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Suppl Figure 3. Immunoblot of Effector Proteins and Markers of Apoptosis in Melanoma Cell Lines

Suppl Figure 2. Immunoblot Analysis of Effector Proteins in Melanoma Cell Lines

Suppl Figure 5. Immunoblot of Effector Proteins in Tumors from PDTX Mice Treated with TAK-733

Suppl Table 1. IC50 Value Matrix

Suppl Figure 4. Representative PDTX Model MB1255 Treated with Vehicle or TAK-733

Suppl Table 2. TAK-733 PD/ Conc. Summary, A375 tumor bearing nude mouse

Suppl Table 4. Antitumor Activity of Orally Administered TAK-733 Against A375 Melanoma Tumor Xenografts in Female Athymic Nude Mice

Suppl Figure 1. Chemical structure and molecular data for TAK-733

Suppl Figure 6. Genomic Analyses of PDTX MB1374 Treated to Resistance

Supplementary Figure Legends

The goal of this study was to investigate the activity of the selective MEK1/2 inhibitor TAK-733 in both melanoma cell lines and patient-derived melanoma xenograft models. In vitro cell proliferation assays using the sulforhodamine B assay were conducted to determine TAK-733 potency and melanoma responsiveness. In vivo murine modeling with eleven patient-derived melanoma explants evaluated daily dosing of TAK-733 at 25 or 10 mg/kg. Immunoblotting was performed to evaluate on-target activity and downstream inhibition by TAK-733 in both in vitro and in vivo studies. TAK-733 demonstrated broad activity in most melanoma cell lines with relative resistance observed at IC50 > 0.1 μmol/L in vitro. TAK-733 also exhibited activity in 10 out of 11 patient-derived explants with tumor growth inhibition ranging from 0% to 100% (P < 0.001–0.03). Interestingly, BRAFV600E and NRAS mutational status did not correlate with responsiveness to TAK-733. Pharmacodynamically, pERK was suppressed in sensitive cell lines and tumor explants, confirming TAK-733–mediated inhibition of MEK1/2, although the demonstration of similar effects in the relatively resistant cell lines and tumor explants suggests that escape pathways are contributing to melanoma survival and proliferation. These data demonstrate that TAK-733 exhibits robust tumor growth inhibition and regression against human melanoma cell lines and patient-derived xenograft models, suggesting that further clinical development in melanoma is of scientific interest. Particularly interesting is the activity in BRAF wild-type models, where current approved therapy such as vemurafenib has been reported not to be active. Mol Cancer Ther; 14(2); 317–25. ©2014 AACR.

Purpose:The purpose of this study was to evaluate the rational combination of TORC1/2 inhibitor TAK-228 and Aurora A kinase inhibitor alisertib in preclinical models of triple-negative breast cancer (TNBC) and to conduct a phase I dose escalation trial in patients with advanced solid tumors.Experimental Design:TNBC cell lines and patient-derived xenograft (PDX) models were treated with alisertib, TAK-228, or the combination and evaluated for changes in proliferation, cell cycle, mTOR pathway modulation, and terminal cellular fate, including apoptosis and senescence. A phase I clinical trial was conducted in patients with advanced solid tumors treated with escalating doses of alisertib and TAK-228 using a 3+3 design to determine the maximum tolerated dose (MTD).Results:The combination of TAK-228 and alisertib resulted in decreased proliferation and cell-cycle arrest in TNBC cell lines. Treatment of TNBC PDX models resulted in significant tumor growth inhibition and increased apoptosis with the combination. In the phase I dose escalation study, 18 patients with refractory solid tumors were enrolled. The MTD was alisertib 30 mg b.i.d. days 1 to 7 of a 21-day cycle and TAK-228 2 mg daily, continuous dosing. The most common treatment-related adverse events were neutropenia, fatigue, nausea, rash, mucositis, and alopecia.Conclusions:The addition of TAK-228 to alisertib potentiates the antitumor activity of alisertib in vivo, resulting in increased cell death and apoptosis. The combination is tolerable in patients with advanced solid tumors and should be evaluated further in expansion cohorts with additional pharmacodynamic assessment.

Pharmacodynamic effects of alisertib and TAK-228 in patient tissue samples

Cardiac surgery with tricuspid valve and potentially pulmonic valve replacement at an experienced center is currently the most effective strategy available for the treatment of carcinoid heart disease. Cardiac surgery for carcinoid heart disease requires a multidisciplinary team including cardiology, medical oncology, cardiothoracic anesthesia, and cardiac surgery. Without cardiac surgery, morbidity and mortality from carcinoid heart disease is high. Aggressive management of carcinoid before and after cardiac surgery is critical. Over time, though, circulating carcinoid hormones can lead to destruction of prosthetic valves as well, resulting in recurrent right heart failure. Percutaneous options for valve repair may be on the horizon for management of carcinoid heart disease.

This study aimed to investigate safety and efficacy of silmitasertib, an oral small molecule casein kinase 2 inhibitor, plus gemcitabine and cisplatin (G+C) versus G+C in locally advanced/metastatic cholangiocarcinoma.This work is a Phase 1b/2 study (S4-13-001). In Phase 2, patients received silmitasertib 1000 mg twice daily for 10 days with G+C on Days 1 and 8 of a 21-day cycle. Primary efficacy endpoint was progression-free survival (PFS) in the modified intent-to-treat population (defined as patients who completed at least one cycle of silmitasertib without dose interruption/reduction) from both phases (silmitasertib/G+C n = 55, G+C n = 29). The response was assessed by Response Evaluation Criteria in Solid Tumors v1.1. The median PFS was 11.2 months (95% confidence interval [CI], 7.6, 14.7) versus 5.8 months (95% CI, 3.1, not evaluable [NE]) (p = 0.0496); 10-month PFS was 56.1% (95% CI, 38.8%, 70.2%) versus 22.2% (95% CI, 1.8%, 56.7%); and median overall survival was 17.4 months (95% CI, 13.4, 25.7) versus 14.9 months (95% CI, 9.9, NE) with silmitasertib/G+C versus G+C. Overall response rate was 34.0% versus 30.8%; the disease control rate was 86.0% versus 88.5% with silmitasertib/G+C versus G+C. Almost all silmitasertib/G+C (99%) and G+C (93%) patients reported at least one treatment emergent adverse event (TEAE). The most common TEAEs (all grades) with silmitasertib/G+C versus G+C were diarrhea (70% versus 13%), nausea (59% vs. 30%), fatigue (47% vs. 47%), vomiting (39% vs. 7%), and anemia (39% vs. 30%). Twelve patients (10%) discontinued treatment because of TEAEs during the study.Silmitasertib/G+C demonstrated promising preliminary evidence of efficacy for the first-line treatment of patients with locally advanced/metastatic cholangiocarcinoma.

TPS257 Background: Refractory microsatellite stable colorectal cancer (MSS CRC) is immunologically cold and single-agent anti-PD-1/PD-L1 drugs are ineffective; novel immune-based approaches are needed. Evorpacept (E, ALX148) is an engineered protein (high-affinity CD47-blocker fused to an inactive IgG Fc region), which blocks the CD47/SIRPα innate immune inhibitory phagocytosis checkpoint expressed on CRC and phagocytes, respectively. The Fc region of E does not bind to Fcγ receptors, thereby limiting hematologic toxicity, and is intended to be given in combination. In CT26 CRC syngeneic models, E ± anti-PD-1 monoclonal antibody decreases tumor growth, reduces myeloid immunosuppression, increases dendritic cell activation, and increases T cell activation (Kauder, 2018); E enhances the antibody-dependent cellular phagocytosis activity of cetuximab (C) in vitro (Kauder, 2018); and E + pembrolizumab (P) was well-tolerated in the first-in-human trial (Lakhani, 2021). Methods: AGICC-ALX148 21CRC01 (NCT05167409) is a phase 2, single-arm, multicenter, investigator-initiated trial of E (15 mg/kg weekly), C (400 mg/m2 then 250 mg/m2 weekly), and P (200 mg every 3 weeks) in 21-day cycles for patients with unresectable MSS/proficient mismatch repair CRC refractory to oxaliplatin, irinotecan, and a fluoropyrimidine, regardless of tumor sidedness and RAS/BRAF status. Additional key eligibility criteria include ECOG performance status 0-1, evaluable disease per RECIST v1.1, adequate hematologic and end organ function, absence of prior checkpoint inhibitor use, and absence of significant autoimmune disease. Six patients will be enrolled in Stage 1 (safety run-in) and treated with ECP. The study will proceed to Stage 2 (dose expansion, N = 42, and all treated with ECP) if less than 33% of patients in Stage 1 experience a dose-limiting toxicity. Otherwise, additional patients will be enrolled in Stage 1 at lower dose level(s). The co-primary objectives are to determine 1) the recommended dose of E with CP, and 2) objective response rate by RECIST v1.1 (by one-sided exact test with α = 0.05, H0 p ≤ 3% [historical controls], HA p ≥ 15%; power is 87%). The study will close for futility if there are no responses (partial or complete) in the first 24 evaluable patients (by MinMax design with α = 0.025 [1-sided]; power is 87%). Secondary and exploratory aims include determination of progression-free survival, overall survival, safety, response assessment by iRECIST, and blood- and tumor-based immune modulation and baseline tumor expression (PD-L1, EGFR, and CD47) for association with tumor response. The study is open through the Academic GI Cancer Consortium and 5 patients have been enrolled at time of submission. Clinical trial information: NCT05167409 .

