Neuroendocrine Markers Synaptophysin Research Articles

Olfactory Neuroblastoma With Divergent Differentiation: Contemporary Management of Unusual Pathology and Literature Review

Olfactory neuroblastoma (ON; Esthesioneuroblastoma) is a malignant tumor that arises from the olfactory neuroepithelium. Very rarely, ON can histologically display a biphenotypic pattern, with only 7 cases reported in the literature to date. We describe a case of this poorly understood entity and review the patient’s histology, pathology, and treatment. An 85-year-old man presented with endoscopy and imaging findings of a sinonasal mass arising from the olfactory cleft. Biopsy and further pathological review established Hyams grade 3 and Kadish stage C tumor. The patient underwent combined endoscopic and open-approach resection due to the involvement of the nasal bone and subsequent adjuvant radiation therapy. Specimen contained Flexner-Wintersteiner rosettes and stained positive for the neuroendocrine marker synaptophysin consistent with ON. A second, epithelial component pattern of goblet cells and mucin was observed suggesting divergent differentiation. We are the first group to report next-generation sequencing of this tumor, which revealed a pathogenic mutation in PIK3CA and a likely pathogenic variant in RUNX1 (AML1). ON with divergent differentiation is very rare, and more robust studies characterizing molecular drivers and pathology may aid in clinical management.

A Frozen Tale of Two Tumors in the Pancreas: An Intraoperative Diagnostic Challenge of Clear Cell Pancreatic Neuroendocrine Tumor Mimicking Metastatic Clear Cell Renal Cell Carcinoma

Abstract Introduction/Objective Clear cell pancreatic neuroendocrine tumor (PanNET) is rare, with very few cases reported in the literature, and is often found in patients with von Hippel-Lindau (VHL) disease. The differential for this entity includes ectopic adrenal tissue and metastatic clear cell neoplasms. These challenges during intraoperative consultation add to the complexity of diagnosing this condition. Distinguishing it from other pancreatic lesions requires careful examination and further ancillary testing. Methods/Case Report We report a case of clear cell PanNET mimicking metastatic clear cell renal cell carcinoma (RCC) in a 56-year-old woman with recently diagnosed VHL. She presented with renal masses and a pancreatic lesion with previous biopsy showing conventional NET. Intraoperative consultation during her pancreaticoduodenectomy identified a 1.3 cm well-circumscribed, golden-yellow solid mass. Further characterization was deferred to permanent sections given the diagnostic challenge of distinguishing between a PanNET and metastatic clear cell RCC. Permanent sections showed nests of clear cells with prominent nucleoli. The neoplastic cells were immunohistochemically positive for neuroendocrine markers synaptophysin, chromogranin and insulinoma- associated protein 1(INSM1), as well as pankeratin and PAX8, with a Ki-67% proliferation index <1%, consistent with low-grade clear cell PanNET. Clear cell RCC marker, CA-IX, as well as adrenocortical marker SF1, were negative. The combination of immunohistochemical stains is key for making the diagnosis. The previously biopsied conventional PanNET was also identified, which was positive for neuroendocrine markers and had a Ki-67 proliferation index <1%. Results (if a Case Study enter NA) NA Conclusion Given the rarity of this entity and its strong association with VHL, clear cell PanNET should be considered in the intraoperative consultation to inform clinical management. Furthermore, literature review indicates that metastatic RCC in the pancreas is exceptionally rare in VHL patients and typically of low nuclear grade, suggesting low metastatic potential. Thus, this case emphasizes the diagnostic challenges and the necessity of deferring to permanent sections with immunohistochemical work-up during intraoperative consultation or when assessing small biopsies.

Rare histologic transformation of a CTNNB1 (β-catenin) mutated prostate cancer with aggressive clinical course

BackgroundCatenin (Cadherin-Associated Protein), Beta 1 (CTNNB1) genomic alterations are rare in prostate cancer (PCa). Gain-of-function mutations lead to overexpression of β-catenin, with consequent hyperactivation of the Wnt/β-catenin signaling pathway, implicated in PCa progression and treatment resistance. To date, successful targeted treatment options for Wnt/β-catenin - driven PCa are lacking.MethodsWe report a rare histologic transformation of a CTNNB1 (β-catenin) mutated metastatic castration resistant prostate cancer (mCRPC), clinically characterized by highly aggressive disease course. We histologically and molecularly characterized the liver metastatic tumor samples, as well as successfully generated patient-derived organoids (PDOs) and patient-derived xenograft (PDX) from a liver metastasis. We used the generated cell models for further molecular characterization and drug response assays.ResultsImmunohistochemistry of liver metastatic biopsies and PDX tumor showed lack of expression of typical PCa (e.g., AR, PSA, PSAP, ERG) or neuroendocrine markers (synaptophysin), compatible with double-negative CRPC, but was positive for nuclear β-catenin expression, keratin 7 and 34βE12. ERG rearrangement was confirmed by fluorescent in situ hybridization (FISH). Drug response assays confirmed, in line with the clinical disease course, lack of sensitivity to common drugs used in mCRPC (e.g., enzalutamide, docetaxel). The casein kinase 1 (CK1) inhibitor IC261 and the tankyrase 1/2 inhibitor G700-LK showed modest activity. Moreover, despite harbouring a CTNNB1 mutation, PDOs were largely insensitive to SMARCA2/4- targeting PROTAC degraders and inhibitor.ConclusionsThe reported CTNNB1-mutated mCRPC case highlights the potential challenges of double-negative CRPC diagnosis and underlines the relevance of further translational research to enable successful targeted treatment of rare molecular subtypes of mCRPC.

