Graham, Rondell P.

Abstract Objectives Sclerosing pneumocytomas are rare pulmonary neoplasms that are typically benign. However, rare patients experience progressive disease, and therapy targeting specific genetic underpinnings could be an attractive therapeutic option. Recent studies have found recurrent AKT 1 mutations in sclerosing pneumocytoma, but little is known about whether oncogenic fusion genes may also be present. Methods To better understand the genetic background, 10 sclerosing pneumocytomas were subjected to next-generation sequencing cancer mutation panel testing (n = 9) and/or RNA sequencing (n = 3). The patients were all women (average age, 47 years; range, 17-74 years). Results Eight patients had solitary sclerosing pneumocytomas, while one had two tumors, and one had many bilateral tumors. Recurrent mutations were noted in genes involved in the mTOR pathway, including AKT1, PIK3R1, and PTEN. AKT1 alterations were particularly common, present in 78%. No recurrent genetic fusions were identified. The patient in our study with multiple bilateral lesions was treated with the mammalian target of rapamycin (mTOR) inhibitor everolimus, with no objective radiographic evidence of treatment response after 4 months. Conclusions Our data further support that abnormal activation of the mTOR pathway is a consistent genetic event in sclerosing pneumocytoma. This warrants further exploration to determine if mTOR pathway inhibitors may be effective in patients with metastatic or recurrent disease.

Abstract Purpose: TNF-related apoptosis inducing ligand (TRAIL) expression by immune cells contributes to antitumor immunity. A naturally occurring splice variant of TRAIL, called TRAILshort, antagonizes TRAIL-dependent cell killing. It is unknown whether tumor cells express TRAILshort and if it impacts antitumor immunity. Experimental Design: We used an unbiased informatics approach to identify TRAILshort expression in primary human cancers, and validated those results with IHC and ISH. TRAILshort-specific mAbs were used to determine the effect of TRAILshort on tumor cell sensitivity to TRAIL, and to immune effector cell dependent killing of autologous primary tumors. Results: As many as 40% of primary human tumors express TRAILshort by both RNA sequencing and IHC analysis. By ISH, TRAILshort expression is present in tumor cells and not bystander cells. TRAILshort inhibition enhances cancer cell lines sensitivity to TRAIL-dependent killing both in vitro and in immunodeficient xenograft mouse models. Immune effector cells isolated from patients with B-cell malignancies killed more autologous tumor cells in the presence compared with the absence of TRAILshort antibody (P < 0.05). Conclusions: These results identify TRAILshort in primary human malignancies, and suggest that TRAILshort blockade can augment the effector function of autologous immune effector cells. See related commentary by de Miguel and Pardo, p. 5546

Background Secondary carcinoma involving the gastrointestinal (GI) tract is an uncommon finding in biopsy specimens. The diagnosis can be challenging for tumours mimicking a primary carcinoma and when the clinical context is unknown. Methods and results A multicentre retrospective study was performed to evaluate the clinical and histological features of a series of secondary carcinoma in GI biopsies. A total of 197 cases from 190 patients (median age = 67 years; 57% females) were reviewed. In 16% of patients, the primary carcinoma was unknown. Most lesions presented endoscopically as mucosal or submucosal masses (58%). In 13%, the endoscopy was non‐suspicious for malignancy. The most common tumours were carcinomas of the breast (38%), kidney (13%), lung (12%), prostate (8%) and ovary (7%). The sites of involvement were the stomach (34%), colon (27%), rectum (18%), duodenum (13%), oesophagus (5%), jejunum (3%) and anus (0.5%). Histological patterns of infiltration were mucosal (76%), submucosal (41%), lymphatic (14%), and epithelial colonisation (8%). Submucosal infiltration was found significantly more frequently in carcinomas of the prostate (67%) and lung (62%), compared with carcinomas of the ovary (27%) and breast (23%). Histological obstructive changes were observed in 36% of all cases, with the highest rate in prostate carcinoma (53%) and the lowest rate in kidney carcinoma (8%). Conclusion Awareness of the main clinical and histological patterns of secondary carcinomas in GI tract biopsies may help pathologists to raise the possibility of this uncommon diagnosis and confirm it with the judicious use of immunohistochemistry.