85 Background: Rising rates of early-onset colorectal cancer (EOCRC) pose a dilemma for clinicians when deciding how to treat early-stage patients to maximize outcomes. While standard-of-care for Stage II colon cancer is largely surgical resection, evidence suggests that treatment selection may differ by patient age. We sought to determine whether rates of adjuvant chemotherapy administration differ between early and later-onset patients with Stage II CRC. Methods: Data were derived from the Flatiron National Database spanning 1/1/2003 to 8/1/2021. Patients 18 years or older with Stage II CRC were grouped into those aged 18-49 (EOCRC) and those aged 50 or older (LOCRC). Demographic characteristics, ECOG score, stage and site of tumor, and chemotherapy were included for all patients. Primary outcomes of interest included rates of adjuvant chemotherapy administration by age and ethnicity. Univariate and multivariable logistic regression models were used to evaluate relationships between chemotherapy administration, age groups, and ethnicity while adjusting for significant covariates. Results: Of 2133 patients with Stage II CRC, 1606 patients with complete data were included. A secondary analysis of 1065 patients with colon cancer was performed to address potential confounding factors related to neoadjuvant and/or adjuvant chemotherapy given in patients with stage II rectal cancer. Mean age of EOCRC patients was 45.0 years (range: 41.0-48.0) vs. 68.0 years (60.0-75.0) for LOCRC. Adjusting for ethnicity, gender, site, and ECOG score, multivariate analysis showed EOCRC patients received chemotherapy significantly more often than LOCRC patients for stage II CRC (adjusted odds ratio 1.85, 95% CI 1.32-2.60, p < 0.001). Similar findings were observed in the colon cancer only cohort (adjusted OR 2.02, 95% CI 1.31-3.09, p < 0.001). By ethnicity, non-Hispanic patients received chemotherapy at significantly lower rates than Hispanic patients in both cohorts (adjusted odds ratio 0.58, 95% CI 0.39-0.88, p = 0.009 and adjusted odds ratio 0.55, 95% CI 0.34-0.91, p = 0.018). In a subgroup analysis of Stage IIA patients, multivariate logistic regression adjusting for gender, ECOG, site, and ethnicity showed that patients with EOCRC were more likely to receive chemotherapy than patients with LOCRC (adjusted odds ratio 1.91, 95% CI 1.21-2.99, p = 0.005). Updated survival data will be presented. Conclusions: Adjuvant chemotherapy is given preferentially in Stage II EOCRC, even in Stage IIA disease, despite deviation from established guidelines. This may expose patients at low risk for recurrence to unnecessary toxicities and reveals potential provider bias in favor of younger patients in aggressively treating CRC, despite unclear evidence for any outcome benefit.

BACKGROUND: MEK inhibitors have limited activity in biliary tract cancers (BTCs) as monotherapy but are hypothesized to enhance responses to programmed death ligand 1 (PD-L1) inhibition. METHODS: This open-label phase II study randomized patients with BTC to atezolizumab (anti–PD-L1) as monotherapy or in combination with cobimetinib (MEK inhibitor). Eligible patients had unresectable BTC with 1 to 2 lines of prior therapy in the metastatic setting, measurable disease, and Eastern Cooperative Oncology Group (ECOG) performance status less than or equal to 1. The primary endpoint was progression-free survival (PFS). RESULTS: Seventy-seven patients were randomized and received study therapy. The trial met its primary endpoint, with a median PFS of 3.65 months in the combination arm versus 1.87 months in the monotherapy arm (HR 0.58, 90% CI 0.35–0.93, 1-tail P = 0.027). One patient in the combination arm (3.3%) and 1 patient in the monotherapy arm (2.8%) had a partial response. Combination therapy was associated with more rash, gastrointestinal events, CPK elevations, and thrombocytopenia. Exploratory analysis of tumor biopsies revealed enhanced expression of antigen processing and presentation genes and an increase in CD8/FoxP3 ratios with combination treatment. Patients with higher baseline or lower fold changes in expression of certain inhibitory ligands (LAG3, BTLA, VISTA) on circulating T cells had evidence of greater clinical benefit from the combination. CONCLUSION: The combination of atezolizumab plus cobimetinib prolonged PFS as compared with atezolizumab monotherapy, but the low response rate in both arms highlights the immune-resistant nature of BTCs. TRIAL REGISTRATION: ClinicalTrials.gov NCT03201458. FUNDING: National Cancer Institute (NCI) Experimental Therapeutics Clinical Trials Network (ETCTN); F. Hoffmann-La Roche, Ltd.; NCI, NIH (R01 CA228414-01 and UM1CA186691); NCI’s Specialized Program of Research Excellence (SPORE) in Gastrointestinal Cancers (P50 CA062924); NIH Center Core Grant (P30 CA006973); and the Passano Foundation.

Background AZD0156 is an oral inhibitor of ATM, a serine threonine kinase that plays a key role in DNA damage response (DDR) associated with double-strand breaks. Topoisomerase-I inhibitor irinotecan is used clinically to treat colorectal cancer (CRC), often in combination with 5-fluorouracil (5FU). AZD0156 in combination with irinotecan and 5FU was evaluated in preclinical models of CRC to determine whether low doses of AZD0156 enhance the cytotoxicity of irinotecan in chemotherapy regimens used in the clinic. Methods Anti-proliferative effects of single-agent AZD0156, the active metabolite of irinotecan (SN38), and combination therapy were evaluated in 12 CRC cell lines. Additional assessment with clonogenic assay, cell cycle analysis, and immunoblotting were performed in 4 selected cell lines. Four colorectal cancer patient derived xenograft (PDX) models were treated with AZD0156, irinotecan, or 5FU alone and in combination for assessment of tumor growth inhibition (TGI). Immunofluorescence was performed on tumor tissues. The DDR mutation profile was compared across in vitro and in vivo models. Results Enhanced effects on cellular proliferation and regrowth were observed with the combination of AZD0156 and SN38 in select models. In cell cycle analysis of these models, increased G2/M arrest was observed with combination treatment over either single agent. Immunoblotting results suggest an increase in DDR associated with irinotecan therapy, with a reduced effect noted when combined with AZD0156, which is more pronounced in some models. Increased TGI was observed with the combination of AZD0156 and irinotecan as compared to single-agent therapy in some PDX models. The DDR mutation profile was variable across models. Conclusions AZD0156 and irinotecan provide a rational and active combination in preclinical colorectal cancer models. Variability across in vivo and in vitro results may be related to the variable DDR mutation profiles of the models evaluated. Further understanding of the implications of individual DDR mutation profiles may help better identify patients more likely to benefit from treatment with the combination of AZD0156 and irinotecan in the clinical setting.

Background: FOLFOX (folinic acid, 5-fluorouracil and oxaliplatin) is the most commonly used chemotherapy regimen for the treatment of colorectal cancer. FOLFOX is administered in 14-day cycles, though toxicities frequently lead to unplanned delays. We report the incidence of unplanned delays among patients receiving FOLFOX and describe the reasons for delays. Methods: We conducted a retrospective analysis of patients receiving FOLFOX chemotherapy for colorectal cancer. Patients were treated at one of two tertiary cancer centers between January 2012 and April 2016. Cycles 2–6 were assessed for delays, and treatments were considered delayed when the interval from prior treatment was >18 days. Reasons for unplanned delays were categorized based on review of clinical records. Results: We identified 214 patients receiving FOLFOX as standard-of-care therapy. The median age was 59 years, and 55% were female. Of 961 evaluable treatment cycles, 124 (13%) had unplanned delays, and 92 of 214 patients (43%) had one or more unplanned delays in cycles 2–6. Cytopenias (neutropenia and/or thrombocytopenia) were the most common cause of unplanned delays, affecting 34% of patients and accounting for 74 of 124 unplanned delays (60%). Conclusions: Delays are common during FOLFOX chemotherapy, with 43% of patients having at least one unplanned delay prior to completing cycle 6. Neutropenia and thrombocytopenia were the leading causes of unplanned delays. Our findings justify the development of systematic approaches for preventing unplanned delays, such as standardized laboratory treatment criteria and/or proactive dose adjustment strategies.