Abstract 4234: Personalized therapeutic screening in patient derived organoids for gastroenteropancreatic neuroendocrine tumors

Abstract Introduction Gastroenteropancreatic Neuroendocrine Neoplasms (GEP-NENs) are a rare subset of cancers which nevertheless are a rising health burden. Development of new therapies suffers from several bottlenecks, including low patient accrual and poor understanding of tumor characteristics. Patient tumor organoids (PTOs) are a novel model capable of improving screening of patient tissue in an accurate, standardized, and high-throughput capacity. In this study, we utilized patient tumors for creation of high-fidelity PTOs from a variety of GEP-NEN primary origins to evaluate therapy responses. Methods Tumors from patients undergoing clinically guided surgeries were processed within two hours of resection and dissociated into single-cell suspension. Cells were encapsulated into Matrigel and cultured into two groups. The first group was grown for 10 days to assess viability then treated with a panel of clinically approved therapies and novel treatments recommended by high throughput NEN cell line screening for treatment sensitivity using the MIPE 5.0 library. The second group was grown for long-term expansion and biobanking, followed by characterization using immunohistochemistry and genetic profiling to ensure tumor cell maintenance. Results From March 2023-November 2023, 14 patients provided 30 tumors for PTO development. These included small intestine (n=5), pancreatic (n=8), and gastric (n=1) neuroendocrine tumors. Long-term culture (>3 passages) was successful for 23/30 (77%) specimens, with passage timing related to tumor grade. PTOs maintained immunohistochemical characteristics of the parent tumor types and demonstrated similar genetic profiles, including neuroendocrine tumor cell markers synaptophysin, chromogranin A, and matched grade-based Ki67 proliferative index. The early-stage therapeutic screening was performed for 12/14 (86%) patients, demonstrating tumor grade dependent treatment efficacy and showing clinically dose relevant sensitivity towards approved small molecule inhibitor therapies including cabozantinib and sunitinib in a patient and tumor origin-dependent manner. Testing of effective therapy classes recommended by cell line treatment panels suggested high level treatment efficacy for proteosome inhibitors, MEK inhibitors, and topoisomerase inhibitors. Conclusion Development of GEP-NEN PTOs is feasible for long term culture and standard of care therapy testing. Further study demonstrated the successful application of novel therapeutic options in PTOs for patients with GEP-NENs. Citation Format: Steven D. Forsythe, Srujana V. Yellapragada, Stephen G. Andrews, Jaydira del Rivero, Jonathan M. Hernandez, Naris Nilubol, James P. Madigan, Samira M. Sadowski. Personalized therapeutic screening in patient derived organoids for gastroenteropancreatic neuroendocrine tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4234.

The Significance of Insulinoma-Associated Protein 1 in the Pathological Diagnosis of Small-Cell Lung Cancer in Biopsy Specimens.

Objective: Our purpose was to investigate the clinicopathological diagnostic value of immunohistochemical antibody for insulinoma-associated protein 1 (INSM1) in biopsy specimens of SCLC. Methods: Biopsy specimens of SCLC diagnosed at the pathology department of Tangshan Gongren Hospital from January 2022 to June 2023 were selected. INSM1 expression was detected and compared with conventional neuroendocrine markers synaptophysin (SYP), chromogranin A (CHGA), and CD56 regarding expression sensitivity and specificity. Results: The sensitivity of INSM1 expression was significantly higher than that of CHGA (95% vs 50%, P = .000), but there was no statistically significant difference in the specificity of INSM1, SYP, CHGA, and CD56 expression (100% vs 94% vs 98% vs 92%, respectively, P = .241, 1.000, .126). Conclusions: INSM1 antibody shows high sensitivity and specificity in the expression of SCLC and serves as a reliable immunohistochemical marker in the clinicopathological diagnosis of SCLC in biopsy specimens.

