Truty, Mark J.

Surgical site infections (SSI) are a significant burden to patients and health care systems. They increase mortality rates, length of hospital stays, and associated health care costs.

463 Background: Molecularly targeted therapy is gaining traction in cholangiocarcinoma with the first FDA approval of a targeted agent in 2020, the FGFR inhibitor pemigatinib. Yet response rates and efficacy are modest, and most cholangiocarcinoma patients still lack targeted therapy options. The hippo pathway effector, YAP, is an oncogene that is activated in the majority of cholangiocarcinoma, yet attempts to develop YAP inhibitors have not yet been clinically successful. Recently, we described a novel mechanism of YAP regulation, via tyrosine phosphorylation by the Src-family kinase LCK. Therefore we hypothesized that LCK inhibition by may be therapeutic in cholangiocarcinoma through suppression of YAP activity. Methods: NTRC 0652-0 is a novel tyrosine kinase inhibitor with specificity for LCK. In vitro pharmacodynamics were defined. In cholangiocarcinoma cells, specificity for LCK inhibition was determined by mass spectrometry-based unbiased analysis of the tyrosine phosphoproteome following treatment with NTRC 0652-0 or genetic deletion of LCK. YAP phosphorylation and cotranscriptional activity, cell viability, and apoptosis were assessed. A panel of eight cholangiocarcinoma patient derived organoids (PDO) were characterized and tested for sensitivity to NTRC 0652-0. Two patient-derived xenograft (PDX) models bearing FGFR2-rearrangements were utilized for in vivo assessment of pharmacokinetics, toxicity, and efficacy. Results: NTRC 0652-0 demonstrated selectivity for LCK inhibition in vitro and in cholangiocarcinoma cells. NTRC 0652-0 treatment led to YAP inhibition and apoptotic cell death in cholangiocarcinoma cell lines, associated with inhibition of MCL1 expression. PDOs demonstrated variable sensitivity to NTRC 0652-0, correlated with basal YAP tyrosine phosphorylation and drug-induced suppression of YAP co-transcriptional activity. FGFR2-altered cholangiocarcinoma was also identified as a subset with enrichment of YAP target genes. Cells from an FGFR2-altered PDX were sensitive to NTRC 0652-0 despite being primarily resistant to pemigatinib. In two PDX models of FGFR2-altered cholangiocarcinoma, daily oral treatment with NTRC 0652-0 inhibited YAP tyrosine phosphorylation, and resulted in stable plasma and tumor drug levels, acceptable toxicity, and significantly decreased tumor growth. Conclusions: A novel LCK inhibitor, NTRC 0652-0, reduced YAP signaling and demonstrated preclinical efficacy in multiple patient-derived models of cholangiocarcinoma. LCK is a novel therapeutic target in cholangiocarcinoma, and YAP activation or FGFR2-alteration are potential biomarkers for response.

Prostate‐specific membrane antigen (PSMA) is a validated target for molecular diagnostics and targeted radionuclide therapy. Our purpose was to evaluate PSMA expression in hepatocellular carcinoma (HCC), cholangiocarcinoma (CCA), and hepatic adenoma (HCA); investigate the genetic pathways in HCC associated with PSMA expression; and evaluate HCC detection rate with 68Ga‐PSMA‐11 positron emission tomography (PET). In phase 1, PSMA immunohistochemistry (IHC) on HCC (n = 148), CCA (n = 111), and HCA (n = 78) was scored. In a subset (n = 30), messenger RNA (mRNA) data from the Cancer Genome Atlas HCC RNA sequencing were correlated with PSMA expression. In phase 2, 68Ga‐PSMA‐11 PET was prospectively performed in patients with treatment‐naïve HCC on a digital PET scanner using cyclotron‐produced 68Ga. Uptake was graded qualitatively and semi‐quantitatively using standard metrics. On IHC, PSMA expression was significantly higher in HCC compared with CCA and HCA (P < 0.0001); 91% of HCCs (n = 134) expressed PSMA, which principally localized to tumor‐associated neovasculature. Higher tumor grade was associated with PSMA expression (P = 0.012) but there was no association with tumor size (P = 0.14), fibrosis (P = 0.35), cirrhosis (P = 0.74), hepatitis B virus (P = 0.31), or hepatitis C virus (P = 0.15). Overall survival tended to be longer in patients without versus with PSMA expression (median overall survival: 4.2 vs. 1.9 years; P = 0.273). FGF14 (fibroblast growth factor 14) mRNA expression correlated positively (rho = 0.70; P = 1.70 × 10‐5) and MAD1L1 (Mitotic spindle assembly checkpoint protein MAD1) correlated negatively with PSMA expression (rho = −0.753; P = 1.58 × 10‐6). Of the 190 patients who met the eligibility criteria, 31 patients with 39 HCC lesions completed PET; 64% (n = 25) lesions had pronounced 68Ga‐PSMA‐11 standardized uptake value: SUVmax (median [range] 9.2 [4.9‐28.4]), SUVmean 4.7 (2.4‐12.7), and tumor‐to‐liver background ratio 2 (1.1‐11). Conclusion: Ex vivo expression of PSMA in neovasculature of HCC translates to marked tumor avidity on 68Ga‐PSMA‐11 PET, which suggests that PSMA has the potential as a theranostic target in patients with HCC.