SummaryVantictumab is a fully human monoclonal antibody that inhibits Wnt pathway signaling through binding FZD1, 2, 5, 7, and 8 receptors. This phase Ib study evaluated vantictumab in combination with nab-paclitaxel and gemcitabine in patients with untreated metastatic pancreatic adenocarcinoma. Patients received vantictumab at escalating doses in combination with standard dosing of nab-paclitaxel and gemcitabine according to a 3 + 3 design. A total of 31 patients were treated in 5 dosing cohorts. Fragility fractures attributed to vantictumab occurred in 2 patients in Cohort 2 (7 mg/kg every 2 weeks), and this maximum administered dose (MAD) on study was considered unsafe. The dosing schedule was revised to every 4 weeks for Cohorts 3 through 5, with additional bone safety parameters added. Sequential dosing of vantictumab followed by nab-paclitaxel and gemcitabine was also explored. No fragility fractures attributed to vantictumab occurred in these cohorts; pathologic fracture not attributed to vantictumab was documented in 2 patients. The study was ultimately terminated due to concerns around bone-related safety, and thus the maximum tolerated dose (MTD) of the combination was not determined. The MAD of vantictumab according to the revised dosing schedule was 5 mg/kg (n = 16).

Background: At our institution, the standard treatment recommendation for esophageal cancer patients with stage IB–IIIB disease is for neoadjuvant chemoradiation per the CROSS regimen prior to surgery. This regimen can be difficult for patients to tolerate, and they may be unable to receive full dose therapy without treatment dose reductions and delays. Methods: We conducted a quality improvement (QI) project, STRENGTH (Seeking to Reactivate Esophageal and Gastric Treatment Health), to implement supportive care interventions in the prehabilitation phase of neoadjuvant treatment. Our QI program included a standardized chemotherapy order template with supportive care interventions implemented at specific time points. Following implementation of the STRENGTH pathway, a retrospective QI analysis assessed an equal number of patients in the pre-STRENGTH and STRENGTH group for chemotherapy and radiation therapy dose intensities, as well as treatment outcomes. Results: During the pre-STRENGTH period, patients received an average of 5 chemotherapy treatments (range, 2–6), with an average relative dose intensity of 91.8% for carboplatin and 86.7% for paclitaxel. During the STRENGTH period, patients received an average of 6 (range, 5–8) chemotherapy treatments, with an average relative dose intensity of 111.4% for carboplatin and 112.9% for paclitaxel. In the pre-STRENGTH group, one patient did not complete their planned radiation dose due to nausea, vomiting, and dehydration. All patients in the STRENGTH group received their planned radiation dose. In the STRENGTH group, there is a trend of improved pathologic response, longer progression-free survival, and shortened time to surgery. Conclusion: Implementation of the STRENGTH pathway improved chemotherapy dose intensity, with potentially improved oncologic outcomes in the STRENGTH group. We plan to further optimize the STRENGTH program with implementation of standardized dose reduction and delay protocols for both chemotherapy and radiation, and assess the effects of STRENGTH interventions on patient quality of life.

3112 Background: In prior work, senescence and up-regulation of genes in the PI3K/AKT/mTor pathway were observed in patient-derived xenograft models treated with alisertib to resistance, and tumor growth inhibition was observed when MLN0128 (sapanisertib) was added to alisertib. In a previously reported dose escalation cohort of patients with advanced solid tumors treated with the combination of alisertib and MLN0128, the maximum tolerated dose (MTD) was alisertib 30mg BID days 1-7 of a 21-day cycle and MLN0128 2mg daily on a continuous schedule. Presented here are final results from the dose expansion portion of this clinical trial. Methods: Three cohorts of patients were treated with the combination at the MTD. Patients with advanced solid tumors, refractory to standard therapy, were assigned to either single-agent treatment with alisertib (Group 1) or MLN0128 (Group 2) on days 1-7 of Cycle 1. For the remainder of the study, patients received combination treatment. Group 3 enrolled patients with refractory pancreatic adenocarcinoma who were treated with standard dosing of the combination. Biopsies were performed in Groups 1 and 2 prior to treatment initiation and after both the single-agent lead-in and 7 days of combination treatment, with assessment of pharmacodynamic markers. Functional imaging was performed pre-treatment and after Cycle 1. Results: A total of 31 patients with refractory cancers were treated. Group 1 included patients with breast (5), colorectal (2), ovarian (2), and pancreatic (1) cancers. Group 2 included patients with breast (4), colorectal (2), pancreatic (2), uterine (1), and kidney (1) cancers. Eleven patients with refractory pancreatic cancer were treated in Group 3. Median time on study was 11.6 weeks in Group 1, 6 weeks in Group 2, and 9 weeks in Group 3. One partial response was documented in Group 1. One patient with pancreatic cancer in Group 1 continued on study for 47 weeks, and another pancreatic cancer patient in Group 3 continued on study for 28 weeks. Toxicity was similar across cohorts, with mucositis, fatigue, hyperglycemia and neutropenia reported as most common. Biopsy results were significant for increased apoptosis and tumor-infiltrating immune cells noted in tissues from 4 patients treated with the MLN0128 lead-in. Decreased F18-FDG uptake on PET/CT, often with increased ADC values in diffusion MRI, was observed in metastatic liver lesions in 4 patients after Cycle 1. Conclusions: In an expansion cohort of 31 patients treated with the combination of MLN0128 and alisertib at the previously defined MTD, treatment was tolerable with an expected toxicity profile. Prolonged stable disease was observed in 2 patients with pancreatic cancer. Increased apoptosis and tumor-infiltrating immune cells were noted in tissues from patients treated with a lead-in of MLN0128. Clinical trial information: NCT02719691.

Introduction: The impact of radiation prescription dose on postoperative complications during standard of care trimodality therapy for operable stage II-III esophageal and gastroesophageal junction cancers has not been established.Methods: We retrospectively reviewed 82 patients with esophageal or gastroesophageal junction cancers treated between 2004 and 2016 with neoadjuvant chemoradiation followed by resection at a single institution. Post-operative complications within 30 days were reviewed and scored using the Comprehensive Complication Index (CCI). Results were compared between patients treated with Results: Twenty-nine patients were treated with Conclusions: In selected esophageal and gastroesophageal junction cancer patients, radiation doses ≥ 50 Gy do not appear to increase 30 day post-operative complication rates. These findings suggest that the use of definitive doses of radiotherapy (50–50.4 Gy) in the neoadjuvant setting may not increase post-operative complications.

Purpose: The purpose of this study was to evaluate the rational combination of TORC1/2 inhibitor TAK-228 and Aurora A kinase inhibitor alisertib in preclinical models of triple-negative breast cancer (TNBC) and to conduct a phase I dose escalation trial in patients with advanced solid tumors. Experimental Design: TNBC cell lines and patient-derived xenograft (PDX) models were treated with alisertib, TAK-228, or the combination and evaluated for changes in proliferation, cell cycle, mTOR pathway modulation, and terminal cellular fate, including apoptosis and senescence. A phase I clinical trial was conducted in patients with advanced solid tumors treated with escalating doses of alisertib and TAK-228 using a 3+3 design to determine the maximum tolerated dose (MTD). Results: The combination of TAK-228 and alisertib resulted in decreased proliferation and cell-cycle arrest in TNBC cell lines. Treatment of TNBC PDX models resulted in significant tumor growth inhibition and increased apoptosis with the combination. In the phase I dose escalation study, 18 patients with refractory solid tumors were enrolled. The MTD was alisertib 30 mg b.i.d. days 1 to 7 of a 21-day cycle and TAK-228 2 mg daily, continuous dosing. The most common treatment-related adverse events were neutropenia, fatigue, nausea, rash, mucositis, and alopecia. Conclusions: The addition of TAK-228 to alisertib potentiates the antitumor activity of alisertib in vivo, resulting in increased cell death and apoptosis. The combination is tolerable in patients with advanced solid tumors and should be evaluated further in expansion cohorts with additional pharmacodynamic assessment.

