Expression of the Serrated Markers Annexin A10 or Gremlin1 in Colonic Adenocarcinomas: Morphology and Prognostic Values.

Describe clinical, histological and molecular charatcteristics and prognosis values of the serrated candidate markers AnnexinA10 and Gremlin1 in colon adenocarcinomas. Immunohistochemical expression of AnnexinA10 and Gremlin1 was evaluated on 346 colonic adenocarcinomas. Clinicopathological, molecular features and prognostic characteristics were then evaluated. A total of 40 colonic adenocarcinomas expressed AnnexinA10 (11.6%) and, 115 expressed Gremlin1 (40.4%). AnnexinA10 expression was significantly associated, on univariate analyses, with female gender (p = 0.03), right tumor location (p < 0.001), differentiation grade 3 (p < 0.001), serrated adenocarcinoma subtype (p < 0.001), serrated (p < 0.001), medullary (p = 0.005), and mucinous component (p = 0.004), cytoplasmic eosinophilia (p < 0.001), discernible nuclei (p = 0.001), preserved polarity (p < 0.001), lymphatic invasion (p = 0.01), BRAFV600E mutation (p < 0.001), MSI-H status (p < 0.001) and CIMP-H status (p = 0.019). Multivariate analyses revealed that mucinous component (p = 0.002), lymphatic invasion (p = 0.02) and BRAFV600E mutation (p < 0.001) were independently associated with AnnexinA10 expression. In addition, AnnexinA10 was an indicator of poorer overall survival (OS) in UICC stage IV adenocarcinomas (p = 0.01) only. Gremlin1 expression was neither associated with serrated adenocarcinoma subtype (p = 0.51) nor with AnnexinA10 expression (p = 0,31), but was significantly associated, in univariate analysis with male gender (p = 0.002), younger age (p = 0.002), left tumor location (p = 0.04), and MSS status (p = 0.03). Gremlin1 expression was associated with better OS only in UICC stage III colon adenocarcinomas (p = 0.006). Colon adenocarcinomas expressing AnnexinA10 have distinct clinico-pathological and molecular features. AnnexinA10 expression is an indicator of poorer OS in UICC stage IV patients. Gremlin1 expression is not associated with serrated adenocarcinomas subtype. Its expression was associated with better OS in UICC Stage III patients.

Is the Mirror Image Method Really Useful in Tumor Tissue Bank Quality Control?

Oncology research projects are highly dependent on the quality of tumor samples stored in the biobank. Microscopic control is important to ensure the quality of the frozen sample (Does the sample correspond to tumor tissue? Does the sample contain a sufficient number of tumor cells for molecular analysis?). The aim of this study was to evaluate the value of the mirror image method in quality control of colonic adenocarcinoma samples stored in a tumor bank. Microscopic concordance for the differentiation grade, malignant and normal cell percentages, necrosis, mucinous component, and ulceration was assessed on 82 colon adenocarcinoma banked samples and their paired, formalin-fixed, paraffin-embedded mirror controls. Molecular concordance for KRAS status was evaluated in 76 of these 82 cases. Morphological correspondence between frozen and mirror samples was good for the mucinous component (intraclass correlation coefficient [ICC] = 0.81), moderate for differentiation (Cohen's kappa coefficient [k] = 0.67), fair for malignant cells (ICC = 0.44), and poor for ulceration (k = 0.08), normal tissue (ICC = 0.36), and necrosis (ICC = 0.13) percentages. Molecular correspondence for KRAS status was almost perfect (95% correspondence, k = 0.88) between frozen and mirror samples. In conclusion, the mirror sample method is not a good alternative for microscopic and molecular control of frozen colonic adenocarcinoma samples.

LRP1 expression in colon cancer predicts clinical outcome.