Abstract Background and aims Biliary tract cancers (BTCs) are uncommon, but highly lethal, gastrointestinal malignancies. Gemcitabine/cisplatin is a standard‐of‐care systemic therapy, but has a modest impact on survival and harbors toxicities, including myelosuppression, nephropathy, neuropathy, and ototoxicity. Whereas BTCs are characterized by aberrations activating the cyclinD1/cyclin‐dependent kinase (CDK)4/6/CDK inhibitor 2a/retinoblastoma pathway, clinical use of CDK4/6 inhibitors as monotherapy is limited by lack of validated biomarkers, diffident preclinical efficacy, and development of acquired drug resistance. Emerging studies have explored therapeutic strategies to enhance the antitumor efficacy of CDK4/6 inhibitors by the combination with chemotherapy regimens, but their mechanism of action remains elusive. Approach and results Here, we report in vitro and in vivo synergy in BTC models, showing enhanced efficacy, reduced toxicity, and better survival with a combination comprising gemcitabine/cisplatin and CDK4/6 inhibitors. Furthermore, we demonstrated that abemaciclib monotherapy had only modest efficacy attributable to autophagy‐induced resistance. Notably, triplet therapy was able to potentiate efficacy through elimination of the autophagic flux. Correspondingly, abemaciclib potentiated ribonucleotide reductase catalytic subunit M1 reduction, resulting in sensitization to gemcitabine. Conclusions As such, these data provide robust preclinical mechanistic evidence of synergy between gemcitabine/cisplatin and CDK4/6 inhibitors and delineate a path forward for translation of these findings to preliminary clinical studies in advanced BTC patients.

Objectives This study sought to better characterize patient characteristics, treatment options, and outcomes for osteoclast-like giant cell carcinoma of the pancreas, a rare subtype of pancreatic adenocarcinoma. Methods This is a retrospective study of all patients with osteoclast-like giant cell carcinoma of pancreatic origin treated at Mayo Clinic from 2000 to present. Baseline patient characteristics, treatment modalities utilized, and outcomes were compiled. Overall survival (OS) and progression-free survival were assessed using Kaplan-Meier analysis with a significance level of P ≤ 0.05. Results Fifteen patients met criteria for inclusion. Four patients had distant metastases at diagnosis, the remaining 11 with locoregional disease. Median OS for the entire cohort was 11 months. Metastatic disease was associated with significantly shorter OS (3.5 vs 14.1 months; P = 0.005). Three patients had no evidence of disease at time of analysis; all 3 were treated with complete resection followed by adjuvant chemotherapy. Conclusions Osteoclast-like giant cell carcinoma of the pancreas is an aggressive malignancy with poor prognosis. For patients with locoregional disease, surgical resection followed by adjuvant chemoradiation may play a role in extended disease-free survival. Metastatic disease presents a challenging entity to treat with little data to support any effective chemotherapy regimens.

Abstract Background and Objectives Pancreatic neuroendocrine tumors (pNETs) in patients with hereditary cancer syndromes are typically multifocal. In contrast, sporadic pNETs are usually unifocal and the incidence of multifocal sporadic pNETs is unknown. The primary aim of this study was to investigate the incidence of multifocality in sporadic pNETs and any associated effect on recurrence risk and survival. Methods Patients who underwent resection of pNETs at Mayo Clinic from 2000 to 2019 were identified and clinical data were obtained from medical records. Syndromic disease was defined as pNETs arising in the setting of a hereditary cancer syndrome. Statistical comparisons were made using χ 2 , Fisher's exact, and Kruskal–Wallis tests and survival was assessed using the Kaplan–Meier method. Results Six hundred and sixty‐one patients with sporadic pNETs and fifty‐nine with syndromic pNETs were identified. Multifocal disease was present in 4.8% of sporadic patients and 84.7% of syndromic patients ( p < .001). Within patients with sporadic pNETs, clinicopathologic features and recurrence‐free and overall survival were similar between patients with unifocal and multifocal disease. Conclusions Multifocal sporadic pNETs are rare and multifocality is not associated with worse survival or increased recurrence risk. Patients with multifocal sporadic pNETs can likely be safely managed with a combination of resection and observation as indicated for each tumor.