118 Background: To date, immune-checkpoint inhibition for microsatellite stable (MSS) mCRC has been ineffective, though targeted therapy combination strategies may be promising. This phase II, investigator-initiated trial (NCT03475004) was designed to evaluate the efficacy and safety of the three-drug combination of pembrolizumab (pembro), binimetinib, and bevacizumab in patients with advanced, MSS treatment-refractory colorectal cancer. Methods: Patients with mCRC locally determined to be MSS and whom have progressed on two prior lines of therapy were enrolled. Treatment consists of pembro (200 mg every 3 weeks), binimetinib (45 mg BID) and bevacizumab (7.5 mg/kg every 3 weeks) until disease progression or unacceptable toxicity. The primary endpoint is PFS using RECIST v1.1 by investigator review. Additional endpoints include objective response rate, disease control rate at time of first re-staging (2 mo), duration of response, and safety and tolerability. Results: 50 patients have been enrolled (accrual is completed). 53% of patients are male and the mean age is 53.6 (range 31-79). The mean number of prior therapies is 5.3. At the time of preliminary data review, 39 patients are evaluable for response data. The median PFS was 5.8 mo (95% CI 4.2 to 8.9). The objective response rate was 13% with 5 partial responses. 24 patients (62%) had stable disease and 10 (26%) had progressive disease as the best response. The disease control rate at the time of first re-staging was 74%. Median duration of response was 6.5 mo. 19 (40%) patients experienced serious adverse events; the most common grade ≥3 adverse events included transaminase elevation (15%), diarrhea (11%), acneiform rash (9%), hypertension (9%), and anemia (9%). Conclusions: Preliminary results from this phase II study indicate that this regimen of pembrolizumab, binimetinib, and bevacizumab has promising activity and acceptable tolerability in this heavily pre-treated population of patients with MSS metastatic colorectal cancer. Final results will be presented as well as ongoing correlative studies. Clinical trial information: NCT03475004.

Aim: Far less is known about colorectal cancer (CRC) incidence in individuals under the age of 50. This study examined CRC incidence in order to better understand the changing CRC population. Methods: This study analyzed 39,525 CRC cases from the Colorado Central Cancer Registry from 1992 through 2013. Age-adjusted incidence, observed and relative 5-year survival, and estimated annual percentage change was analyzed. Results: Age-adjusted rates averaging 1.7% per year were observed in the under-50 population, while falling on average 4.3% per year (p < 0.05) in the over-50 population. Average-adjusted incidence rose in males under 50 by 2.7% per year (p < 0.05). Conclusion: The absolute incidence of CRC continues to fall in Colorado, however incidence is rising in individuals under 50, particularly males.

OBJECTIVES The role of radiotherapy (RT) in locally advanced pancreatic cancer (LAPC) is uncertain. This study examines patterns of care and survival outcomes of LAPC undergoing chemotherapy alone versus chemotherapy plus RT (C + RT). METHODS The National Cancer Database was queried for nonmetastatic LAPC patients who received chemotherapy alone or C + RT. RESULTS Of the 13,695 patients included, 5306 underwent chemotherapy alone and 4971, C + RT. Use of C + RT declined from 2003 to 2011 (73%-53%), whereas chemotherapy alone increased. Of those receiving RT, rates of intensity-modulated radiotherapy (IMRT) increased (27%-72%), whereas 3-dimensional (3D) RT decreased (73%-28%). Unadjusted 1-year overall survival (OS) was longer for versus chemotherapy (45.6% vs 38.7%), as was 2-year OS (12.9% vs 11.9%) (hazard ratio, 0.88; 0.85-0.91; P < 0.001). Under multivariate analysis, C + RT was associated with improved OS (hazard ratio, 0.84; 0.81-0.87; P < 0.001). On subgroup analysis comparing C + IMRT, C + 3D RT, and chemotherapy alone, 1-year OS was 49.1%, 45.1%, and 38.7%, and 2-year OS was 13.1%, 11.6%, and 11.9% accordingly. CONCLUSIONS Utilization of RT in LAPC is decreasing, whereas chemotherapy alone is increasing. Of patients undergoing RT, rates of IMRT are increasing. Whereas C + IMRT appeared to be associated with improved OS compared with chemotherapy alone, 3D RT was not.

3075 Background: OKI-179 is a novel, oral pro-drug analog of largazole, a compound in the romidepsin-depsipeptide class of natural products. OKI-006, the active metabolite of OKI-179, inhibits HDAC 1,2,3 (IC50 = 1.2, 2.4, 2.0 nM, respectively), with no significant inhibition of Class IIa HDACs and has shown promising activity in preclinical models of solid tumors. We conducted a first-in-human dose escalation study of OKI-179 in patients with advanced solid tumors. Methods: Patients with advanced solid tumors, ECOG ≤1, normal QTc, and disease refractory to or with no available standard therapy options were treated with OKI-179 with either intermittent dosing (once daily for 4 days on 3 days off) or continuous dosing (once daily). Dose escalation was conducted using a standard 3+3 design. Pharmacokinetic (PK) and pharmacodynamic (PD) testing was performed at various time points after dosing. Results: As of Feb 4, 2021, 26 patients (19 female, 7 male) were enrolled with mean age of 63 (range 41-83). Patients received a median of 5 (range 1-11) prior lines of therapy and most common tumor types included pancreatic (N = 5), breast (N = 4), lung (N = 4), and ovarian cancer (N = 4). Twenty patients were treated in intermittent dosing cohorts from 30-450 mg. One DLT (Grade 2 [G2] thrombocytopenia) occurred in the 450 mg cohort which was expanded to 6 patients without subsequent DLTs. Six patients were treated in 2 continuous dosing cohorts of 200 mg and 300 mg. Two of 3 patients in the 300 mg cohort had DLTs of G3-4 thrombocytopenia and no DLTs were observed in 3 patients treated at 200 mg PO daily. The most common adverse events (AEs) were nausea (62%), fatigue (42%), anemia (39%), anorexia (27%), and vomiting (23%). These AE’s were G1-2 except for G3 anemia (12%), G3 fatigue (12%), and G3 anorexia (4%). No other G4-5 treatment-related AEs occurred. Median time on study was 79 days and best response was stable disease (SD) in 10 of 24 patients evaluable for efficacy (42%). Prolonged SD was observed in patients with platinum-resistant serous ovarian cancer (446 days) and adenoid cystic nasopharyngeal carcinoma (256 days). OKI-006 achieved consistent exposure with Cmax > 2,000 ng/ml and AUC > 8,000 hr*ng/ml, well above the targeted exposure for efficacy based on pre-clinical studies in murine models. Tmax was 2 hours and T1/2 was 6-8 hours. OKI-179 treatment resulted in > 3X increased T cell histone H3K9 and H3K27 acetylation within circulating PBMCs at doses of 180 - 450 mg. Conclusions: OKI-179 has a manageable safety profile, with thrombocytopenia being the on-target DLT. It has a favorable PK profile and demonstrated on-target PD effects at tolerable doses. The MTD and RP2D for OKI-179 was 450 mg daily for intermittent dosing and 200 mg daily for continuous dosing. Phase 2 studies are being designed, with a focus on combination with endocrine therapy in ER+ breast cancer and in NRAS-mutant melanoma. Clinical trial information: NCT03931681.

e14559 Background: While TMB is very dependent on methodology, tissue TMB-H (≥10 mutations/megabase) may predict benefit with ICIs. Pembrolizumab received tissue-agnostic approval for TMB-H unresectable cancers in 2020, but little is known about TMB as a predictive biomarker in mGI cancers. We hypothesized that tissue TMB will correlate with efficacy of ICIs in mGI cancers. Methods: A retrospective chart review identified patients with mGI cancers who received an anti-PD-(L)1 drug and had known TMB at a single academic center from 2012 to 2020. The association of TMB with objective response rate (ORR), progression-free survival (PFS), and overall survival (OS) was analyzed using the Fisher’s exact and Log-rank tests. Survival curves were generated using the Kaplan-Meier method. Cox proportional hazard and logistic regression models were used to adjust for microsatellite status. Significance was prespecified at 0.05. Results: 83 patients were identified and included. The most common cancer types were colorectal adenocarcinoma (AC, n = 29), esophageal/gastric AC (n = 21) and SCC (n = 4), cholangiocarcinoma (n = 11), anal SCC (n = 7), and pancreas AC (n = 7). Average age was 61, average number of lines of prior systemic therapy for advanced disease was 1.3 (range 0-4), and 37% of patients were treated on a clinical study. All patients received an anti-PD-(L)1 drug; 6%, 2%, and 36% also received ipilimumab, cytotoxic chemotherapy, and other combinations, respectively. Among those with esophageal/gastric cancer, 76% had known PD-L1 CPS (84% ≥1, 63% ≥5, 42% ≥10). TMB was primarily determined by Foundation One CDx (87%). TMB ranged from 0 to 54; n = 22 (27%) were TMB-H (of these, n = 10 were microsatellite instability-high (MSI-H)), and n = 61 were TMB-L ( < 10 mutations/megabase; of these, n = 2 were MSI-H). The prevalence of TMB-H and microsatellite stable (MSS) was 14.4%. TMB-L, compared to TMB-H, was associated with inferior ORR (3.5% vs 55.6%; odds ratio (OR) 0.045; p < 0.001) and PFS (median 12.7 vs 29.3 weeks; hazard ratio (HR) 2.70; p = 0.001), but not OS (HR 1.20; p = 0.60). In patients with MSS disease, TMB-L, compared to TMB-H, was associated with inferior ORR (OR 0.13; p = 0.04) but not PFS (HR 1.76; p = 0.07) or OS (HR 0.89; p = 0.79). In subgroup analyses, ORR was not significantly associated to tumor type in all or MSS patients. TMB as a continuous variable, in patients with MSS disease, was positively correlated with ORR (p = 0.02) and PFS (p = 0.04), but not OS (p = 0.59). Among all patients, PFS and OS data is immature (median follow-up 13 and 31 weeks). Conclusions: In a single center retrospective study of patients with mGI cancers treated with ICIs, TMB-H was associated with improved ORR and PFS compared to TMB-L. In patients with MSS disease, ORR remained significant. PFS and OS data are immature. TMB as a biomarker of efficacy with ICIs in mGI cancers warrants further study to guide clinical use.