Comparison of INSM1 immunostaining with established neuroendocrine markers Synaptophysin and Chromogranin A in over 14,000 neuroendocrine and non-neuroendocrine tumors

Abstract Introduction/Objective INSM1 is a transcription factor protein which is increasingly used as an immunohistochemical marker for neuroendocrine differentiation. Methods/Case Report To determine the prevalence of INSM1 expression in tumors and its expression pattern in normal tissues, tissue microarrays containing 14,908 samples from 117 different tumor types/subtypes as well as 76 different normal tissues were analyzed by immunohistochemistry. Results (if a Case Study enter NA) INSM1 was positive in 89.2% of 471 neuroendocrine neoplasms (NEN) and in 3.5% of 11,815 non-neuroendocrine neoplasms that were successfully analyzed. INSM1 positivity occurred in 59 non- neuroendocrine tumor entities, of which 15 entities contained at least one strongly positive case. Comparison with synaptophysin and chromogranin A revealed that in NEN, synaptophysin showed the highest sensitivity (93.3%), followed by INSM1 (89.2%) and chromogranin A (87.5%). In neuroendocrine carcinomas (NEC), sensitivity was highest for INSM1 (88.0%), followed by synaptophysin (86.5%) and chromogranin A (66.4%). The additional use of INSM1 increased the sensitivity for detecting neuroendocrine differentiation in NEN from 88.2% (synaptophysin and chromogranin A) to 91.2% (synaptophysin, chromogranin A and INSM1). Conclusion Our study shows that INSM1 is a useful additional marker for neuroendocrine differentiation that shows a particularly high sensitivity in NEC. The additional use of INSM1 results in a higher sensitivity for the identification of a neuroendocrine differentiation than what can be obtained by using only synaptophysin and chromogranin A.

Comparison of INSM1 immunostaining with established neuroendocrine markers synaptophysin and chromogranin A in over 14,000 neuroendocrine and non-neuroendocrine tumors

INSM1 is a transcription factor protein which is increasingly used as an immunohistochemical marker for neuroendocrine differentiation. To determine the prevalence of INSM1 expression in tumors and its expression pattern in normal tissues, tissue microarrays containing 14,908 samples from 117 different tumor types/subtypes as well as 76 different normal tissues were analyzed by immunohistochemistry. INSM1 was positive in 89.2% of 471 neuroendocrine neoplasms (NEN) and in 3.5% of 11,815 non-neuroendocrine neoplasms that were successfully analyzed. At least an occasional weak INSM1 positivity was observed in 59 different non-neuroendocrine tumor entities, of which 15 entities contained at least one case with strong INSM1 staining. A comparison with synaptophysin and chromogranin A staining revealed that in NEN, synaptophysin showed the highest sensitivity (93.3%), followed by INSM1 (89.2%) and chromogranin A (87.5%). In neuroendocrine carcinomas (NEC), sensitivity was highest for INSM1 (88.0%), followed by synaptophysin (86.5%) and chromogranin A (66.4%). If INSM1 was used as an additional marker, the sensitivity for detecting neuroendocrine differentiation in NEN increased from 96.6% (synaptophysin and chromogranin A) to 97.2% (synaptophysin, chromogranin A and INSM1). Our study shows that INSM1 is a useful additional marker for neuroendocrine differentiation with high sensitivity, particularly in NEC.

High expression of insulinoma-associated protein 1 (INSM1) distinguishes colorectal mixed and pure neuroendocrine carcinomas from conventional adenocarcinomas with diffuse expression of synaptophysin.

Complementary to synaptophysin and chromogranin A, insulinoma-associated protein 1 (INSM1) has emerged as a sensitive marker for the diagnosis of neuroendocrine neoplasms. Since there are no comparative data regarding INSM1 expression in conventional colorectal adenocarcinomas (CRCs) and colorectal mixed adenoneuroendocrine carcinomas/neuroendocrine carcinomas (MANECs/NECs), we examined INSM1 in a large cohort of conventional CRCs and MANECs/NECs. In conventional CRC, we put a special focus on conventional CRC with diffuse expression of synaptophysin, which carry the risk of being misinterpreted as a MANEC or a NEC. We investigated INSM1 according to the immunoreactive score in our main cohort of 1,033 conventional CRCs and 21 MANECs/NECs in comparison to the expression of synaptophysin and chromogranin A and correlated the results with clinicopathological parameters and patient survival. All MANECs/NECs expressed INSM1, usually showing high or moderate expression (57% high, 34% moderate, and 9% low), which distinguished them from conventional CRCs, which were usually INSM1 negative or low, even if they diffusely expressed synaptophysin. High expression of INSM1 was not observed in conventional CRCs. Chromogranin A was negative/low in most conventional CRCs (99%), but also in most MANECs/NECs (66%). Comparable results were observed in our independent validation cohorts of conventional CRC (n = 274) and MANEC/NEC (n = 19). Similar to synaptophysin, INSM1 expression had no prognostic relevance in conventional CRCs, while true MANEC/NEC showed a highly impaired survival in univariate and multivariate analyses (e.g. disease-specific survival: p < 0.001). MANECs/NECs are a highly aggressive variant of colorectal cancer, which must be reliably identified. High expression of INSM1 distinguishes MANEC/NEC from conventional CRCs with diffuse expression of the standard neuroendocrine marker synaptophysin, which do not share the same dismal prognosis. Therefore, high INSM1 expression is a highly specific/sensitive marker that is supportive for the diagnosis of true colorectal MANEC/NEC.

