tumor cells in the body to destroy cells or slow their growth. The DNA-dependent protein kinase (DNA-PK) complex plays a pivotal role in non-homologous end-joining (NHEJ) repair after radiation therapy. A novel, clinical-stage DNA-PK inhibitor, M3814, potently and selectively blocks the NHEJ repair pathway for DNA double strand breaks (DSB). Here, we investigated the feasibility of radiosensitizing neuroendocrine (NET) cells with the DNA-PK inhibitor M3814, both in vitro and in preclinical NET models. Methods: 96-well clonogenic assays were used to evaluate efficacy of M3814 in combination with radiation therapy (XRT) in vitro in QGP-1 and BON pancreatic NET cell lines. DSB-induced DNA repair was visualized with confocal laser scanning microscopy using γ-H2AX immunofluorescence after M3814 treatment. Western blot was used to confirm activation of DNA-PKcs (pDNA-PKcs S2056) after M3814 treatment. The efficacy of M3814 in combination with XRT was evaluated in QGP-1 and BON human xenograft models in athymic nude mice. Tumor cells were injected subcutaneously into athymic nude mice, and treatment started when palpable tumors (∼200 mm3) were established. M3814 was given orally (100 mg/kg) 30 min prior to XRT, which was applied using a radiation therapy device calibrated to deliver 2 Gy daily for 4 d. Results: Clonogenic assays demonstrated lack of cytotoxicity of M3814 alone at doses below 1000 nM and a strong radiosensitizing effect at 500 nM in both QGP-1 and BON cell lines. Immunofluorescence analysis revealed high numbers of γH2AX foci, marking unrepaired DSBs inside the micronuclei. Western blot showed that activation of DNA-PKcs, induced by XRT treatment, was quickly suppressed by M3814 treatment. In combination with XRT, M3814 showed efficacy in both QGP-1 and BON xenografts as demonstrated by a strong potentiation of the effect of XRT. In all models, a dose of 2 Gy administered daily for 4 d, in combination with M3814, induced a significant tumor growth inhibition compared to XRT alone. M3814, alone or in combination with XRT, did not induce significant weight loss or visual signs of toxicity in the mice in any of the studies. Conclusions: Our results demonstrate a benefit of adding M3814 to XRT in NET treatment. Selective DNA-PK inhibition by M3814 provides a potent therapeutic strategy for disruption of NHEJ DSB repair and may offers a novel therapeutic approach in advanced NET. Citation Format: Piotr Rychahou, Aman Chauhan, Zeta Chow, Tadahidi Izumi, Quan Chen, Eun Y. Lee, Dana Napier, Lowell B. Anthony, Michael J. Cavnar, Charles Kunos, B. Mark Evers. DNA-PK inhibitor, M3814, as a radiation sensitizer in the treatment of neuroendocrine tumors [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6402.

e15038 Background: Neuroendocrine neoplasms (NENs) are heterogeneous tumors which originate from various organs and are of variable aggressiveness based on grade and morphology. Current biomarkers for NENs lack sensitivity and specificity, especially for high-grade NENs (small and large cell neuroendocrine carcinomas). hPG80, progastrin, is a novel bio-marker which is easily measured in plasma using an ELISA test. Physiologically, hPG80, an 80 amino acid protein, is the precursor of the gastrointestinal hormone gastrin. It is synthetized by gastric antrum G cells, and then processed into gastrin by multiple enzymatic processes. In pathological conditions, the GAST gene, which encodes hPG80, was shown to be over-expressed in human solid tumors from various primary sites. hPG80 is unprocessed and released from the tumor cells and becomes detectable in the blood. This study is the first to explore hPG80 in NENs. Methods: hPG80 was quantified in the plasma from 31 NEN patients using DxPG80 technology (ECS-Progastrin, Switzerland). Additional 69 samples are currently undergoing analysis. Progastrin concentrations in 18-70 YO (n = 557) and 18-25 YO (n = 137) healthy blood donors were compared to 31 stage IV NENs patients. The study was IRB approved. Results: Current data are for 31 patients. Data on total 100 patients will be presented at ASCO 2020. Mean age of study cohort at the time of blood collection was 60.9 years. 21 patients had grade 1 and 2 well differentiated NET. 10 patients had high grade NEN (Small cell, large cell and poorly differentiated NEC). High grade sub cohort also included two well differentiated grade 3 NET patients. Mean hPG80 in NENs was 14.17 pM as compared to 2.04 pM and 0.99 pM in 18-70 and 18-25 YO control groups (p < 0.0001), respectively. Subgroup analysis of NENs revealed mean hPG80 of 24.61 pM in high-grade NENs (n = 10) vs 10.88 pM in G1/2 NETs (n = 21). Conclusions: This first-ever study of plasma hPG80 in NENs suggests hPG80 may be a diagnostic blood-based biomarker in both low and high-grade NENs and further study is warranted. A prospective trial is ongoing in high-grade NEN to evaluate its role in monitoring of disease (NCT03958045) and further studies in low-grade NETs are underway. This research was supported by Cancer Center Support Grant (CCSG) from the National Cancer Institute (P30 CA177558) and ECS Progastrin.