e16276 Background: Appendiceal adenocarcinoma is relatively rare and often diagnosed incidentally during operations for acute appendicitis. It is commonly associated, either at time of initial presentation or upon recurrence, with peritoneal metastases. A typical treatment strategy for patients with peritoneal disease includes cytoreductive surgery (CRS) with hyperthermic intraperitoneal chemotherapy (HIPEC). Extrapolating largely from literature in colorectal cancer, chemotherapy is frequently given before and/or after CRS/HIPEC though high-level evidence to support this is lacking. We sought to evaluate the effect of systemic chemotherapy on survival. Methods: Utilizing a database of CRS/HIPEC procedures at University of Colorado Hospital from 2008 to present we retrospectively reviewed cases of appendiceal adenocarcinoma. Data collected included staging, histologic grade, chemotherapy given, surgical outcomes, and time to disease recurrence. Patients without adequate information regarding treatment, or without at least 1 year of clinical follow-up, were excluded. Associations between administration of chemotherapy or histologic grade and 1-year DFS were analyzed using Fisher’s exact test, and logistic regression was used to assess whether 1-year DFS were different in chemotherapy-treated patients when adjusted for histologic grade. Results: In total, 117 cases reviewed indicated an appendiceal pathology. Of these, 54 cases in a total of 51 patients met the specified criteria for pathology and completeness and length of follow-up information. The average age was 58 years (range 26-81 years). Adenocarcinoma was graded as low in 15 (28%) cases, intermediate in 18 (33%) cases, and high in 21 (39%) cases. 23 (43%) patients received no chemotherapy while 31 (57%) received chemotherapy before and/or after surgery. In the overall population, there was no significant effect of chemotherapy on survival, with 1-year DFS demonstrated in 74.2% of patients receiving some chemotherapy and 70% in patients not receiving chemotherapy (p = 0.765). One-year DFS was achieved in 86% of low-grade cases, 61% of intermediate-grade cases, and 71% of high-grade cases, though this was also not statistically significant (p = 0.254). Furthermore, when 1-year DFS between chemotherapy and non-chemotherapy patients was adjusted for grade, there was again no significant interaction (odds ratio = 0.48, 95% C.I. (0.13-1.64), p = 0.763). Conclusions: In this small, single-institution experience of patients with peritoneal appendiceal adenocarcinoma, there was no significant effect of chemotherapy administration on 1-year DFS. These findings are likely affected by significant confounding with the small sample size and retrospective nature of the data. Further investigation on a larger scale is warranted.

64 Background: The management of locally advanced rectal cancer has historically included preoperative chemoradiation followed by surgery and then adjuvant chemotherapy. Recently there has been an increasing utilization of preoperative chemotherapy in addition to standard chemoradiation, a strategy known as total neoadjuvant therapy (TNT). TNT has been offered to patients at the University of Colorado Cancer Center since 2015. Methods: Records of all patients presenting to the University of Colorado colorectal multidisciplinary clinic since 2015 were screened for treatment with TNT. Data collected on these patients included demographic information, diagnosis and initial staging, preoperative treatment received, and surgical outcomes including treatment response and pathological stage. TNT included preoperative chemotherapy with oxaliplatin combined with either 5-FU (FOLFOX) or capecitabine (CAPOX) as well as chemoradiation, generally given with concurrent capecitabine. Patients then underwent surgical resection; if a complete clinical response was achieved with TNT, non-operative management (NOM) was offered. Results: A total of 81 patients thus far have undergone TNT followed by resection or, if complete clinical response and preferred by the patient, NOM. The mean age of patients was 56 years, ranging from 23 to 87, and 60% of patients were male. The majority of patients (67) had stage III disease at presentation while 1 had stage 1 (T2N0) disease, 11 had stage II disease and 2 patients had oligometastatic disease. Ultimately 13 patients (16%) opted for non-operative management after being found to have a complete clinical response following TNT. Of the 68 patients who underwent surgical resection, 21 (31%) had a pathological complete response, with another 14 (21%) with near-complete response. 28 patients (41%) had a partial treatment response and 5 (7%) had no treatment response. In total, the rate of complete clinical or pathologic response was 42%. Treatment was overall well-tolerated with 90% of patients receiving the full planned dose of radiation and 98% of patients completing all planned cycles of chemotherapy, though most of them with typical dose reductions needed. Of the patients who underwent surgery, 49 (72%) had low anterior resection and 19 (28%) had an abdominoperineal resection. Of patients with temporary ileostomies, 85% of them had their ileostomy reversed within 10 weeks of surgery. Conclusions: Treatment of locally advanced rectal cancer by a total neoadjuvant approach is well-tolerated and results in a high rate of clinical and pathological complete response.

// Todd M. Pitts 1, 3, * , Erica L. Bradshaw-Pierce 2, 3, 5, * , Stacey M. Bagby 1 , Stephanie L. Hyatt 1 , Heather M. Selby 1 , Anna Spreafico 1 , John J. Tentler 1, 3 , Kelly McPhillips 1 , Peter J. Klauck 1 , Anna Capasso 1 , Jennifer R. Diamond 1, 3 , S. Lindsey Davis 1, 3 , Aik Choon Tan 1, 3 , John J. Arcaroli 1, 3 , Alicia Purkey 1 , Wells A. Messersmith 1, 3 , Jeffery A. Ecsedy 4 , S. Gail Eckhardt 1, 3 1 Division of Medical Oncology, School of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, CO, USA 2 Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado, Anschutz Medical Campus, Aurora, CO, USA 3 University of Colorado Cancer Center, University of Colorado Anschutz Medical Campus, Aurora, CO, USA 4 Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Limited, Cambridge, MA, USA 5 Takeda California, San Diego, CA, USA * These authors have contributed equally to this work Correspondence to: Todd M. Pitts, email: Todd.Pitts@ucdenver.edu Keywords: colorectal cancer, aurora kinase a Received: January 20, 2016 Accepted: June 17, 2016 Published: July 1, 2016 ABSTRACT Background: The Aurora kinases are a family of serine/threonine kinases comprised of Aurora A, B, and C which execute critical steps in mitotic and meiotic progression. Alisertib (MLN8237) is an investigational Aurora A selective inhibitor that has demonstrated activity against a wide variety of tumor types in vitro and in vivo , including CRC. Results: CRC cell lines demonstrated varying sensitivity to alisertib with IC 50 values ranging from 0.06 to > 5 umol/L. Following exposure to alisertib we observed a decrease in pAurora A, B and C in four CRC cell lines. We also observed an increase in p53 and p21 in a sensitive p53 wildtype cell line in contrast to the p53 mutant cell line or the resistant cell lines. The addition of alisertib to standard CRC treatments demonstrated improvement over single agent arms; however, the benefit was largely less than additive, but not antagonistic. Methods: Forty-seven CRC cell lines were exposed to alisertib and IC 50 s were calculated. Twenty-one PDX models were treated with alisertib and the Tumor Growth Inhibition Index was assessed. Additionally, 5 KRAS wildtype and mutant PDX models were treated with alisertib as single agent or in combination with cetuximab or irinotecan, respectively. Conclusion: Alisertib demonstrated anti-proliferative effects against CRC cell lines and PDX models. Our data suggest that the addition of alisertib to standard therapies in colorectal cancer if pursued clinically, will require further investigation of patient selection strategies and these combinations may facilitate future clinical studies.