LRP1 (low-density lipoprotein receptor-related protein 1), a multifunctional endocytic receptor, has recently been identified as a hub within a biomarker network for multi-cancer clinical outcome prediction. As its role in colon cancer has not yet been characterized, we here investigate the relationship between LRP1 and outcome. MATERIALS AND METHODS: LRP1 mRNA expression was determined in colon adenocarcinoma and paired colon mucosa samples, as well as in stromal and tumor cells obtained after laser capture microdissection. Clinical potential was further investigated by immunohistochemistry in a population-based colon cancer series (n = 307). LRP1 methylation, mutation and miR-205 expression were evaluated and compared with LRP1 expression levels. RESULTS: LRP1 mRNA levels were significantly lower in colon adenocarcinoma cells compared with colon mucosa and stromal cells obtained after laser capture microdissection. Low LRP1 immunohistochemical expression in adenocarcinomas was associated with higher age, right-sided tumor, loss of CDX2 expression, Annexin A10 expression, CIMP-H, MSI-H and BRAFV600E mutation. Low LRP1 expression correlated with poor clinical outcome, especially in stage IV patients. While LRP1 expression was downregulated by LRP1 mutation, LRP1 promoter was never methylated. CONCLUSIONS: Loss of LRP1 expression is associated with worse colon cancer outcomes. Mechanistically, LRP1 mutation modulates LRP1 expression.

Doxorubicin Drug-Eluting Embolic Chemoembolization of Hepatocellular Carcinoma: Study of Midterm Doxorubicin Delivery in Resected Liver Specimens.

This study evaluated the midterm delivery of doxorubicin in liver specimens from patients (N = 4) with hepatocellular carcinoma treated with drug-eluting embolic (DEE) transarterial chemoembolization. The patients had surgical resection 57, 79, 80 and 105 days after doxorubicin DEE chemoembolization. Doxorubicin concentrations inside embolic particles and in surrounding tissues were assessed by infrared microspectroscopy and microspectrofluorimetry, respectively. Embolic particles still contained doxorubicin and provided sustained drug delivery within targeted tissues 80 days after chemoembolization. Doxorubicin was undetectable after 105 days. In addition, aggregation of embolic particles inside vessel lumina was associated with slower doxorubicin elution and higher tissue concentrations when the number of aggregated embolic particles increased.

Development and validation of the Nancy histological index for UC.

OBJECTIVE: We developed a validated index for assessing histological disease activity in UC and established its responsiveness. METHODS: Two hundred biopsies were scored. The outcome was the Global Visual Evaluation (GVE). Eight histological features were tested. The Nancy index was developed by multiple linear regression and bootstrap process to create an index that best matched the GVE. Goodness of fit was assessed by the adjusted R squared (adjusted R2). The second step was the validation of the index: 100 biopsies were scored for the Nancy index by three pathologists from different centres. Inter-reader reliability was evaluated for each reader. The relationship between the change of the Nancy index and the Geboes index was assessed to assess the responsiveness. RESULTS: After backward selection with bootstrap validation, 3/8 items were selected: ulceration (adjusted R2=0.55), acute inflammatory infiltrate (adjusted R2=0.88) and chronic inflammatory infiltrate (adjusted R2=0.79). The Nancy index is defined by a 5-level classification ranging from grade 0 (absence of significant histological disease activity) to grade 4 (severely active disease). The intraclass correlation coefficient (ICC) for the intrareader reliability was 0.88 (95% CI 0.82 to 0.92) and the index had good inter-reader reliability (ICC=0.86 (0.81 to 0.99)). The correlation between the Nancy index and the Geboes score or the GVE was very good. The index had a good responsiveness with a high correlation between changes in the Geboes score and changes in the Nancy index (0.910 (0.813 to 0.955)). CONCLUSIONS: A three descriptor histological index has been validated for use in clinical practice and clinical trials.

Familial small-intestine carcinoids: Chromosomal alterations and germline inositol polyphosphate multikinase sequencing.

