title: Principal component analysis, hierarchical clustering, and decision tree assessment of plasma mRNA and hormone levels as an early detection strategy for small intestinal neuroendocrine (carcinoid) tumors.
abstract: Incidence of neuroendocrine tumors (NETs) is increasing (approximately 6%/year), but clinical presentation is nonspecific, resulting in delays in diagnosis (5-7 years; approximately 70% have metastases). This reflects absence of a sensitive plasma marker. The aim of this study is to investigate whether detection of circulating messenger RNA (mRNA) alone or in combination with circulating NET-related hormones and growth factors can detect gastrointestinal NET disease. The small intestinal (SI) NET cell line KRJ-I was used to define the sensitivity of real-time polymerase chain reaction (PCR) for mRNA detection in blood. NSE, Tph-1, and VMAT ( 2 ) transcripts were identified from one KRJ-I cell/ml blood. mRNA from the tissue and plasma of SI-NETs (n = 12) and gastric NETs (n = 7), and plasma from healthy controls (n = 9) was isolated and real-time PCR performed. Tph-1 was a specific marker of SI-NETs (58%, p < 0.03) whereas CgA transcripts did not differentiate tumors from controls. Patients with metastatic disease expressed more marker transcripts than localized tumors (75% versus 18%, p < 0.02). Plasma 5-hydroxytryptamine (5-HT), chromogranin A (CgA), ghrelin, and connective tissue growth factor (CTGF) fragments were measured, combined with mRNA levels, and a predictive mathematical model for NET diagnosis developed using decision trees. The sensitivity and specificity to diagnose SI-NETs and gastric NETs were 81.2% and 100%, and 71.4% and 55.6%, respectively. We conclude that mRNA from one NET cell/ml blood can be detected. Circulating plasma Tph-1 is a promising marker gene for SI-NET disease (specificity 100%) while an increased number of marker transcripts (>2) correlated with disease spread. Including NET-related circulating hormones and growth factors in the algorithm increased the sensitivity of detection of SI-NETs from 58 to 82%.
authors.affiliation: Department of Gastroenterological Surgery, Yale University School of Medicine, New Haven, CT 06520-8062, USA. imodlin@optonline.net
authors.forename: Irvin M
authors.initials: IM
authors.lastname: Modlin
authors.forename: Björn I
authors.initials: BI
authors.lastname: Gustafsson
authors.forename: Ignat
authors.initials: I
authors.lastname: Drozdov
authors.forename: Boaz
authors.initials: B
authors.lastname: Nadler
authors.forename: Roswitha
authors.initials: R
authors.lastname: Pfragner
authors.forename: Mark
authors.initials: M
authors.lastname: Kidd
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000097:blood* / Q000235:genetics; D002276:Carcinoid Tumor / Q000097:blood*; D016022:Case-Control Studies; D053379:Chromogranin A / Q000097:blood*; D016000:Cluster Analysis; D055513:Connective Tissue Growth Factor / Q000097:blood*; D003663:Decision Trees; D042241:Early Diagnosis; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D007414:Intestinal Neoplasms / Q000097:blood*; D007421:Intestine, Small; D008297:Male; D008875:Middle Aged; D009367:Neoplasm Staging; D025341:Principal Component Analysis; D011379:Prognosis; D012333:RNA, Messenger / Q000097:blood* / Q000235:genetics; D012334:RNA, Neoplasm / Q000097:blood / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
chemical_list: D014408:Biomarkers, Tumor; D053379:Chromogranin A; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D055513:Connective Tissue Growth Factor