Hepatoid adenocarcinoma masquerading as adrenocortical carcinoma in an elderly male

Background: Hepatoid adenocarcinoma is a malignant lesion arising from an anatomic site other than the liver with histological features similar to hepatocellular carcinoma. Hepatoid adenocarcinoma has been reported from various anatomical sites, but adrenal hepatoid adenocarcinomas are extremely rare.Case Description: A 70-year-old male with well-controlled hypertension presented with chronic abdominal pain and was found to have a large left-sided supra renal mass. The lesion showed radiological features suggestive of malignancy with local invasion. There was biochemical evidence of primary aldosteronism and a non-suppressed overnight dexamethasone suppression test. He underwent open left-sided adrenalectomy: histologically the tumour demonstrated features typical of a hepatoid carcinoma with bile production and immunohistochemical staining for Hep Par-1 and CD10 demonstrating a hepatic canalicular pattern. Adrenocortical-specific immunohistochemical markers (Inhibin and melan A) and neuroendocrine markers (synaptophysin and chromogranin) were negative. He did not have clinical or biochemical evidence of cirrhosis, Hepatitis B or C infection. Triple-phase CT scanning of the abdomen before resection of the adrenal lesion and five months following surgery did not show any significant lesion in the liver suggestive of primary hepatocellular carcinoma, except an 8 mm non enhancing benign-appearing cystic lesion.Conclusion: Most hepatoid adenocarcinomas originate from the gastro-intestinal tract. We present a patient with hepatoid adenocarcinoma masquerading as adrenocortical carcinoma due to misleading imaging and biochemical evidence. Very little is known about their pathogenesis, especially those with an atypical site of origin such as the adrenal gland.

Two Cases of Duodenal Neuroendocrine (Carcinoid) Tumor Concurrent with Invasive Gastric Adenocarcinoma and Pancreatic Intraductal Papillary Mucinous Neoplasm

Abstract Introduction/Objective Neuroendocrine (carcinoid) tumors of the duodenum are extremely rare and account for only 2-4% of gastrointestinal tract neuroendocrine tumors. We report two cases of incidentally discovered duodenal neuroendocrine tumor which respectively occurred in a patient with invasive gastric adenocarcinoma and in a patient with pancreatic intraductal papillary mucinous neoplasm (IPMN). Methods/Case Report The first case involved a 60-year old male who presented with melena and symptomatic anemia and was found on EGD to have a large circumferential mass at the gastric antrum/pylorus, which was revealed to be an invasive gastric adenocarcinoma on biopsy. At the time of distal gastrectomy for this malignancy, a concurrent, 0.6 cm-sized, low grade neuroendocrine tumor was incidentally discovered at the duodenal margin. The duodenal tumor consisted of nests and trabeculae of monotonous cells with ovoid/round nuclei, finely speckled “salt and pepper” chromatin, and low mitotic rate (&amp;lt; 2 mitoses/2 mm2), and invaded only into submucosa. The tumor cells were strongly and diffusely positive for neuroendocrine markers chromogranin, synaptophysin, and CD56, positive for CK7 and CDX2, negative for CK20, and with low Ki67 proliferation index (&amp;lt; 3%). The second case involved a 69-year old female who presented with epigastric pain and was found on abdominal CT to have a 4 cm-sized cystic mass in the pancreatic head. While biopsy of the pancreatic mass revealed an IPMN, a separate biopsy of an incidentally discovered, 1 cm-deep ulcer in the duodenum bulb revealed a low-grade neuroendocrine tumor with similar morphologic features, mitotic rate, and immunohistochemical staining pattern as the prior case while also negative for IMP3 stain. The patient had normal serum gastrin and urine HIAA levels, but elevated serum chromogranin-A levels. Results (if a Case Study enter NA) NA. Conclusion Duodenal carcinoids are rare (1-3% of primary duodenal tumors) and are mostly non-functional and unifocal. The duodenum is also the least common site (2-4%) for GI tract neuroendocrine tumors. The pathophysiology of these tumors is still poorly understood, with the majority occurring sporadically and a minority in the setting of hereditary cancer syndromes. This report documents two very rare instances of duodenal neuroendocrine tumor incidentally discovered with invasive gastric adenocarcinoma and pancreatic IPMN.

S3080 Metastatic Merkel Cell Carcinoma Presenting as Silent Infiltrative Liver Disease: A Lesson Emphasizing the Utility of Liver Biopsy in the Exclusion of Immune Checkpoint Inhibitor-Mediated Hepatotoxicity