TPS4660 Background: Radiolabeled somatostatin analogues provide a means of delivering targeted radiation with a high therapeutic index to NETs that express somatostatin receptors (SSTRs). Radiolabeled somatostatin analogue Lutetium Lu 177 Dotatate (Lutathera) is a beta-emitting radionuclide, recently FDA approved for use in SSTR positive gastroenteropancreatic neuroendocrine tumors (GEPNETS) in the US based on the NETTER-1 Phase III trial. Despite favorable PFS and safety profile, the drug has limited cytoreductive capability with a 17% ORR. We hypothesize that addition of an effective radiation sensitizer could help improve antitumor activity of Lutathera. Ribonucleotide reductase (RNR) is the only enzyme responsible for conversion of ribonucleoside diphosphate to deoxyribonucleotide diphosphate (dNDP), the key building blocks for DNA synthesis. Radiation is a potent inducer of DNA double-strand breaks (DSBs), and RNR is the rate-limiting enzyme in the repair of DNA in this setting. Triapine is an inhibitor of RNR. This study will test the hypothesis that radiation sensitizer triapine can be safely combined with peptide receptor radionuclide therapy and ultimately may improve antitumor activity of Lutetium Lu 177 Dotatate. Methods: This study is an investigator initiated, NCI sponsored, multicenter phase 1 trial of triapine and Lutetium Lu 177 Dotatate in well-differentiated somatostatin receptor-positive gastroenteropancreatic neuroendocrine tumor (GEP-NETs) after the failure of at least one line of prior systemic cancer treatment. A total of 29 patients will be enrolled in the dose escalation with help of Bayesian optimal interval design (BOIN) and dose expansion cohorts. The study will be open through the NCI ETCTN (National Cancer Institute Experimental Therapeutics Clinical Trials Network) program. Patients will be treated with 177 lutetium dotatate in combination with triapine. Triapine will be administered orally (100 mg once a day starting dose) from D1-14 with each dose of PRRT [200 mCi]. Primary endpoint is to evaluate recommended phase II dose (RP2D). Secondary endpoints are to evaluate safety, pharmacokinetics, and clinical activity (ORR and PFS). We are also evaluating NETEST, a novel blood based test that evaluates levels of 51 neuroendocrine tumor gene transcripts. In addition, the study will correlate clinical outcome with baseline somatostatin receptor density, somatic tumor mutations and germline mutations. Clinical trial information: 04234568 .

e24107 Background: Gastrointestinal toxicity is a common side effect of cancer therapy. Pre-clinical studies using a proprietary mixture of amino acids, demonstrated a reduction in mucositis and gastrointestinal toxicity following irradiation via tightening of the mucosal barrier, increasing proliferation of crypt cells, increasing villous height and increasing absorption of fluid, electrolytes and nutrients. A novel 5-amino acid mixture “enterade” was previously evaluated in a Phase II clinical trial for anti-diarrheal effects in post bone marrow transplant patients at Dana Farber Cancer Institute (NCT02919670). Signs of anti-diarrheal efficacy were found in treatment-compliant patients.Enterade is currently being tested in two prospective Phase II studies (NCT03722511 and NCT04073017) in neuroendocrine tumor patients with quality of life limiting diarrhea. We conducted a pilot study to evaluate early antidiarrheal signals in oncologic treatment-induced diarrhea. Methods: An IRB-approved retrospective chart review was conducted at Markey Cancer Center between Sept 2019 and Dec 2019. Medical records were retrospectively reviewed for all solid tumor patients who received enterade for chemotherapy or immunotherapy-induced diarrhea. Patients were instructed to consume one 8-oz bottle of enterade twice a day for at least one week in addition to standard of care antidiarrheal medications. Results: A total of 46 patients were offered enterade. Antidiarrheal efficacy data was available on 17 patients. 11 were female and the mean age of the cohort was 64 years. Six out of 17 patients had a previous history of bowel resection. Four patients had immune checkpoint induced colitis. Three patients developed diarrhea from TKI use and rest were on cytotoxic chemotherapy at the time of the diarrheal episode. Eighty-eight percent (15/17) of patients reported a reduction in diarrhea frequency after consuming enterade. On average, patients noticed an improvement in diarrhea frequency after 3.6 days of enterade consumption, with 86% (13/15) of responders reporting at least 50% reduction in diarrhea frequency. Conclusions: Patient-reported diarrheal improvement after enterade consumption is consistent with pre-clinical data. Results suggest the amino acid mixture may alleviate symptoms of cancer therapy-induced gastrointestinal toxicity (i.e. diarrhea). A prospective clinical trial is warranted to substantiate the observed improvement in chemotherapy and immunotherapy induced diarrhea.