Background: BTCs are aggressive cancers with a poor prognosis. In preclinical models, MEK inhibition modulates the tumor immune microenvironment and enhances responses to programmed death-ligand 1 (PD-L1) inhibition. We report a randomized, open-label, multicenter phase 2 trial of atezolizumab (anti-PD-L1) as monotherapy or in combination with cobimetinib (MEK inhibitor) in BTC (NCT03201458). Methods: Eligible patients had advanced BTC [intrahepatic cholangiocarcinoma (ICC), extrahepatic cholangiocarcinoma (ECC), and gallbladder cancer (GBC)], with 1-2 lines of prior therapy in the metastatic setting, measurable disease by RECIST v1.1, and ECOG performance status ≤1. Patients randomized to Arm A received atezolizumab 840 mg IV Q2w. Patients randomized to Arm B received oral cobimetinib 60 mg daily (21 days on/7 days off) plus atezolizumab 840 mg IV Q2w. The primary endpoint was progression free survival (PFS) using the Kaplan-Meier method and compared between groups under the assumption of Cox proportional hazards, stratified for primary tumor site. Secondary endpoints included objective response rate (ORR), safety and tolerability, and overall survival (OS). Results: 86 patients were enrolled at 23 centers in the United States; 77 patients were randomized and received at least one dose of study therapy (Arm A: n=37, ICC=21, ECC=7, GBC=11; Arm B: n=38, ICC=22, ECC=8, GBC=8). Median age was 63 (range 44-86), and 48 (62%) were female. The trial met its primary endpoint, with a median PFS of 3.65 months (cobimetinib+atezolizumab) vs 1.87 months (atezolizumab monotherapy) (p=0.027). OS data are not mature at the time of analysis. There was 1 PR (3.2%), 13 SD (41.9%), and 17 PD (54.8%) in the combination arm and 1 PR (2.9%), 10 SD (29.4%), and 23 PD (67.6%) in the atezolizumab monotherapy arm. Two patients in the combination arm remain on therapy 15+ months from enrollment. One patient in each treatment arm had known mismatch repair deficiency (MMRd), of whom 1 had PD as a best response and the other withdrew prior to response evaluation. Grade 3-4 treatment-related adverse events were similar in both arms, and there were no treatment-related deaths. 4 (10%) of patients receiving atezolizumab monotherapy and 8 (22.2%) receiving cobimetinib+atezolizumab discontinued therapy due to adverse events. Changes in tumor CD8, CD4, FoxP3, PDL1, and MHC expression from paired tumor biopsies will be presented at the conference. Conclusions: We report the first randomized trial of immunotherapy in BTC. The combination of atezolizumab plus cobimetinib met its primary endpoint and significantly prolonged PFS as compared to atezolizumab monotherapy in BTC. The combination of atezolizumab and cobimetinib had manageable toxicity and warrants further investigation in BTC. Citation Format: Mark Yarchoan, Leslie Cope, Robert A. Anders, Anne Noonan, Laura W. Goff, Lipika Goyal, Jill Lacy, Daneng Li, Anuj Patel, Aiwu R. He, Ghassan Abou-Alfa, Kristen Spencer, Edward Kim, Stephanie Xavier, Amanda Ruggieri, S. Lindsey Davis, Autumn McRee, Paul Kunk, Qingfeng Zhu, Andrea Wang-Gillam, Andrew Poklepovic, Helen Chen, Elad Sharon, Gregory B. Lesinski, Nilo Azad. A multicenter randomized phase 2 trial of atezolizumab as monotherapy or in combination with cobimetinib in biliary tract cancers (BTCs): A NCI Experimental Therapeutics Clinical Trials Network (ETCTN) study [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 CT043.

Immunotherapies that target immune regulatory checkpoints such as CTLA-4 and PD-1 are widely used among many cancer types and have shown positive results in CRC with high microsatellite instability. However, in microsatellite stable (MSS) CRC there is a dismal response rate of 0%. The limited efficacy has shown to be partially due to the lack of T-cells in the tumor microenvironment and/or no activation/regulation of paramount cells in the immune system. The Wnt pathway is the most commonly altered pathway in CRC and is highly involved in driving tumor initiation and progression. Recent evidence also demonstrates the Wnt pathway is involved in T-lymphocyte development, maturation/activation of CD8+ effector T cells and recruitment of dendritic cells. Therefore, targeting the Wnt pathway utilizing a Porcupine (PORCN) inhibitor (ETC-159) in MSS CRC may be a promising strategy to sensitize tumors to immune checkpoint inhibition. Human Immune System BRGS (BALB/c, Rag2−/−, IL2RγC−/−, NODSIRPα) mice were engrafted with MSS CRC PDX (hPDX). The hPDX were randomized according to human chimerism into the following drug treatments groups: Vehicle, ETC-159, nivolumab, and the combination. Treatments began when tumors reached 100-300mm3 and tumors were measured twice weekly. At the end of study, sera, lymph nodes, spleen, and tumor tissue were collected for immunohistochemistry, single cell suspensions, and flow cytometry analysis. Combination therapy resulted in a significant decrease in tumor volume compared to both single agents and vehicle. Flow cytometric analysis demonstrated an increase in human immune cells, in particular human CD4 and CD8 cells in the combination compared to the vehicle and nivolumab treated groups. Additionally, these T-cells showed increased signs of activation and effector function, as indicated by increased CD69+ expression, effector memory subsets, and granzyme B+ cells in the TILs, with a further reduction in Treg populations, suggesting an overall increase in inflammation. An increase in MHC II expression on tumor cells was observed in the ETC-159 single agent with a statistically significant increase in the combination treated tumors demonstrating enhanced antigen presentation. Furthermore, PD-1 expression was upregulated on CD4+ T-cells in the ETC-159 single agent. Lastly, VECTRA analysis corroborates the flow cytometry data showing a changing tumor immune landscape through an increase in CD4+ and CD8+ T cells in the tumor and surrounding stroma. Our data demonstrates the combination treatment of ETC-159 + nivolumab in MSS CRC hPDX show increased tumor infiltration of human immune cells. Further preclinical data is compulsory but these results support further development of this combination in clinical trials. Citation Format: Stacey M. Bagby, Sarah J. Hartman, Natalie M. Navarro, Betelehem W. Yacob, Jeremy Shulman, Jessica Barkow, Christopher H. Lieu, S. Lindsey Davis, Alexis D. Leal, Wells A. Messersmith, Angela Minic, Kimberly R. Jordan, Julie Lang, Todd M. Pitts. Sensitizing microsatellite stable colorectal cancer to immune checkpoint therapy utilizing Wnt pathway inhibition [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 6647.

Pancreatic ductal adenocarcinoma (PDAC) is currently the fourth leading cause of cancer deaths with more than 56,000 new cases estimated to be diagnosed in 2019. Current treatment options for PDAC include radiation and chemotherapeutic regimens, however these targeted therapies are ineffective for patients with advanced disease progression. Additionally, the dense stromal nature of PDAC tumors create challenges to target the cancer cells resulting in incomplete cell killing and eventual drug resistance. Recent evidence has shown that CPT1A, an enzyme that regulates the entry of lipids into mitochondria for β-oxidation, is strongly expressed in several cancers. CPT1A is located on the mitochondrial membrane and potentially interacts with BCL-2, an anti-apoptotic protein that promotes tumor maintenance and metastasis. Metabolic stress can activate the anti-apoptotic effects of BCL-2, reprograming metabolism to use fat oxidation for cancer survival. Therefore, a co-inhibition using the selective BCL-2 inhibitor, venetoclax, with agents that inhibit CPT1A and β-oxidation, could be a novel strategy for PDAC. There are few studies considering CPT1A as a therapeutic target for PDAC. Current available drugs to target these pathways include the anti-anginal ranolazine, and CPT1A inhibitors etomoxir and perhexiline. Previous studies have shown that expression of BCL-2 by tumor cells is necessary for BCL-2 inhibitors to be effective. We initially wanted to determine the expression of BCL-2 and CPT1A in PDAC cells utilizing western blot and rtPCR, and to confirm their proximity using a proximity ligation assay (PLA). PDAC cells were then plated in 96 well plates and Cell Titer-Glo assays were performed to determine effective concentrations of single agent venetoclax, etomoxir, and perhexiline. The effects of these drugs in combination were then evaluated using a clonogenic assay, which was analyzed using the ImageJ colony area plugin. PDAC cells were then exposed to the combinations and western blots were performed to evaluate changes downstream effectors. We have confirmed the expression of BCL-2 and CPT1A on the mitochondrial membrane using Westerns, rtPCR, and a PLA on several PDAC lines. Though single agent drugs had little effect on cell viability, the combination of venetoclax with CPT1A and β-oxidation inhibitors decreased colony formation in some PDAC cell lines. Western blot analysis revealed the drug combinations affected the phosphorylation of AKT and 4E-BP1 and expression of the pro-apoptotic protein BID. These data suggest that co-targeting BCL-2 and CPT1A have potential for anti-tumor effects in PDAC. Additional research into the role of CPT1A in PDAC biology will elucidate the optimal dosing concentrations and mechanisms for further studies. Citation Format: Sarah J. Hartman, Nathalie Nadales, Stacey M. Bagby, Betelehem W. Yacob, Brian L. Gittleman, Adriana Estrada-Bernal, Anh T. Le, Christopher H. Lieu, S. Lindsey Davis, Alexis D. Leal, Jennifer R. Diamond, Wells A. Messersmith, Isabel R. Schlaepfer, Todd M. Pitts. Therapeutic targeting of lipid oxidation and apoptosis in pancreatic ductal adenocarcinoma [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 6387.