BACKGROUND: Familial small-intestine neuroendocrine tumors (SI-NETs) are an exceptional inherited entity. Underlying predisposing mechanisms are unelucidated, but inositol polyphosphate multikinase (IPMK) gene alterations might promote their tumorigenesis. METHODS: A retrospective-prospective nationwide cohort was constituted, by including patients with proven SI-NETs and at least one relative with the same disease. We performed constitutional and somatic IPMK sequencing, and somatic DNA comparative genomic hybridization (CGH). RESULTS: We included 17 patients from 8 families, who were characterized by high prevalence (57%) of multiple SI-NETs, and high frequency of distant metastases (82%) and carcinoid syndrome (65%). No IPMK mutation was found in constitutional or tumor DNA. CGH array revealed recurrent chromosome-18 deletions but no alteration in the IPMK region. CONCLUSION: We report here the first European series of patients with familial SI-NETs. Predisposing mechanisms may not involve the IPMK-encoding sequence or chromosomal region and might not differ from those of sporadic SI-NETs.

Matricellular TSP-1 as a target of interest for impeding melanoma spreading: towards a therapeutic use for TAX2 peptide.

Thrombospondin-1 (TSP-1) is a matricellular glycoprotein known for being highly expressed within a tumor microenvironment, where it promotes an aggressive phenotype particularly by interacting with the CD47 cell-surface receptor. While it originates from the stromal compartment in many malignancies, melanoma is an exception as invasive and metastatic melanoma cells overexpress TSP-1. We recently demonstrated that a new molecular agent that selectively prevents TSP-1 binding to CD47, called TAX2, exhibits anti-cancer properties when administered systemically by decreasing viable tumor tissue within subcutaneous B16 melanoma allografts. At the same time, emerging evidence was published suggesting a contribution of TSP-1 in melanoma metastatic dissemination and resistance to treatment. Through a comprehensive systems biology approach based on multiple genomics and proteomics databases analyses, we first identified a TSP-1-centered interaction network that is overexpressed in metastatic melanoma. Then, we investigated the effects of disrupting TSP-1:CD47 interaction in A375 human malignant melanoma xenografts. In this model, TAX2 systemic administrations induce tumor necrosis by decreasing intra-tumoral blood flow, while concomitantly making tumors less infiltrative. Besides, TAX2 treatment also drastically inhibits B16F10 murine melanoma cells metastatic dissemination and growth in a syngeneic experimental model of lung metastasis, as demonstrated by histopathological analyses as well as longitudinal and quantitative µCT follow-up of metastatic progression. Altogether, the results obtained by combining bioinformatics and preclinical studies strongly suggest that targeting TSP-1/CD47 axis may represent a valuable therapeutic alternative for hampering melanoma spreading.

Portal hypertension with extensive fibrosis and plasma cell infiltration in multiple myeloma.

Plasma cell infiltration of the liver has been described in about 45% of patient with multiple myeloma in autopsy review; however, it is usually not associated with significant liver dysfunction. Indeed, only rare cases of massive plasma cell infiltration leading to non-obstructive cholestasis and hepatic failure have been described. Here, we report a case with a history of 8 years of MM with extensive liver fibrosis and portal hypertension with no other evidence aetiology unless massive plasma cell infiltration who presented a significant regression of both biological liver abnormalities and liver stiffness after ten months of chemotherapy concomitantly to a significant decrease of the IgG serum monoclonal band.

Impact of Visceral Obesity on Microvascular Invasion in Hepatocellular Carcinoma.

BACKGROUND: Our objective was to determine whether visceral obesity is associated with increased microvascular invasion (MVI) in patients surgically treated for hepatocellular carcinoma (HCC). METHODS: Data were collected retrospectively in a series of 79 patients treated by surgical resection for HCC, using CT-scan for evaluation of visceral obesity. RESULTS: There was no significant association between visceral obesity and MVI (OR = 1.20 (0.38-3.75), p = 0.75). Independent predictive factors of MVI were moderate/poor differentiation, tumor size above 50 mm and underlying cirrhosis. CONCLUSION: This study did not support the hypothesis that visceral obesity might promote MVI in patients with HCC.