Introduction: Merkel cell carcinoma (MCC) is a rare neuroendocrine tumor that arises on skin exposed to the sun and occurs in patients who are Caucasian, elderly, and immunocompromised. The mortality of MCC is about 33% and disseminated disease suggests poor prognosis. Metastasis to the liver is exceedingly rare, with only few reported cases presenting as infiltrative liver disease. We present a case of MCC with “silent” infiltration in the liver effectively differentiated from suspected immune checkpoint inhibitor-mediated hepatotoxicity (IMH) by liver biopsy Case Description/Methods: A 76-year-old man with a history of metastatic MCC of the left distal thigh status-post stem cell transplant presented to a cancer care center for evaluation of fatigue, nausea, and dyspnea. He reported consuming 10 alcoholic drinks a week for many years until 2 years ago. He was started on a treatment regimen of pembrolizumab and plinabulin (a phase I microtubule inhibitor) with radiation to the liver 2 months ago. On admission, serum transaminases were elevated from baseline as indicated in Table 1. After an initial period of observation, the patient was started on oral budesonide 9 mg/d and ursodiol 1000 mg/d for empiric treatment of IMH without subsequent improvement in liver enzymes. Liver imaging was unrevealing for liver lesions. A diagnostic parenchymal liver biopsy was performed to clarify the underlying diagnosis, including possible IMH, drug-induced liver injury from plinabulin toxicity, radiation-induced liver injury, or metastatic disease. Histology showed that the tumor cells were strongly positive for synaptophysin, chromogranin, and extensive metastatic MCC. The steroids were discontinued, and the worsening hepatitis was attributed to Merkel cell liver infiltration. The patient subsequently passed away 3 months later from cancer. (Figure) Discussion: Traditionally when there is high suspicion, IMH is clinically diagnosed without expectation for liver biopsy, and steroids are often prescribed upfront. One critique regarding the role of liver biopsy may be that histological features associated with IMH are nonspecific, with features of lobular to pan-lobular hepatitis. However, the absence of an alternative findings on the biopsy confers additional confidence towards the diagnosis of IMH. This case highlights the importance of the diagnostic liver biopsy prior to immunosuppressive treatment for suspected IMH to exclude other differentials and guide the decision for empiric steroids.Figure 1.: Solid nests and lobules of poorly differentiated neuroendocrine cells are seen in this liver biopsy A) Merkel cell chromogranin - Tumor cells are strongly and diffusely positive for chromogranin. B) Merkel cell CK20- Tumor cells show perinuclear dot-like reactivity to Cytokeratin20 C) Merkel cell synaptophysin- Tumor cells are strongly and diffusely positive for neuroendocrine marker Synaptophysin. Table 1. - Prior to initiating immunotherapy, serum transaminases rose slowly but remained within 3x upper limits of normal (ULN) Laboratory Value Prior to Admission On Immunotherapy Day 1 Of Admission After Initiating Steroids Aspartate Aminotransferase (AST) 41 U/L 269 U/L 281 U/L Alanine Aminotransferase (ALT) 46 U/L 315 U/L 240 U/L Alkaline Phosphatase (ALP) 87 U/L 226 U/L 227 U/L Total Bilirubin 0.3 mg/dL 0.7 mg/dL 1.1 mg/dL International Normalized Ratio (INR) 1.06 1.15 1.18 At the end of treatment, transaminases reached >5 times ULN. No improvement was seen in liver enzymes after initiating empiric steroid therapy for suspected IMH.

Human fetal skin derived merkel cells display distinctive characteristics in vitro and in bio-engineered skin substitutes in vivo.

Human skin contains specialized neuroendocrine Merkel cells responsible for fine touch sensation. In the present study, we performed in-depth analysis of Merkel cells in human fetal back skin. We revealed that these Merkel cells expressed cytokeratin 20 (CK20), were positive for the neuroendocrine markers synaptophysin and chromogranin A, and the mechanosensitive ion channel Piezo2. Further, we demonstrated that Merkel cells were present in freshly isolated human fetal epidermal cells in vitro, and in tissue-engineered human dermo-epidermal skin substitutes 4 weeks after transplantation on immune-compromised rats. Merkel cells retained the expression of CK20, synaptophysin, chromogranin A, and Piezo2 after isolation and in culture, and in the skin substitutes after transplantation. Interestingly, we observed that in fetal skin and in skin substitutes, only Merkel cells were positive for CK8, while in culture, also non-Merkel cells showed positivity for CK8. In summary, human fetal Merkel cells showed phenotypical features confirming their cell identity. This findings are of pivotal importance for the future application of fetal tissue-engineered skin in clinics.

Synaptophysin and chromogranin A expression analysis in human tumors

The expression of the neuroendocrine markers synaptophysin and chromogranin A was analyzed by immunohistochemistry in 14,584 samples from 103 different tumor types and subtypes in a tissue microarray format. At least one of these markers was found to be positive in 96.7% of tumors from various subtypes of neuroendocrine neoplasms. In non-neuroendocrine tumors, synaptophysin and/or chromogranin A staining was seen in 6.3% (n = 584), specifically in 41 of 88 non-neuroendocrine tumor entities. Basal cell carcinomas of the skin (50% positive for chromogranin A alone) and adrenocortical carcinomas (91.7% positive for synaptophysin alone) stood out due to a frequent expression of only one specific marker. A subdivision of non-neuroendocrine neoplasms revealed “neuroendocrine differentiation” most commonly in adenocarcinomas from the female genital tract (18.9%), from pancreatico-/hepato-/biliary tract (15.8%) and the prostate (14.9%) while it was rare in urothelial (1.0%) and squamous cell carcinomas (0.6%). A comparison with clinico-pathological parameters of tumor aggressiveness did not suggest a clinical significance of neuroendocrine marker expression in 204 endometrium cancers, 249 pancreatic adenocarcinomas, 233 gastric adenocarcinomas and 1,182 colorectal adenocarcinomas. Within a cohort of 1,073 breast cancers of no special type, synaptophysin positivity was seen in 4.9% of cases and it was significantly linked to advanced tumor stage (p = 0.0427), high tumor grade (p = 0.0319) and loss of estrogen receptor expression (p = 0.0061) but unrelated to patient outcome. In conclusion, “neuroendocrine differentiation” can be observed in many different tumor types with non-neuroendocrine morphology. Evidence for a statistically significant association (p < 0.0001) between such a “neuroendocrine differentiation” and tumor aggressiveness could not be found.