258 Background: Patients with clinical stage II or III locally advanced rectal cancer may be treated with the total neoadjuvant therapy (TNT) approach; chemotherapy with 4 mths of FOLFOX followed by chemoradiation (chemo/XRT) with capecitabine for 5 wks administered before surgery. We hypothesized that full dose intensity is not necessary for treatment benefit. Methods: A retrospective chart review was conducted on patients with newly diagnosed rectal cancer recommended to receive TNT by the multidisciplinary (multiD) colorectal cancer tumor board at the University of Colorado Cancer Center (UCCC). The primary objective was to evaluate dose intensity of TNT and its impact on response assessed by biopsy and/or imaging (MRI). Results: Between January 31, 2016 and January 31, 2019, 80 patients were recommended the TNT approach for cancer management by the multiD team. Of those, 48 completed their neoadjuvant treatment at UCCC and were included in the analysis. The average age was 55 years (range 23-80) and 61% were male. Thirty-one patients had an ECOG of 0 and 17 had an ECOG of 1. Overall responses were 44% complete response (CR, n = 21), 15% near complete response (nCR, n = 7), 35% partial response (PR, n = 17), and 6% no response (NR, n = 3). See Table for responses seen by dose intensity for chemotherapy. Two patients did not receive their full planned XRT course, and 9 patients had their capecitabine doses held/decreased during chemoradiation. Conclusions: This single center retrospective analysis of patients receiving the TNT approach for rectal cancer provides data supporting that achieving full dose intensity is not necessary to achieve treatment benefit. [Table: see text]

MEK inhibition is of interest in cancer drug development, but clinical activity in metastatic colorectal cancer (mCRC) has been limited. Preclinical studies demonstrated Wnt pathway overexpression in KRAS-mutant cell lines resistant to the MEK inhibitor, selumetinib. The combination of selumetinib and cyclosporin A, a noncanonical Wnt pathway modulator, demonstrated antitumor activity in mCRC patient-derived xenografts. To translate these results, we conducted a NCI Cancer Therapy Evaluation Program–approved multicenter phase I/IB trial (NCT02188264) of the combination of selumetinib and cyclosporin A. Patients with advanced solid malignancies were treated with the combination of oral selumetinib and cyclosporin A in the dose escalation phase, followed by an expansion cohort of irinotecan and oxaliplatin-refractory mCRC. The expansion cohort utilized a single-agent selumetinib “run-in” to evaluate FZD2 biomarker upregulation and KRAS-WT and KRAS-MT stratification to identify any potential predictors of efficacy. Twenty and 19 patients were enrolled in dose escalation and expansion phases, respectively. The most common adverse events and grade 3/4 toxicities were rash, hypertension, and edema. Three dose-limiting toxicities (grade 3 hypertension, rash, and increased creatinine) were reported. The MTD was selumetinib 75 mg twice daily and cyclosporin A 2 mg/kg twice daily on a 28-day cycle. KRAS stratification did not identify any differences in response between KRAS-WT and KRAS-MT cancers. Two partial responses, 18 stable disease, and 10 progressive disease responses were observed. Combination selumetinib and cyclosporin A is well tolerated, with evidence of activity in mCRC. Future strategies for concept development include identifying better predictors of efficacy and improved Wnt pathway modulation. Significance: These findings translate preclinical studies combining selumetinib and cyclosporin into a phase I first-in-human clinical trial of such a combination in patients with advanced solid malignancies. Cancer Res; 78(18); 5398–407. ©2018 AACR.

Cholangiocarcinoma, or cancer arising from bile duct epithelium, represents about 10–25% of primary liver cancers worldwide. Intrahepatic cholangiocarcinoma occurs in the bile ducts within the hepatic parenchyma, while extrahepatic tumors occur from the junction of the hepatic ducts to the common bile duct. Hilar cholangiocarcinoma is a subset of extrahepatic disease occurring at or near the junction of the left and right hepatic ducts. Chronic inflammation of the biliary tree from such processes such as liver fluke infection and primary sclerosing cholangitis confers a higher risk of developing cholangiocarcinoma. Diagnosis generally requires multiple modalities including laboratory tests, imaging studies, and often endoscopic procedures and biopsy. Cholangiocarcinoma is now staged according to subtype, with unique systems specific to intrahepatic, hilar, and extrahepatic diseases. As the only confirmed curative intervention for the treatment of cholangiocarcinoma, surgical intervention in the form of resection or liver transplant should be considered in all patients with localized disease. Locoregional therapies may provide some benefit to patients with localized disease who are not surgical candidates. Patients with metastatic cholangiocarcinoma may benefit from combination chemotherapy with gemcitabine and cisplatin, though prognosis remains poor. This chapter reviews the epidemiology, risk factors, and pathogenesis of intrahepatic and hilar cholangiocarcinomas, as well as the basic principles of diagnosis, staging, and treatment.

Introduction: Pancreatic ductal adenocarcinoma (PDA) is the fourth leading cause of cancer death and has a 5-year survival rate of less than 7%. The poor prognosis associated with PDA is related in part to a lack of screening tests to promote early detection and ineffective systemic targeted therapies. Adavosertib (AZD1775, MK1775) is a selective Wee1 inhibitor with promising preclinical activity in PDA and synergy with cytotoxic chemotherapy in other cancer types. Wee1 is a tyrosine kinase that activates in the G2M cell cycle checkpoint in response to DNA damage. Inhibition of Wee1 with adavosertib prevents the phosphorylation of CDC2, thus allowing unrepaired DNA to enter mitosis and ultimately succumb to mitotic catastrophe. The purpose of this study was to investigate adavosertib in combination with standard chemotherapy and other targeted agents in preclinical models of PDA. Methods: Athymic nude mice were implanted with PDA PDX models on the right and left flanks. When the average tumor volume reached 100-300 mm3, mice were randomized into one of the following treatments: vehicle, adavosertib, irinotecan, navitoclax, capecitabine, adavosertib + irinotecan, or adavosertib + navitoclax, adavosertib + capecitabine. Tumor volume was calculated using the following equation: volume = (length × width) × 0.52. Four pancreatic cancer cell lines were plated in 96-well plates and Cell Titer-Glo proliferation assays were performed to determine the most effective combination doses of irinotecan, 5FU, or navitoclax with adavosertib in vitro. Combination effects were analyzed using CalcuSyn software. The most effective doses within each cell line were selected and used for Caspase 3/7 apoptosis assays and cell cycle analyses by flow cytometry. Western blots were performed to evaluate changes in downstream effectors. Results: In vivo, the combination of adavosertib with either irinotecan or navitoclax resulted in decreased tumor growth compared to the respective single agents. The combination of adavosertib with irinotecan, 5FU, or navitoclax in vitro resulted in greater antiproliferative effects in all cell lines, and the several combinations were synergistic in all cell lines as determined by CI values less than 1. Navitoclax increased apoptosis in several cell lines both as a single agent and was enhanced in combination with adavosertib. Irinotecan proved to be more cell cycle dependent and significantly altered the cell cycle in all cell lines. Irinotecan increased phospho-CDC2 and decreased PHH3, while adavosertib increased gamma-H2AX as a single agent and in combination. Conclusions: The combination of adavosertib with either irinotecan, 5FU, or navitoclax in vivo decreased tumor growth and had enhanced antiproliferative effects in vitro. These data support future studies with adavosertib in combination with standard therapies or navitoclax to treat PDA. Citation Format: Sarah J. Hartman, Stacey M. Bagby, Betelehem W. Yacob, Dennis M. Simmons, Tonia E. Tse, Christopher H. Lieu, S. Lindsey Davis, Alexis D. Leal, Jennifer R. Diamond, Wells A. Messersmith, Todd M. Pitts. Combination of Wee1 inhibition with targeted and standard chemotherapy in preclinical models of pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1315.

