RESUMO
BACKGROUND: Current diagnostic and prognostic blood-based biomarkers for neuroendocrine tumors are limited. MiRNAs have tumor-specific expression patterns, are relatively stable, and can be measured in patient blood specimens. We performed a multi-stage study to identify and validate characteristic circulating miRNAs in patients with metastatic small intestine neuroendocrine tumors, and to assess associations between miRNA levels and survival. METHODS: Using a 742-miRNA panel, we identified candidate miRNAs similarly expressed in 19 small intestine neuroendocrine tumors and matched plasma samples. We refined our panel in an independent cohort of plasma samples from 40 patients with metastatic small intestine NET and 40 controls, and then validated this panel in a second, large cohort of 120 patients with metastatic small intestine NET and 120 independent controls. RESULTS: miRNA profiling of 19 matched small intestine neuroendocrine tumors and matched plasma samples revealed 31 candidate miRNAs similarly expressed in both tissue and plasma. We evaluated expression of these 31 candidate miRNAs in 40 independent cases and 40 normal controls, and identified 4 miRNAs (miR-21-5p, miR-22-3p, miR-29b-3p, and miR-150-5p) that were differently expressed in cases and controls (p<0.05). We validated these 4 miRNAs in a separate, larger panel of 120 cases and 120 controls. We confirmed that high circulating levels of miR-22-3p (p<0.0001), high levels of miR 21-5p, and low levels of miR-150-5p (p=0.027) were associated with the presence of metastatic small intestine NET. While levels of 29b-3p were lower in cases than in controls in both the initial cohort and the validation cohort, the difference in the validation cohort did not reach statistical significance. We further found that high levels of circulating miR-21-5p, high levels of circulating miR-22-3p and low levels of circulating miR-150-5p were each independently associated with shorter overall survival. A combined analysis using all three markers was highly prognostic for survival (HR 0.47, 95% CI 0.27-0.82). CONCLUSIONS: Our study suggests that elevated circulating levels of miR-21-5p and miR-22-3p and low levels of miR-150-5p are characteristic in patients with metastatic small intestine neuroendocrine tumors, and further suggests that levels of these miRNAs are associated with overall survival. These observations provide the basis for further validation studies, as well as studies to assess the biological function of these miRNAs in small intestine neuroendocrine tumors.
RESUMO
CONTEXT: Serum IGF-I levels are often low in patients with short stature (SS) without defined etiology. Hence, genetic investigations have focused on the GH-IGF-I axis. OBJECTIVE: Our objectives were to characterize IGF-I axis status and search for a broader range of genetic associations in children with SS and normal GH. DESIGN AND SETTING: We conducted a prospective, cross-sectional, epidemiogenetic case-control study in 9 European countries (2008-2010). PARTICIPANTS: Children (n = 275) aged ≥2 years with SS without defined etiology (≤-2.5 height SD score [SDS]) and ≥1 peak GH ≥7 µg/L) were recruited. METHODS: Serum IGF-I, IGF-binding protein-3 (IGFBP-3), and acid-labile subunit (ALS) levels were measured in a central laboratory. Candidate gene exome sequencing was performed in this cohort and ethnicity-matched controls. RESULTS: Serum IGF-I, IGFBP-3, and ALS levels were highly correlated, but there was a discrepancy between prevalence of IGF-I, IGFBP-3, and ALS deficiencies (53%, 30%, and 0.8%, respectively). An insertion-deletion (Indel) on the IGF1 gene (P = 1.2 × 10(-5), Bonferroni-corrected; case vs control frequency: 0.04 vs 0.112), an Indel on NFKB1 (P = 1.36 × 10(-10); case vs control frequency: 0.464 vs 0.272), and 2 single-nucleotide polymorphisms on ZBTB38 (P < 2.3 × 10(-6)) were associated with SS. At P < 10(-4), single-nucleotide polymorphisms on genes related to protein kinase regulation, MAPK, and Fanconi pathways were also associated with SS. CONCLUSIONS: IGF-I deficiency is a common feature in SS without defined etiology; an Indel in the IGF1 gene was associated with SS. However, genes involved in transcriptional regulation (NFKB1 and ZBTB38) and growth factor signaling were also associated, providing further candidates for genetic investigations on individual patients.