Biobanking of Fresh-Frozen Human Adenocarcinomatous and Normal Colon Tissues: Which Parameters Influence RNA Quality?

Medical research projects become increasingly dependent on biobanked tissue of high quality because the reliability of gene expression is affected by the quality of extracted RNA. Hence, the present study aimed to determine if clinical, surgical, histological, and molecular parameters influence RNA quality of normal and tumoral frozen colonic tissues. RNA Quality Index (RQI) was evaluated on 241 adenocarcinomas and 115 matched normal frozen colon tissues collected between October 2006 and December 2012. RQI results were compared to patients' age and sex, tumor site, kind of surgery, anastomosis failure, adenocarcinoma type and grade, tumor cell percentage, necrosis extent, HIF-1α and cleaved caspase-3 immunohistochemistry, and BRAF, KRAS and microsatellites status. The RQI was significantly higher in colon cancer tissue than in matched normal tissue. RQI from left-sided colonic cancers was significantly higher than RQI from right-sided cancers. The RNA quality was not affected by ischemia and storage duration. According to histological control, 7.9% of the samples were unsatisfactory because of inadequate sampling. Biobanked tumoral tissues with RQI ≥5 had lower malignant cells to stromal cells ratio than samples with RQI <5 (p <0.05). Cellularity, necrosis extent and mucinous component did not influence RQI results. Cleaved caspase-3 and HIF-1α immunolabelling were not correlated to RQI. BRAF, KRAS and microsatellites molecular status did not influence RNA quality. Multivariate analysis revealed that the tumor location, the surgical approach (laparoscopy versus open colectomy) and the occurrence of anastomotic leakage were the only parameters influencing significantly RQI results of tumor samples. We failed to identify parameter influencing RQI of normal colon samples. These data suggest that RNA quality of colonic adenocarcinoma biospecimens is determined by clinical and surgical parameters. More attention should be paid during the biobanking procedure of right-sided colon cancer or laparoscopic colectomy specimen. Histological quality control remains essential to control sampling accuracy.

Signaling of Serum Amyloid A Through Receptor for Advanced Glycation End Products as a Possible Mechanism for Uremia-Related Atherosclerosis.

OBJECTIVE: Cardiovascular disease is the leading cause of death in patients with end-stage renal disease. Serum amyloid A (SAA) is an acute phase protein and a binding partner for the multiligand receptor for advanced glycation end products (RAGE). We investigated the role of the interaction between SAA and RAGE in uremia-related atherogenesis. APPROACH AND RESULTS: We used a mouse model of uremic vasculopathy, induced by 5 of 6 nephrectomy in the Apoe(-/-) background. Sham-operated mice were used as controls. Primary cultures of Ager(+/+) and Ager(-/-) vascular smooth muscle cells (VSMCs) were stimulated with recombinant SAA, S100B, or vehicle alone. Relevance to human disease was assessed with human VSMCs. The surface area of atherosclerotic lesions at the aortic roots was larger in uremic Apoe(-/-) than in sham-operated Apoe(-/-) mice (P<0.001). Furthermore, atherosclerotic lesions displayed intense immunostaining for RAGE and SAA, with a pattern similar to that of α-SMA. Ager transcript levels in the aorta were 6× higher in uremic animals than in controls (P<0.0001). Serum SAA concentrations were higher in uremic mice, not only after 4 weeks of uremia but also at 8 and 12 weeks of uremia, than in sham-operated animals. We investigated the functional role of RAGE in uremia-induced atherosclerosis further, in animals lacking RAGE. We found that the induction of uremia in Apoe(-/-) Ager(-/-) mice did not accelerate atherosclerosis. In vitro, the stimulation of Ager(+/+) but not of Ager(-/-) VSMCs with SAA or S100B significantly induced the production of reactive oxygen species, the phosphorylation of AKT and mitogen-activated protein kinase-extracellular signal-regulated kinases and cell migration. Reactive oxygen species inhibition with N-acetyl cysteine significantly inhibited both the phosphorylation of AKT and the migration of VSMCs. Similar results were obtained for human VSMCs, except that the phosphorylation of mitogen-activated protein kinase-extracellular signal-regulated kinases, rather than of AKT, was subject to specific redox-regulation by SAA and S100B. Furthermore, human aortic atherosclerotic sections were positively stained for RAGE and SAA. CONCLUSIONS: Uremia upregulates SAA and RAGE expression in the aortic wall and in atherosclerotic lesions in mice. Ager(-/-) animals are protected against the uremia-induced acceleration of atherosclerosis. SAA modulates the functions of murine and human VSMCs in vitro in a RAGE-dependent manner. This study, therefore, identifies SAA as a potential new uremic toxin involved in uremia-related atherosclerosis through interaction with RAGE.