AbstractIntroductionAlthough surgical resection is necessary, it is not sufficient for long‐term survival in pancreatic ductal adenocarcinoma (PDAC). We sought to evaluate survival after up‐front surgery (UFS) in anatomically resectable PDAC in the context of three critical factors: (A) margin status; (B) CA19‐9; and (C) receipt of adjuvant chemotherapy.MethodsThe National Cancer Data Base (2010–2015) was reviewed for clinically resectable (stage 0/I/II) PDAC patients. Surgical margins, pre‐operative CA19‐9, and receipt of adjuvant chemotherapy were evaluated. Patient overall survival was stratified based on these factors and their respective combinations. Outcomes after UFS were compared to equivalently staged patients after neoadjuvant chemotherapy on an intention‐to‐treat (ITT) basis.ResultsTwelve thousand and eighty‐nine patients were included (n = 9197 UFS, n = 2892 ITT neoadjuvant). In the UFS cohort, only 20.4% had all three factors (median OS = 31.2 months). Nearly 1/3rd (32.7%) of UFS patients had none or only one factor with concomitant worst survival (median OS = 14.7 months). Survival after UFS decreased with each failing factor (two factors: 23 months, one factor: 15.5 months, no factors: 7.9 months) and this persisted after adjustment. Overall survival was superior in the ITT‐neoadjuvant cohort (27.9 vs. 22 months) to UFS.ConclusionDespite the perceived benefit of UFS, only 1‐in‐5 UFS patients actually realize maximal survival when known factors highly associated with outcomes are assessed. Patients are proportionally more likely to do worst, rather than best after UFS treatment. Similarly staged patients undergoing ITT‐neoadjuvant therapy achieve survival superior to the majority of UFS patients. Patients and providers should be aware of the false perception of ‘optimal’ survival benefit with UFS in anatomically resectable PDAC.

Pancreatic ductal adenocarcinoma is a recalcitrant tumor with minimal response to conventional chemotherapeutic approaches. Oncogenic signaling by activated tyrosine kinases has been implicated in cancers resulting in activation of diverse effector signaling pathways. Thus, the discovery of aberrantly activated tyrosine kinases is of great interest in developing novel therapeutic strategies in the treatment and management of pancreatic cancer. Patient-derived tumor xenografts (PDXs) in mice serve as potentially valuable preclinical models as they maintain the histological and molecular heterogeneity of the original human tumor. Here, we employed high-resolution mass spectrometry combined with immunoaffinity purification using anti-phosphotyrosine antibodies to profile tyrosine phosphoproteome across 13 pancreatic ductal adenocarcinoma PDX models. This analysis resulted in the identification of 1199 tyrosine-phosphorylated sites mapping to 704 proteins. The mass spectrometric analysis revealed widespread and heterogeneous activation of both receptor and non-receptor tyrosine kinases. Preclinical studies confirmed ephrin type-B receptor 4 (EphB4) as a potential therapeutic target based on the efficacy of human serum albumin-conjugated soluble EphB4 in mice bearing orthotopic xenografts. Immunohistochemistry-based validation using tissue microarrays from 346 patients with PDAC showed significant expression of EphB4 in >70% of patients. In summary, we present a comprehensive landscape of tyrosine phosphoproteome with EphB4 as a promising therapeutic target in pancreatic ductal adenocarcinoma.