614 Background: Diarrhea is a common quality of life limiting symptom seen in neuroendocrine tumor patient. Diarrhea in these patients could be due to excessive serotonin production, secondary to post-operative short gut syndrome, steatorrhea from somatostatin analogs, bile acid colitis or intestinal bacterial overgrowth. In this study we summarize our single center experience with enterade. Methods: Medical records of all the NET patients treated with enterade for symptomatic diarrhea were retrospectively reviewed after appropriate IRB approval. Patients were treated at Markey cancer center between May 2017-June 2019. Results: Total 98 patients were offered enterade. enterade was instructed to be taken as one 8 Oz bottle BID for 1 week. Antidiarrheal efficacy data was available on 49 patients at the time of abstract submission. 37 (75%) patients had GEPNETs, 8 had bronchial NETs, 1 gynecological NET and 3 patients had unknown primary. 28 (57%) patients had history of prior bowel resection either for primary neuroendocrine tumor resection or debulking. 28 (57%) patients were on somatostatin analog at the time of initiation of enterade. 42 (85%) out of 49 patients reported subjective improvement in diarrheal symptoms. 27 (64%) out of these 42 responders reported at least 50 percent reduction in diarrhea frequency. Conclusions: 85% (42/49) neuroendocrine tumor patients reported improvement in diarrhea with enterade. 64% (27/42) reported more than 50% reduction in diarrhea frequency. A prospective Phase II study of enterade in neuroendocrine tumor patients with quality of life limiting diarrhea is currently being conducted (NCT03722511).

632 Background: Incidence and prevalence of neuroendocrine tumors (NETs) are on the rise. However, with the increasing burden of the disease, there is a need to understand financial toxicity associated with NET management. Especially the out of pocket spending or expenses made by patients and family members, which affects the management and treatment adherence. We report results of our prospective online survey based study looking into determinants and the burden of financial toxicity in NET patients. Methods: We collaborated with NCAN (Neuroendocrine Cancer Awareness Network) a national NET patient support group for our prospective online questionnaire-based survey. After IRB approval, NCAN disseminated online link to the survey on their webpage. The survey had twenty-one questions with the goal of at least 200 complete responses. Results: We received 215 responses before we decided to stop the survey for analysis. The average age of the respondents was 58.4 years. 75% of respondents were females with most repsondents of the survey (37%) being graduates. About 41% of respondents had small bowel NET while 27% had pancreas NET as their NET diagnosis. The median income of the respondents was $65000/year. Median oncology visits were reported to be 4 per year. Median co-pay for the NET specialist visit was found to be $30. The median spending on travel for the physician`s appointment was $250 per visit to see NET specialists. Median monthly insurance premium was $300. Study participants reported spending $75 out of pocket on NET related care every month on top of insurance premiums. More than half of the respondents were on Somatostatin analogues only as their main treatment. About 21% mentioned that they were not able to get required treatment due to cost related issues. Conclusions: The preliminary analysis of the survey gave us an insight into financial toxicity related to NET management. Although most NETs are indolent, the chronic nature of disease and associated, symptom management is often time and resource-intensive. Many patients mentioned having issues with the coverage of insurance and having to travel a long distance to see a NET specialist, which demonstrate a lack of access to NET specialists.