Cytopathology of Vitreous Humor Samples in Routine Practice.

OBJECTIVE: To describe findings in vitreous fluid samples in routine cytology practice. STUDY DESIGN: The pathology archives from 1988 to 2008 at our institution were searched for vitreous samples. The slides were reviewed and clinical and follow-up information was obtained. RESULTS: One hundred and eighty-two vitreous fluid samples from 166 patients were analyzed. Most of the samples had been collected for vitreous hemorrhage (75 cases). The second reason for vitreous sample cytological evaluation was an intraocular inflammatory process (55 cases). A specific cause of inflammation was found by combining clinical, microbiological and cytopathological findings in 19 cases, i.e. infection in 7, sarcoidosis in 3, retinal necrosis in 3, lens-induced endophthalmitis in 2, uveitis associated with systemic disease in 2, retrobulbar neuritis in 1 and sympathetic ophthalmia in 1. Among the 19 samples from 16 patients collected to rule out malignancy, 8 had a confirmed intraocular malignancy. Malignant cells were observed in 5 cases (3 lymphomas, 1 melanoma and 1 carcinoma). Undiagnosed malignancies included 2 lymphomas and 1 choroidal melanoma. Other samples were collected during surgery for retinal detachment and cataracts. CONCLUSIONS: In routine practice, cytology of the vitreous fluid is performed in many and varied situations that are most often nonneoplastic. In nonneoplastic cases, cytology is a useful adjunct to vitrectomy, in spite of its limitations.

BRAFV600E Gene Mutation in Colonic Adenocarcinomas. Immunohistochemical Detection Using Tissue Microarray and Clinicopathologic Characteristics: An 86 Case Series.

The detection of BRAF mutation in colorectal cancer has several clinical applications: enabling the discrimination between sporadic and Lynch syndrome-related colorectal carcinoma, and providing warning of a poorer prognosis. Few immunohistochemical studies using whole-tissue tumor section staining have recently been performed on colorectal cancer. The aim of this study was to evaluate the detection of BRAF mutation by immunohistochemistry (IHC) on tissue microarray (TMA). IHC was performed with the BRAF-specific antibody using TMA on a retrospective series of 86 colonic adenocarcinomas with known BRAF status. IHC using BRAF-specific antibody allowed to detect 20/21 BRAF mutated colonic adenocarcinomas and 60/65 BRAF wild-type cases. The staining was equivocal because of equivocal staining in 4 cases and heterogeneity in 3 cases. When compared with TaqMan real-time PCR, the sensitivity and specificity were 95.2% and 92.3%, respectively. Comparison with the whole section immunostaining improved sensitivity to 100% and specificity to 95.4%. Furthermore, in this study we found that BRAF mutated colonic adenocarcinoma were significantly more frequent in women, older patients, and right-sided. Moreover, morphologic features significantly associated with BRAF mutation were: serrated adenocarcinoma subtype, adenocarcinomas with a mucinous component, high histologic grade, pushing margins, stromal inflammation. BRAF-specific antibody can be used on TMA to screen BRAF-mutated colorectal carcinomas. Cases with equivocal or heterogenous staining must be compared with whole section staining. Moreover, BRAF mutated colonic carcinomas have distinct clinical and histopathologic features.