Abstract Objectives Mucinous cystic neoplasm of the liver is characterized by neoplastic mucinous and/or biliary epithelium surrounded by ovarian-type stroma. Immunohistochemical studies have shown that the ovarian-type stroma expresses estrogen receptor, suggesting potential hormonal responsiveness. The molecular biology of mucinous cystic neoplasm of the liver remains poorly studied. Methods Transcriptome sequencing and immunohistochemistry were performed on a series of mucinous cystic neoplasms. Results Mucinous cystic neoplasm of the liver exhibited significantly increased RNA expression of ovarian stromal markers WT1, PR, and ER2 and sex cord stromal markers SF-1, inhibin-α, and calretinin compared with nonneoplastic liver. Immunohistochemistry confirmed the RNA-level data. Evidence for sex hormone biosynthesis was identified by significant overexpression of multiple estrogen biosynthetic enzymes. Expression of 17β-hydroxysteroid dehydrogenase 1 was confirmed immunohistochemically. Pathway analysis also identified significant upregulation of the hedgehog and Wnt pathways and significant downregulation of T-helper 1 and T-helper 2 pathways. Conclusions Mucinous cystic neoplasm of the liver recapitulates ovarian stroma at the morphologic, DNA, RNA, and protein levels. These data support the concept that this tumor likely arises from ectopic primitive gonadal tissue and/or stromal cells with capacity to transdifferentiate to ovarian cortical cells.

AbstractObjectivesSteatohepatitic hepatocellular carcinoma is a distinct variant of hepatocellular carcinoma strongly associated with underlying nonalcoholic steatohepatitis. The molecular biology of steatohepatitic hepatocellular carcinoma is not fully elucidated, and thus we aimed to investigate the molecular underpinnings of this entity.MethodsTranscriptomic analysis using RNAseq was performed on eight tumor-nonneoplastic pairs of steatohepatitic hepatocellular carcinoma with comparison to conventional hepatocellular carcinoma transcriptomes curated in The Cancer Genome Atlas. Immunohistochemistry was used to validate key RNA-level findings.ResultsSteatohepatitic hepatocellular carcinoma demonstrated a distinctive differential gene expression profile compared with The Cancer Genome Atlas curated conventional hepatocellular carcinomas (n = 360 cases), indicating the distinctive steatohepatitic hepatocellular carcinoma morphology is associated with a unique gene expression profile. Pathway analysis comparing tumor-nonneoplastic pairs revealed significant upregulation of the hedgehog pathway based on GLI1 overexpression and significant downregulation of carnitine palmitoyltransferase 2 transcript. Glutamine synthetase transcript was significantly upregulated, and fatty acid binding protein 1 transcript was significantly downregulated and immunohistochemically confirmed, indicating steatohepatitic hepatocellular carcinoma tumor cells display a zone 3 phenotype.ConclusionsSteatohepatitic hepatocellular carcinoma demonstrates a distinctive morphology and gene expression profile, phenotype of zone 3 hepatocytes, and activation of the hedgehog pathway and repression of carnitine palmitoyltransferase 2, which may be important in tumorigenesis.

Anaplastic pancreatic cancer (APC) is a rare undifferentiated variant of pancreatic ductal adenocarcinoma with poor overall survival (OS). The aim of this study was to evaluate the clinical outcomes of APC compared with differentiated pancreatic ductal adenocarcinoma. We conducted a retrospective review of all patients treated at the Mayo Clinic with pathologically confirmed APC from 1987 to 2011. After matching with control subjects with pancreatic ductal adenocarcinoma, OS was evaluated using Kaplan-Meier estimates and log-rank test. Sixteen patients were identified with APC (56.3% male, median age 57 years). Ten patients underwent exploration of whom eight underwent pancreatectomy. Perioperative morbidity was 60 per cent with no mortality. The median OS was 12.8 months. However, patients with APC who underwent resection had longer OS compared with those who were not resected, 34.1 versus 3.3 months ( P = 0.001). After matching age, sex, tumor stage, and year of operation, the median OS was similar between patients with APC and those with ductal adenocarcinoma treated with pancreatic resection, 44.1 versus 39.9 months, ( P = 0.763). Overall survival for APC is poor; however, when resected, survival is similar to differentiated pancreatic ductal adenocarcinoma.