Patient-derived tumor organoids for personalized medicine in a patient with rare hepatocellular carcinoma with neuroendocrine differentiation: a case report

BackgroundHepatocellular carcinoma with neuroendocrine differentiation (HCC-NED) is a very rare subtype of primary liver cancer. Treatment allocation in these patients therefore remains a challenge.MethodsWe report the case of a 74-year-old man with a HCC-NED. The tumor was surgically removed in curative intent. Histopathological work-up revealed poorly differentiated hepatocellular carcinoma (Edmondson-Steiner grade IV) with diffuse expression of neuroendocrine markers synaptophysin and chromogranin. Three months after resection, multifocal recurrence of the HCC-NED was observed. In the meantime, tumor organoids have been generated from the resected HCC-NED and extensively characterized. Sensitivity to a number of drugs approved for the treatment of HCC or neuroendocrine carcinomas was tested in vitro.ResultsBased on the results of the in vitro drug screening, etoposide and carboplatin are used as first line palliative combination treatment. With genomic analysis revealing a NTRK1-mutation of unknown significance (kinase domain) and tumor organoids found to be sensitive to entrectinib, a pan-TRK inhibitor, the patient was treated with entrectinib as second line therapy. After only two weeks, treatment is discontinued due to deterioration of the patient’s general condition.ConclusionThe rapid establishment of patient-derived tumor organoids allows in vitro drug testing and thereby personalized treatment choices, however clinical translation remains a challenge. To the best of our knowledge, this report provides a first proof-of-principle for using organoids for personalized medicine in this rare subtype of primary liver cancer.

Abstract 5715: Epigenomic profiling identifies distinct neuroendocrine subtypes in lung cancer with neuroendocrine differentiation

Abstract Lung cancer is the leading cause of cancer-related death worldwide, among which high-grade neuroendocrine tumors including small-cell lung cancer (SCLC) and pulmonary large-cell neuroendocrine carcinoma (LCNEC) are especially aggressive with a dismal prognosis. Although molecular subtypes of SCLC based on the expression of lineage transcription factors such as ASCL1, NEUROD1, POU2F3 and YAP1 have been identified and explored recently, LCNEC has not been well characterized so far, and in clinic, patients with LCNEC have been treated with both chemotherapy regimens for SCLC and regimens for non-small cell lung cancer. To make efficient treatment strategies for LCNEC, further advances in molecular characterization of LCNEC are warranted. Therefore, in this study, we aimed to elucidate the heterogeneity of neuroendocrine differentiation in lung cancer with LCNEC components by using epigenomic profiling. We investigated super-enhancer profiles of 24 formalin-fixed paraffin-embedded tumor tissues from patients with primary lung cancer containing LCNEC components. Unsupervised hierarchical clustering of these specimens using H3K27 acetylation signals over super-enhancer regions near transcriptional regulators identified four distinct clusters. Principal component analysis supported this classification. Immunohistochemical staining for ASCL1, NEUROD1, POU2F3 and YAP1 on these tissues was also performed and the staining results were compatible with our epigenomic profiling. Cluster 1 tissues were positive for ASCL1 and NKX2-1 while the high expression of POU2F3 or YAP1 was found in some samples among Cluster 3 and Cluster 4. None of the four transcription factors were positive in Cluster 2. Additional genomic profiling revealed no clear associations between the clusters and genetic alterations such as RB1 loss and STK11 loss. Neuroendocrine markers synaptophysin, chromogranin A and CD56 were found to be more commonly positive in tissues among Cluster 1. These findings suggest the existence of the distinct differentiation subtypes of lung cancer with LCNEC components, different from the previously reported classifications, which provides new insight into the regulation of neuroendocrine differentiation in lung cancer. Citation Format: Takashi Sato, Junko Hamamoto, Katsura Emoto, Takahiro Fukushima, Kai Sugihara, Masayuki Shirasawa, Yoshiro Nakahara, Satoshi Igawa, Yoshiki Murakumo, Takashi Kohno, Kouya Shiraishi, Hiroyuki Yasuda, Kenzo Soejima, Hideo Watanabe, Katsuhiko Naoki. Epigenomic profiling identifies distinct neuroendocrine subtypes in lung cancer with neuroendocrine differentiation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5715.

Adjuvant chemotherapy, extent of resection, and immunoistochemical neuroendocrine markers as prognostic factors of early-stage large-cell neuroendocrine carcinoma.

