Section 5 -

Well Differentiated Neuroendocrine Carcinoma of Small Intestine (WDNET) with Metastases in the Liver and Mesentery Robert Yoshiyuki Osamura Tokai University School of Medicine Kanagawa, Japan

Case 2 Diagnosis Well differentiated neuroendocrine carcinoma of small intestine (WDNET) with metastases in the liver and mesentery Case History The patient was a 71 year old male who had had a history of Parkinson's disease. Approximately one and half year prior to admission, he noticed abdominal pain and was pointed out by CT scan to have a mass in the abdomen. CT scan also revealed many metastatic foci in the liver. On admission, he had a palpable abdominal mass. Othewise, his physical examination, erythrocyte analysis, and biochemical data are within normal limit. unremarkable serum tumor markers showed high serum serotonin(579ng/ml). He was operated with the clinical diagnoses of retroperitoneal tumor and hepatic metastases. His tumor in the liver, small intestine and part of hepatic metastases were excised. He was first given by 5-FU 500mg(twice IA) followed by Octreotide (100μ once subcutaneous) and thereafter LAR20mg.IM(once a month). At 17 months after the operation, no recurrence was noted and no enlargement of the hepatic metastases was observed. Pathology : Immunohisochemically, CGA, synaptophysin and NSE were positive in the tumor cells. SSTR2a was also diffusely positive on the cell membrane. RNA extracted from the paraffin sections were subjected RT-PCR for SSTR2a. The tumor revealed SSTR2a mNA. Discussion Approximately a third of al gastrointestinal carcinoids are found in the small intestine, ad these constitute a third of all small intestine tumors. The majority of lesions are localized to the ileum. From 15% to 30% of patients with carcinoids are reported to have a second primary tumor, such as a carcinoma, usually within the gut. These lesions are found predominantly in middle-aged and elderly patients,who foten have a long history of symptoms, averaging about 4 years. The majority have liver metastases at the time of diagnosis. Patients usually present with features of intestinal obstruction, massive hemorrhage,or carcinoid syndrome(20% of the time). Liver metastases are required to produce the carcinoid syndrome wbich is attributed by the serotonin produced by the tumor cells. The classical carcinoid syndrome consists of vasomotor, cardiopulmonary, and GI symptoms. High levels of serotonin and tachykinins probably lead to hyperperistalsis, flushing, bronchial constriction and endocardial fibrosis(Rosai's textbook). Immunohistochemical staining commonly shows chromogranin A, synaptophysin. Many tumors also stain with serotonin(5-HT). The tumor cells are stongly stained for cytokeratin 18(Weidner, Weiss…p718-719) Wang GG et al(MD Anderson) 47 patients with neuroendocrine tumors including 16 with pancreatic endocrine tumors, 15 with nonileal carcinoid tumors and 16 with ileal carcinoid tumors. patients with ileal carcinoid tumors more frequently had liver metastasis compared to patients with nonileal carcinoid tumors and pancreatic endocrine tumors (P=0.02). Allelic loss of chromosome 11q was present in 21% of tumors, chromosome 16q in 13%, and chromosome 18 in 30%. allelic loss of chromosome 18 was present in 69% of ileal carcinoid tumors, 13% of nonileal carcinoid tumors and 6% of pancreatic endocrine tumors (P=0.001). Complete allelic loss of chromosome 18 was associated with smaller tumor size (P=0.02). Our study indicates that genetic alterations vary by tumor subsite and clinicopathologic features, and ileal carcinoid tumors have distinctive clinicopathologic and genetic profiles. Detection of somatostatin receptors(SSTRs) and therapy of metastatic PNETs Since its discovery three decades ago as an inhibitor of GH release from the pituitary gland, somatostatin has attracted much attention because of its functional role in the regulation of a wide variety of physiological functions in the brain, pituitary, pancreas, gastrointestinal tract, adrenals, thyroid, kidney and immune system. Its actions include inhibition of endocrine and exocrine secretions, modulation of neurotransmission, motor and cognitive functions, inhibition of intestinal motility, absorption of nutrients and ions and vascular contractility. In addition, the peptide controls the proliferation of normal and tumor cells. Its action is mediated by a family of G protein-coupled receptors [somatostatin receptor (SSTR)1-SSTR5] that are widely distributed in normal and cancer cells. Direct antitumor activities, mediated through SSTR expressed in tumor cells, include blockade of autocrine/paracrine growth-promoting hormone and growth factor production, inhibition of growth factor-mediated mitogenic signals and induction of apoptosis. Indirect antitumor effects include inhibition of growth-promoting hormone and growth factor secretion, and antiangiogenic actions. Many human tumors express more than one SSTR subtype, with SSTR2 being predominant. These receptors represent the molecular basis for the clinical use of somatostatin analogs in the treatment of endocrine tumors and their in vivo localization . Papotti M.et reported that t he vast majority expressed SSTR types 1,2,3,5 and SSTR4 was detected in a small minority. There was a good correlation