Abstract Background: Emerging colorectal cancer trends demonstrate increased incidence and mortality in younger populations, prompting consideration of average-risk colorectal cancer screening initiation at age 45 versus 50 years. However, screening test performance characteristics in adults 45–49 years have been minimally described. To inform the biologic rationale for multi-target stool DNA (mt-sDNA) screening in younger patients, we analyzed and compared tissue levels of methylation (BMP3, NDRG4) and mutation (KRAS) markers included in the FDA-approved, mt-sDNA assay (Cologuard; Exact Sciences Corporation). Methods: Within 40–44, 45–49, and 50–64 year age groups, archived colorectal tissue specimens were identified for 211 sporadic colorectal cancer cases, 123 advanced precancerous lesions (APLs; adenomas >1 cm, high-grade dysplasia, ≥25% villous morphology, or sessile serrated polyp; 45–49 and 50–64 age groups only), and 204 histologically normal controls. Following DNA extraction, KRAS, BMP3, and NDRG4 were quantified using QuARTS assays, relative to ACTB (reference gene). Results: None of the molecular marker concentrations were significantly associated with age (P > 0.05 for all comparisons), with the exception of NDRG4 concentration in APL samples (higher in older vs. younger cases; P = 0.008). However, NDRG4 levels were also statistically higher in APL case versus normal control samples in both the 45–49 (P < 0.0001) and 50–64 (P < 0.0001) year age groups. Conclusions: Overall, these findings support the potential for earlier onset of average-risk colorectal cancer screening with the mt-sDNA assay. Impact: These novel data address an identified knowledge gap and strengthen the biologic basis for earlier-onset, average-risk screening with the mt-sDNA assay.

e16752 Background: Undifferentiated osteoclast-like giant cell carcinoma of the pancreas is an aggressive malignancy only described by a few case reports in the literature. In this study, we sought to better characterize this entity by examining patients seen at Mayo Clinic. Methods: This study identified patients with osteoclast-like giant cell carcinoma of the pancreas using Mayo Clinic databases (MN, AZ, FL) from the year 2000 to present. Patient demographics, genetic data, and treatment modalities were reviewed. Kaplan-Meier analysis was used to evaluate median overall survival (mOS) for the cohort as well as mOS and mPFS for treatment subgroups. Results: 15 patients were identified (9 female, 6 male). Median age at diagnosis was 59 years and mOS for all patients was 11.0 mos (95% CI 6.2-15.7 mos). 3 patients (20%) had metastatic disease at diagnosis with the liver being the most common site (n = 3). Metastatic disease was associated with significantly shorter OS (3.5 vs. 14.1 mos, p = 0.005; HR 7.98 [95% CI 1.43-44.4]) compared to locoregional disease (LRD). 4 patients underwent genetic testing. The most common mutation was CDKN2A (n = 3), followed by TP53 (n = 2) and KRAS (n = 2). 13/15 patients had detailed follow-up information. 6/7 patients undergoing chemotherapy received a gemcitabine-based regimen as first line: with capecitabine (n = 2), with nab-paclitaxel (n = 2), or as monotherapy (n = 2). 1 patient received FOLFIRINOX. In patients with LRD and adequate follow-up (n = 11), 8/11 underwent surgical resection and had longer OS compared to those without resection (17.0 vs. 8.4 mos, p = 0.09). No surgical patients received neoadjuvant chemotherapy. 5/8 received adjuvant chemotherapy, 2 did not undergo chemotherapy, and 1 was lost to follow-up after surgery. PFS from time of surgery was 15.3mos (95% CI 5.0-25.6mos). 6 patients (40%) were alive at time of analysis; all 6 underwent surgical resection. Conclusions: Osteoclast-like giant cell carcinoma of the pancreas is a rare malignancy with a poor prognosis, even when diagnosed at an early stage. OS for patients of all stages in this study was less than 1 year suggesting prognosis may be even worse than that of pancreatic adenocarcinoma. The optimal therapy remains unknown but most patients received similar chemotherapy as in adenocarcinoma. Patients with LRD amenable to surgical resection experienced a PFS over 1 year from surgery, possibly associated with OS benefit, however overall prognosis is poor. Further study is warranted on a larger scale to better understand disease course and treatment options.