Abstract Background: Mixed acinar neuroendocrine carcinoma (MANEC) of the pancreas is an extremely rare histopathologic subtype. MANEC is an amphicrine malignancy composed of at least 25% of acinar cell carcinoma (PACC) and neuroendocrine carcinoma (pNET). No standard systemic treatment is available for MANEC and current recommendations are derived from standard of care for PACC or pNET. Platinum based regimens have been approved for PACC while targeted agents (sunitinib and everolimus) are approved for pNET. This study aimed to investigate the antitumor effect and toxicities of both platinum based and targeted agents on a successfully first described patient derived xenograft (PDX) of metastatic MANEC of the pancreas. Methods: Several generations of PDX from a surgically resected metastatic MANEC of the pancreas was histologically validated. PDX-MANEC tumors were heterotopically implanted into NOD/SCID mice and randomized into 6 groups (4 mice each) as in table 1. The study continued for 3 weeks. Antitumor efficacy was determined based on change in the tumor volume. Toxicity was assessed change in mice weights. Results: Table 1 shows the changes in tumor volume and mouse weight in all study arms compared to controls end point. Only FOLFIRINOX and gemcitabine/cisplatin arms revealed overall decrease in tumor volume lower than starting volume in controls. All arms other than sunitinib, led to decrease in overall tumor volume compared to control tumor volumes over time. All treatment arms led to lower trending mouse weights (toxicities) except sunitinib, however only the everolimus arm had statistical significance. Conclusion: Both platinum-based regimens and everolimus revealed significant tumor control. No synergistic effect observed in sunitinib/everolimus. Preclinical studies are undergoing to investigate other effective and less toxic therapeutics for MANEC. Table 1.Delta in tumor volumes of PDX-MANEC-P after multiple platinum-based chemotherapy regimens and targeted therapeutics from the standard of care for other pancreatic cancer subtypes including PACC and pNET.ControlFOLFIRINOXGemcitabine and CisplatinSunitinibEverolimusSunitinib and EverolimusDelta in tumor volume in mm3 at the study end (mean ± standard deviation)3225 ± 2054-609.3 ± 351.8-654.7 ± 92.561583 ± 950.3-178.1 ± 61.58-160.8 ± 86.55P value (column group vs. control)&lt;0.0010.0030.0950.0010.001Delta in mice weight in grams at the study end (mean ± standard deviation)0.84 ± 1.68-1.26 ± 0.84-8.5 ± 1.630.86 ± 1.22-2.2 ± 1.61-1.42 ± 1.27P value (column group vs. control)0.058&lt;0.0010.9810.0070.050 Citation Format: Amro M. Abdelrahman, Jennifer A. Yonkus, Roberto Alva-Ruiz, Jennifer L. Leiting, Nellie A. Campbell, Lewis R. Roberts, Michael L. Kendrick, David M. Nagorney, Sean P. Cleary, Travis E. Grotz, Gregory J. Gores, Jun Yin, Rondell P. Graham, Thorvardur R. Halfdanarson, Rory L. Smoot, Mark J. Truty. Preclinical efficacy of platinum based chemotherapy regimens and targeted therapeutics in the first successful described patient derived xenograft model of mixed acinar neuroendocrine carcinoma of the pancreas [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 413.

Abstract Cancer cells increase lipid metabolism by up-regulation lipogenic enzymes such as stearoyl-CoA desaturase 1 (SCD1). It is the rate-limiting enzyme in the de novo synthesis of fatty acids (FAs), catalyzing the conversion and biosynthesis of saturated fatty acids (SFAs) into monounsaturated fatty acids (MUFAs). We developed SSI-4, a novel highly specific SCD1 inhibitor. The anti-tumor activity ofSSI-4 was examined against different cholangiocarcinoma (CCA) models that included human and mouseCCA cell lines, CCA patient-derived xenografts (PDX) NOD SCID gamma (NSG) mouse models, and CCA PDX derived organoids. Using cell proliferation assay, we examined twelve CCA cell lines and 4 were very sensitive to SSI-4 (1-40 nM IC50). The remaining CCA cell lines showed moderate to no sensitivity to SSI-4. Studies at the protein level have shown that SCD1 is present in both sensitive and non-sensitive cells. Thus, SCD1 must be present in sensitive cells but is not predictive of response to an SCD1 inhibitor. In an effort to identify biomarkers predicting response to therapy, we performed assays, such as lipid, lactate, or energy phenotype which identified elevated lactate and unsaturated fatty acid levels as well as a glycolytic energy phenotype. In vivo, CCA PDX models have demonstrated that SSI-4strongly inhibited tumor growth. Immunohistochemical analysis of CCA tumors revealed elevated glycolytic genes in sensitive cells and thus, a potential predictive biomarker. We are currently pursuing a better understanding of the relationship between glycolysis and fatty acid synthesis/metabolism. We are also testing novel combination therapy of SSI-4 with the standard of care and other drugs to develop novel treatments for CCA. Using CCA PDX derived organoids showed synergistic antitumor activity of SSI-4in combination with cancer standard of care drugs such as gemcitabine and cisplatin. Collectively, we expect to develop a biomarker-driven selection of CCA patients sensitive to SCD1 inhibitors and novel SSI-4 combination therapy leading to improved patient survival. Citation Format: Justyna J. Gleba, Laura A. Marlow, Erin E. Miller, James L. Miller, Aylin Alasonyalilar-Demirer, Yi Guo, Kabir Mody, Lewis R. Roberts, Steven R. Alberts, Mark J. Truty, Tushar C. Patel, John A. Copland. Defining stearoyl-CoA desaturase 1 as a molecular therapeutic target against cholangiocarcinoma [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 1004.