between RT-PCR and IHC. By immunohistochemistry on 35 GEP tumors, pancreatic insulinomas showed heterogeneous exhibited SSTR expression, 100% of somatostatinomas:SSTR5 and 100% of gastrinomas and glucagonomas possessed SSTR2. It is concluded that SSTRs 1-5 are heterogeneously expressed in GEP endocrine tumors and that IHC is a reliable tool to detect SSTR types 2, 3 and 5 in surgical and biopsy specimens. Nasir A, et al studied the epxpression of SSTRs in the hepatic metastases of pancreastic and intestinal NETs. Eleven (61%) of 18 hepatic metastases from small intestinal and pancreatic ECAs were positive for SSTR-1, 15 (83%) for SSTR-2, 13 (72%) for SSTR-3, 10 (56%) for SSTR-4, and 15 (83%) for SSTR-5. Among 11 hepatic ECA metastases from small intestinal ECAs (carcinoids), 7 (63%) expressed SSTR-1, 9 (81%) expressed SSTR-2, 8 (72%) expressed SSTR-3, 6 (54%) expressed SSTR-4, and 10 (91%) expressed SSTR-5. Of 7 hepatic ECA metastases from pancreatic ECAs, 4 expressed SSTR-1, 6 expressed SSTR-2, and 5 expressed SSTR-3 and SSTR-5 each. SSTR subtype expression needs to be correlated to somatostatin analog therapy. Immunohistochemical profiling of various SSTR subtypes as a part of routine surgical pathologic analysis of enteropancreatic ETs may become a useful predictor of responsiveness ETs for various SSTR analogues. Zar et al. reported that young age and diagnosis in recent years are positive predictors of survival for patients with midgut caricinoids. Akerstom et al. reported that the midgut caricinoids have typically slow proliferation and extended disease course, and surgical treatment has become increasingly important for their management. For abdominal complication, surgery is combined with long-acting somatostatin analoques and interferon. Molecular Endocrine Pathology In the first case, we have performed DNA mutation analysis of RET gene and found a point mutation at codon 918. In the second case , SSTR2a mRNA was detected by RT-PCR. It should be emphasized that both studies were performed on the formalin fixed paraffin embedded(FFPE) tissue(sections). FFPE materials have used successfully for not only DNA studies bu RNA studies. It is particularly important for pathologists, because the paraffin blocks which have been stored in the pathology departments are the target to perform genetic analysis. Scicchitano 　et al. have reported that their results show that FFPE samples are amenable to Affymetrix GeneChip analysis opening up the possibility for expression profiling on archived tissue blocks in pathology laboratories. Gjerdrum et al. found a marked decrease in the mRNA yield from 500 microdissected cells from frozen and paraffin sections after immunostaining. Recovery of mRNA decreased by up to 89%, comparing the immunostained with the routinely stained sections. Slides mounted with paraffin sections could be stored at room temperature for up to 90 days with no significant decrease in mRNA recovery. Abrahamsen et al. have reported that t he polymerase chain reaction product from FFPE tissues could be increased up to 100-fold amplifying short (<136 bp) compared with long amplicons. Variations in time before tissue processing and in fixation length seem to be less important sources of imprecision than previously assumed. Their findings suggest ed that quantitative analysis of mRNA in archive and routine diagnostic tissues may be possible. References Wang GG, Yao JC, Worah S, White JA, Luna R, Wu TT, Hamilton SR, Rashid A. Comparison of genetic alterations in neuroendocrine tumors: frequent loss of chromosome 18 in ileal carcinoid tumors. Mod Pathol. 2005 Aug;18(8):1079-87. Zar N et al. Long-term survival of patients with small intestinal carcinoid tumors. World J Sug 28:1163-8, 2004. Akerstrom G et al. Management of midgut carcinoids. J Surg Oncol 89:161-9,2005. Papotti M, Bongiovanni M, Volante M, Allia E, Landolfi S, Helboe L, Schindler M, Cole SL, Bussolati G: Expression of somatostatin receptor types 1-5 in 81 cases of gastrointestinal and pancreatic endocrine tumors A correlative immunohistochemical and reverse-transcriptase polymerase chain reaction analysis. . Virchows Arch. 2002 440(5):461-75 Nasir A, Stridsberg M, Strosberg J, Su PH, Livingston S, Malik HA, Kelley ST, Centeno BA, Coppola D, Malafa ME, Yeatman TJ, Kvols LK. Somatostatin receptor profiling in hepatic metastases from small intestineal and pancreatic neuroendocrine neoplasms: immunohistochemical approach with potential clinical utility. Cancer Control. 2006 Jan;13(1):52-60. Abrahamsen HN, Steiniche T, Nexo E, Hamilton-Dutoit SJ, Sorensen BS. Towards quantitative mRNA analysis in paraffin-embedded tissues using real-time reverse transcriptase-polymerase chain reaction: a methodological study on lymph nodes from melanoma patients J Mol Diagn. 2003 Feb;5(1):34-41 Gjerdrum LM, Abrahamsen HN, Villegas B, Sorensen BS, Schmidt H, Hamilton-Dutoit SJ. The influence of immunohistochemistry on mRNA recovery from microdissected frozen and formalin-fixed, paraffin-embedded sections Diagn Mol Pathol. 2004 Dec;13(4):224-33 Scicchitano MS, Dalmas DA, Bertiaux MA, Anderson SM, Turner LR, Thomas RA, Mirable R, Boyce RW. Preliminary Comparison of Quantity, Quality, and Microarray Performance of RNA Extracted from Formalin-fixed Paraffin- embedded and Unfixed Frozen Tissue Samples