BackgroundWe investigated whether adjuvant chemotherapy, extent of resection, and immunoistochemical neuroendocrine markers affected survival of patients with the early stage of large‐cell neuroendocrine cancer.MethodsThis was a retrospective multicenter study including consecutive patients undergoing resection of node negative large‐cell neuroendocrine carcinoma. Five‐year survival and disease‐free survival rate were evaluated by the Kaplan–Meier method and the log‐rank test in relation to adjuvant chemotherapy, extent of resection, and immunoistochemical neuroendocrine markers (synaptophysin, chromogranin A, and neuron‐specific enolase).ResultsOur study population included 117 patients; 47 (40%) of these received adjuvant chemotherapy. Patients treated with adjuvant chemotherapy had better survival (74% vs. 45%, p = 0.002) and disease‐free survival (79% vs. 40%, p = 0.001) in all cases except patients with tumor <20 mm (79.5% vs. 57.4%, p = 0.43). Lobectomy compared to sublobar resection was associated with better survival (67% vs. 0.1%, p < 0.0001) and disease‐free survival (65% vs. 0.1%, p < 0.0001) also in patients with tumor <20 mm (79% vs. 28%, p = 0.001). Patients with triple‐positive neuroendocrine markers had better survival (79% vs. 35%, p = 0.0001) and disease‐free survival (69% vs. 42%, p = 0.0008). Regression analysis showed that tumor size <20 mm, lobectomy, adjuvant chemotherapy, and triple‐positive immunistochemical neuroendocrine markers were significant favorable prognostic factors for survival outcomes.ConclusionsLobectomy seems to be the management of choice in patients with large‐cell neuroendocrine cancer <20 mm while adjuvant chemotherapy should be administered only in patients with tumor >20 mm.

Exploration of INSM1 and hASH1 as additional markers in lung cytology samples of high-grade neuroendocrine carcinoma with indeterminate neuroendocrine differentiation.

Traditional neuroendocrine (NE) markers synaptophysin, chromogranin, and CD56 play an integral role in affirming the diagnosis of high-grade lung NE carcinoma, however promising markers, INSM1, and hASH1, have been identified. We investigated the utility of these markers in pulmonary cytology specimens, particularly in cases where results of traditional NE markers were equivocal. A retrospective search of cytology cases obtained via endobronchial ultrasound (EBUS)-guided FNA revealed 26 cases of high-grade lung carcinoma where an indeterminate diagnosis of small-cell lung carcinoma (SCLC) was based on equivocal IHC staining with traditional NE markers. A separate cohort of 23 cases positive for all traditional markers with a definitive diagnosis of SCLC was also selected. Cytology cellblock sections were immunostained with INSM1 and hASH1 and analyzed using H-score methodology (score range 0-300). A score of ≥95 was considered "positive." INSM1 was positive in 19/24 (79.2%) of cases of high-grade lung carcinoma with indeterminate NE differentiation, while hASH1 was positive in 6/24 (25.0%). Chromogranin was seen only focally positive (<10% of cells) in 4/24 (16.7%), synaptophysin positive in 16/24 (66.7%), and CD56 positive in 14/21 (66.7%). Among unambiguous cases, INSM1 was positive in all cases with an average score of 233.9, while hASH1 was positive in 21/23 (91.3%) with an average score of 196.3. Compared with traditional NE stains and to hASH1, INSM1 was expressed in a higher number of cases of high-grade lung NE carcinomas in cytology cellblock specimens, making it a superior, more sensitive NE marker.

Primary neuroendocrine carcinomas of the breast and neuroendocrine differentiated breast cancers: Relationship between histopathological and radiological features

PurposeThe aim of this study wasto investigate whole-breast imaging findings (mammography, ultrasonography (US), magnetic resonance imaging (MRI),clinical, and histopathological findings of primary neuroendocrine carcinomas of the breast (NEC) and neuroendocrine differentiated breast cancers (NEBC). MethodsPatients withadiagnosis of breast cancer with histopathological neuroendocrine features between the years 2010 and 2021 were retrospectively screened.The lesions were divided into two main groups depending on staining with neuroendocrine markers (synaptophysin and chromogranin A). Those showing focal staining were categorized as NEBC while those with diffuse staining as NEC.The mammography, US, and MRI of the lesionswere reviewed in consensus by two breast radiologists in order to assess imaging featuresretrospectively according to the Breast Imaging Reporting and Data System (BI-RADS) 5th lexicon.The findings were compared with breast cancers without neuroendocrine features (BC-WNE) which were randomly selected from the same database. ResultsA total of 105 lesions [NEBC (n = 44), NEC(n = 11), BC-WNE (n = 50)] were evaluated.Patients with neuroendocrine tumors were older (p < 0.001) than those with BC-WNE. Compared with BC-WNE tumors, radiological findings typical of malignancy such as irregular shape [NEBC (7/20); NEC(3/7) vs BC-WNE (35/43); p < 0.001], spiculation [NEBC (2/20); NEC(0/7) vs BC-WNE (21/43); p < 0.001], architectural distortion [(NEBC (3/24); NEC(0/9) vs BC-WNE (31/50); p < 0.001)], calcification [(NEBC (6/24), NEC(0/9) vs BC-WNE (n = 27/50); p = 0.001)] on mamography, non-parallel orientation to skin [(NEBC (n = 17/29), NEC(n = 4/9), BC-WNE (n = 35/42); p = 0.008)], acoustic shadowing [(NEBC (n = 12/29), NEC(1/9), BC-WNE (n = 29/42); p = 0.009)], axillary lymphadenopathy [(NEBC(n = 3/30), NEC(n = 1/9), BC-WNE (21/50); p < 0.001)]on US were less common features of the neuroendocrine carcinomas of breast. Aside from shape features, there was no significant difference in contrast pattern (p = 0.866), kinetic curve (p = 0.454) and diffusion restriction (p = 0.242) on MRI. ConclusionCharacteristic malignant imaging features, including irregular shape, spiculated margins, suspicious calcifications, and posterior acoustic shadowing, are uncommon in neuroendocrine carcinomas of breast. These carcinomas tend to show more benign imaging features when compared with BC-WNE.