e16705 Background: The prevalence of pancreatic neuroendocrine tumors (PNETs) has increased in the last decade. Despite being clinically asymptomatic PNETs can be biologically aggressive. There is currently no reliable non-invasive diagnostic biomarker for PNETs. In this study we aimed to identify and validate methylated DNA marker (MDM) candidates that differentiate PNET from normal pancreas. Methods: For discovery, reduced representation bisulfite sequencing (RRBS) was performed on DNA extracted from frozen normal pancreas (n = 13) and PNET (n = 51) tissues. Area under the receiver operator characteristic curve (AUC), fold-change, and p-value criteria selected candidates MDMs for blinded validation in independent FFPE tissues from primary PNET (n = 67; solid = 50, cystic = 17), normal pancreas controls (n = 24), and normal buffy coat (n = 36) using methylation specific PCR. MDM distributions in primary PNETs were compared to primary lung (n = 36) and small bowel (n = 36) NETs and metastatic PNET tissue (n = 25). The discrimination accuracy of candidate markers was summarized as the AUC with corresponding 95% confidence intervals (CI). Results: From the RRBS discovery, 31 candidate MDMs were selected for validation. Four MDMs ( SRRM3, HCN2, SPTBN4 and TMC6) achieved individual AUCs ≥0.95 in the validation set (Table). These MDMs were similarly discriminant in metastatic PNET tissue and in primary lung and small bowel NETs. Three out of these 4 MDMs perfectly differentiated PNET tissue from buffy coat with AUC of 1 and may be ideally suited for further development of a blood-based assay. Conclusions: We identified and validated novel MDMs in tissue that discriminate PNETs from controls with normal pancreas and buffy coat with high accuracy. These MDMs also differentiated metastatic PNETs from normal pancreas tissue. Further exploration of these candidate tissue MDMs in plasma can potentially guide diagnosis and management of PNETs. Funding: P30DK084567. [Table: see text]

4541 Background: Anti-VEGFR2 therapy (ramucirumab/paclitaxel [RAM/TAX]) and ICI are approved as 2nd- and 3rd-line therapy (Tx), respectively, for pts with mGEA. We unexpectedly saw durable responses in 2 pts on RAM/TAX after progression on an ICI trial (KN-059; PMID 29674442). We performed a pilot to examine the clinical activity of ICI followed by RAM/TAX. Then we retrospectively compared the outcomes of pts who received this serial Tx to pts who received RAM/TAX without prior ICI. Methods: All pts with mGEA at Mayo Clinic who received RAM/TAX (2014-19) were included (N = 87). Outcomes were best objective response rate (ORR: complete [CR] or partial response) per RECIST1.1, progression-free survival (PFS), duration of response (DOR), and overall survival (OS). Chi square and multivariate (MV) logistic and Cox regression were used. Results: 15 consecutive pts with measurable mGEA received ICI immediately followed by RAM/TAX after irRECIST progression. Most pts (95%) did not respond to ICI. Yet on RAM/TAX, 100% (15/15) had tumor reduction (range -8% to -100%) with an ORR of 73% (11/15), including 3 CRs. In these pts (who received ICI followed by RAMTAX), PFS on RAMTAX was longer than on last chemotherapy before ICI (12.3 vs 3.0 m, P < .001). Outcomes on RAM/TAX in these pts were significantly better than in pts who received RAM/TAX alone (see Table). Associations were strengthened after adjusting for total lines of Tx, line of Tx of RAM/TAX, age, and ECOG PS. Exploratory analysis of paired tumor biopsies collected pre-ICI and on RAM/TAX in a small subset revealed that the frequency of intratumoral immunosuppressive FOXP3+ Tregs decreased on RAM/TAX, whereas the frequency of antitumor CD8+ T cells was preserved. Conclusions: RAM/TAX immediately preceded by ICI was associated with significantly higher OS, ORR, and DOR than RAM/TAX alone, suggesting ICI may enhance efficacy of subsequent anti-VEGFR/taxane therapy. This novel sequence of therapy will be tested prospectively in a new randomized phase 2 trial (NCT04069273). [Table: see text]