Background: AZD0156 is an oral inhibitor of ATM, a serine threonine kinase that plays a key role in DNA damage response associated with DNA double strand breaks. Topoisomerase-I inhibitors like irinotecan induce single-strand DNA breaks, which are converted to double-strand breaks during DNA replication. Thus the combination of AZD0156 and irinotecan is a rational combination for clinical use. Irinotecan is used clinically to treat a variety of malignancies, including colorectal cancer (CRC), usually in combination with 5-fluorouracil (5FU) as FOLFIRI. An ongoing phase 1 clinical trial is evaluating AZD0156 in combination with single-agent irinotecan and FOLFIRI in patients with refractory cancers (NCT02588105). The purpose of this study is to evaluate AZD0156 in combination with irinotecan and 5FU in preclinical models of CRC to help inform clinical use. Methods: Anti-proliferative effects of single-agent AZD0156 and combination therapy with SN38 (active metabolite of irinotecan) and 5FU were evaluated in CRC cell lines using the Cell-Titer Glo assay. Immunoblotting and cell cycle analysis were performed to determine the mechanism of enhanced combination effects. Four CRC patient derived xenograft (PDX) models were treated with AZD0156, irinotecan, and 5FU alone and in combination for assessment of tumor growth inhibition (TGI). Results: An enhanced antiproliferative effect was observed with the combination treatment over either single agent. A more significant synergistic effect was demonstrated with the combination of AZD0156 and SN38 as compared with the combination of AZD0156 and 5FU. Cell cycle data demonstrated enhanced cell cycle arrest with combination therapy as compared to single agents. Immunoblotting results suggest a decrease in phosphorylated gamma-H2AX in cell lines treated with combination therapies. Increased TGI was observed in CRC PDX models treated with the combination of AZD0156 and irinotecan as compared to single-agent therapy in 3 of 4 models. There was not a significant change in TGI with the addition of 5FU for triplet therapy in the majority of models. Conclusions: The combination of AZD0156 with irinotecan is synergistic in in vitro models and is associated with increased TGI in CRC PDX in vivo models. The addition of 5FU to AZD0156 and irinotecan did not result in increased TGI as compared to doublet therapy in CRC PDX models, though did not decrease the AZD0156/irinotecan combination effect. An ongoing clinical trial is evaluating this combination in patients with cancers refractory to standard treatments (NCT02588105). Citation Format: S. Lindsey Davis, Marina I. Schlaepfer, Stacey M. Bagby, Sarah J. Hartman, Betelehem W. Yacob, Tonia Tse, Dennis M. Simmons, Jennifer R. Diamond, Christopher H. Lieu, Alexis D. Leal, Elaine B. Cadogan, Gareth D. Hughes, Stephen T. Durant, Wells A. Messersmith, Todd M. Pitts. Ataxia telangiectasia mutated (ATM) kinase inhibitor AZD0156 in combination with 5-fluorouracil and irinotecan in preclinical models of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4720.

3576 Background: In contrast to the older population, the incidence of colorectal cancer (CRC) in younger patients (aged < 50 years) has been increasing in the last three decades. Younger patients tend to present with more advanced disease, thought to be in part related to lack of routine screening colonoscopies. The goal of this study was to examine characteristics of young-onset CRC and potentially identify factors that may aid in earlier diagnosis and treatment. Methods: We collected data for patients available through the University of Colorado Cancer Center Cancer Registry. Inclusion criteria included: 1) Diagnosis of colon or rectal cancer between the years 2012-2018 and 2) age at diagnosis of less than 50 years. Pertinent data including baseline characteristics, clinical presentation, family history, pathology, molecular testing, staging, and treatment were collected. Results: 211 patients with young-onset CRC were available for review. Mean age at diagnosis was 42.4 years and 55.5% were males. A total of 42.1% had rectal cancer and a majority of the colon cancer diagnoses had left-sided tumors (66%). Regarding clinical presentation, 52.2% presented with rectal bleeding prior to diagnosis. Of those who presented with rectal bleeding, the average time from the onset of bleeding to diagnosis was 271.17 days. 42.9% of young-onset CRC were stage IV at the time of initial diagnosis. Evaluation of the pathology specimens showed that 89.6% were adenocarcinomas and 63.5% were grade 2 or higher. At diagnosis, the mean BMI was 26.6 and the mean CEA was 135.5. A total of 72.5% of young-onset patients had a positive family history of any cancer. KRAS or NRAS mutations were present in 49.6% of patients, BRAF V600E mutations were present in 3.8%, and 10.8% were MSI-H. Conclusions: Prolonged rectal bleeding history prior to diagnosis was noted in a significant proportion of young-onset patients with colorectal cancer. Patients and primary care physicians should be made aware of this finding in order to facilitate timely referral for colonoscopy which may lead to earlier diagnosis, less advanced disease at diagnosis, and improved outcomes.

1099 Background: Phase I clinical trials have traditionally enrolled patients with advanced solid tumors and many providers perceive the likelihood of clinical benefit as low. The purpose of this study was to evaluate clinical outcomes for patients with metastatic breast cancer enrolled on Phase I clinical trials and explore differences in outcomes for patients enrolled in all-comer versus breast cancer-specific cohorts. Methods: We performed a retrospective chart review of patients with metastatic breast cancer enrolled in Phase I clinical trials at the University of Colorado Cancer Center from 2012-2018. We included trials with Phase I and/or Phase Ib in the title. Studies or cohorts enrolling patients with ≥ 3 tumor types were considered all-comer and those with enrollment restricted to breast cancer or a breast cancer subtype were considered breast cancer-specific. Results: A total of 208 patients were enrolled in Phase I clinical trials, 168 in breast cancer-specific cohorts and 40 in all-comer trials. Patients on average were 56.9 years old (range 31-79), 98.6% (205/208) female, 1.4% (3/208) male, 57.2% ER+/Her2-, 30.1% ER-/Her2- and 11.1% Her2+. Patients received on average 2.1(range 0-10) prior lines of chemotherapy in the metastatic setting. Patients enrolled on Phase I clinical trials remained on study without progression on average for 138 days (CI 95%, 112.64 to 163.91). Patients enrolled on breast cancer-specific studies remained on study for 152 days (CI 95%, 120.66 to 182.56) compared to 82 days (CI 95%, 59.43 to 105.13) for those enrolled on all-comer trials, p< 0.05. Patients went off study for disease progression (83.17%), adverse events (7.69%), and other (9.14%), including withdrawal of consent. Conclusions: Patients with metastatic breast cancer previously treated with multiple lines of chemotherapy in the metastatic setting enrolled in Phase I clinical trials received clinical benefit from treatment that is favorable compared to historical controls of late-line chemotherapy. The majority of patients were treated on breast cancer-specific cohorts consistent with trends in Phase I trial design including more tumor specific cohorts.

131 Background: Immune-related adverse events (IRAE) occur commonly with immune checkpoint inhibitor therapy for the treatment of cancer, although the specific event and severity can vary widely. Little is known regarding factors that may predict which patients will develop an IRAE. The goal of this study is to identify blood biomarkers predictive of IRAE associated with immune checkpoint inhibitor therapy. Methods: Blood samples collected from patients with melanoma prior to receiving therapy with immune checkpoint inhibitors were obtained from the University of Colorado Skin Cancer Biorepository. Testing for a panel of autoantibodies and cytokines (ANA, CCP 3.1, IL-1 beta, IL-2, IL-6, IL-10, IL-12, IP-10, MCP-1, TNF alpha, IFN alpha 2, IFN gamma) in serum samples from patients who had at least one documented IRAE was performed by Exsera BioLabs. Descriptive statistics were used to evaluate biomarker levels in relation to type, grade, and number of adverse events. Results: Pre-treatment samples from 45 patients were evaluated. Median age was 55; 26 were male and 19 were female. The most common IRAEs were colitis (n = 22), thyroid dysfunction (n = 21), and dermatitis (n = 20). Most IRAEs were grade 2 in severity, and the majority of patients (n = 36) experienced more than 1 IRAE. TNF alpha was elevated in 60% of patient samples, while IFN alpha 2 was elevated in 44%. Borderline ANA was detected in 27% of samples and ANA was positive in 11%. No samples had elevation of IL-2. Between 9% and 18% of samples had elevation of the other immune markers tested (IFN gamma, IL-1 beta, IL-6, IL-10, IL-12, and CCP 3.1). Elevation of TNF alpha and IFN alpha 2 were associated with higher grades of IRAEs. No associations between immune markers and the number or type of adverse events in an individual patient were noted. Results from 15 patients who did not have a documented IRAE on immune checkpoint inhibitor therapy are currently pending to confirm these findings are unique to patients developing IRAE. Conclusions: This preliminary data suggests that baseline elevations of TNF alpha and IFN alpha 2 may predict development of IRAEs with immune checkpoint inhibitor therapy. Results from samples from patients who did not develop an IRAE on therapy will be reported at the meeting.