Abstract 235: Identifying potential drug targets using patient-derived, tissue specific, gene regulatory networks

Abstract Recent developments in sequencing have allowed consortiums like ENCODE and ROADMAP to probe the epigenome of normal and cancer cell lines. Applying these approaches in a broad fashion to patient data has been difficult but a pioneering TCGA study using ATAC-Seq, an assay for probing open chromatin and thus regulatory regions, provides an opportunity to analyze cancer patient epigenomes for key regulators and potential drug targets. Utilizing this TCGA dataset, we identify the regulatory targets of open chromatin regions as assayed by ATAC-Seq and RNA-Seq across 371 patients on a patient-specific basis. We subsequently utilize transcription factor (TF) footprinting to identify regions bound by 850 TF &amp;TF-families in each patient. The final output of this approach identifies which TF regulates individual genes for every patient. Using a combination of network metrics and TF abundance, we can identify known and rare subtypes of multiple cancers with similar underlying networks. We next identify a set of TFs with greater importance and predicted activity within each cluster of samples relative to others. To expand the set of potentially druggable genes, we use data from the Achilles/DepMap project to identify a set of genes whose essentiality is most highly correlated with the set of critical TFs for each cluster, i.e. “coessential genes”. We next combine our list with two datasets, the list of known druggable genes and the Drug Interaction Database to identify compounds that target this gene set. We prioritize genes highly correlated with our key TFs' essentiality, are not pan-cancer essential genes or targeted by drugs with high inferred toxicity. We identify a set of neuroendocrine-like cancers in lung and stomach adenocarcinoma, rare disease subtypes in their representative cancers. Further investigation finds these samples expressing high levels of known neuroendocrine markers synaptophysin and PCSK1. Within well-studied cancers like breast and colon adenocarcinoma we identify separate ER+ and ER- sample clusters as well as MSI-H samples as separate and featuring different key TFs than MSI-L and MSS clusters. Our drug candidate pipeline identified known targeted drugs in breast cancer such as Tamoxifen and Fulvestrant, non-specific chemotherapies like doxorubicin in addition to the novel candidates Nelfinavir and Indinavir as potential therapeutic agents. In summary we identify key TFs in multiple cancers and novel drugs for targeting those TFs or downstream effectors. Citation Format: Andre Neil Forbes, Duo Xu, Ekta Khurana. Identifying potential drug targets using patient-derived, tissue specific, gene regulatory networks [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 235.

Urinary Large Cell Neuroendocrine Carcinoma: A Clinicopathologic Analysis of 22 Cases.

Large cell neuroendocrine carcinoma (LCNEC) of the urinary tract is a rare disease. We present a relatively large retrospective cohort of urinary LCNEC, 20 from the urinary bladder, and 2 from the ureter, from a single institution. The patients included 16 men and 6 women with a median age of 74.5 years. Most LCNEC presented at an advanced stage with tumors invading the muscularis propria and beyond (21/22). Eight cases were pure LCNEC, while 14 cases were mixed with other histologic types, including conventional urothelial carcinoma (n=9), carcinoma in situ (n=7), small cell carcinoma (n=6), and urothelial carcinoma with glandular (n=3) features. Most LCNEC expressed neuroendocrine markers synaptophysin (22/22), chromogranin (13/16), CD56 (7/7), TTF1 (8/8), and INSM1 (2/3). They were negative for common urothelial markers including HMWCK (0/3), p40/p63 (0/6), CK20 (0/10), and had variable GATA3 staining (4/8). Ki-67 stained 25% to nearly 100% tumor cell nuclei. Patient survival was associated with cancer stage, and pure LCNEC showed worse survival than mixed LCNEC. Compared with small cell carcinoma at similar stages from a prior study, LCNEC had a worse prognosis only when patients developed metastatic disease. For organ-confined LCNEC, neoadjuvant chemotherapy followed by radical resection is the treatment option to achieve long-term survival.