Ménétrier's disease: Long-term remission with lanreotide.

Ménétrier's disease is a rare hypertrophic gastropathy, causing protein leak. An overexpression of transforming growth factor alpha is involved. In inhibiting the epidermal growth factor receptor, cetuximab and somatostatin analogues are the two most promising treatments, allowing to avoid radical gastrectomy. We report the case of a patient with a sustained clinical remission after treatment with lanreotide, but without complete endoscopic healing. We discuss the available therapeutic options and present a literature review of somatostatin analogues for the treatment of Ménétrier's disease.

Induction and regulation of murine emphysema by elastin peptides.

Emphysema is the major component of chronic obstructive pulmonary disease (COPD). During emphysema, elastin breakdown in the lung tissue originates from the release of large amounts of elastase by inflammatory cells. Elevated levels of elastin-derived peptides (EP) reflect massive pulmonary elastin breakdown in COPD patients. Only the EP containing the GXXPG conformational motif with a type VIII ß-turn are elastin receptor ligands inducing biological activities. In addition, the COOH-terminal glycine residue of the GXXPG motif seems a prerequisite to the biological activity. In this study, we endotracheally instilled C57BL/6J mice with GXXPG EP and/or COOH-terminal glycine deleted-EP whose sequences were designed by molecular dynamics and docking simulations. We investigated their effect on all criteria associated with the progression of murine emphysema. Bronchoalveolar lavages were recovered to analyze cell profiles by flow cytometry and lungs were prepared to allow morphological and histological analysis by immunostaining and confocal microscopy. We observed that exposure of mice to EP elicited hallmark features of emphysema with inflammatory cell accumulation associated with increased matrix metalloproteinases and desmosine expression and of remodeling of parenchymal tissue. We also identified an inactive COOH-terminal glycine deleted-EP that retains its binding-activity to EBP and that is able to inhibit the in vitro and in vivo activities of emphysema-inducing EP. This study demonstrates that EP are key actors in the development of emphysema and that they represent pharmacological targets for an alternative treatment of emphysema based on the identification of EP analogous antagonists by molecular modeling studies.

Investigating the relationship between changes in collagen fiber orientation during skin aging and collagen/water interactions by polarized-FTIR microimaging.

Upon chronological aging, human skin undergoes structural and molecular modifications, especially at the level of type I collagen. This macromolecule is one of the main dermal structural proteins and presents several age-related alterations. It exhibits a triple helical structure and assembles itself to form fibrils and fibers. In addition, water plays an important role in stabilizing the collagen triple helix by forming hydrogen-bonds between collagen residues. However, the influence of water on changes of dermal collagen fiber orientation with age has not been yet understood. Polarized-Fourier Transform Infrared (P-FTIR) imaging is an interesting biophotonic approach to determine in situ the orientation of type I collagen fibers, as we have recently shown by comparing skin samples of different ages. In this work, P-FTIR spectral imaging was performed on skin samples from two age groups (35- and 38-year-old on the one hand, 60- and 66-year-old on the other hand), and our analyses were focused on the effect of H2O/D2O substitution. Spectral data were processed with fuzzy C-means (FCM) clustering in order to distinguish different orientations of collagen fibers. We demonstrated that the orientation was altered with aging, and that D2O treatment, affecting primarily highly bound water molecules, is more marked for the youngest skin samples. Collagen-bound water-related spectral markers were also highlighted. Our results suggest a weakening of water/collagen interactions with age. This non-destructive and label-free methodology allows us to understand better the importance of bound water in collagen fiber orientation alterations occurring with skin aging. Obtaining such structural information could find benefits in dermatology as well as in cosmetics.

Identification of TAX2 peptide as a new unpredicted anti-cancer agent.

The multi-modular glycoprotein thrombospondin-1 (TSP-1) is considered as a key actor within the tumor microenvironment. Besides, TSP-1 binding to CD47 is widely reported to regulate cardiovascular function as it promotes vasoconstriction and angiogenesis limitation. Therefore, many studies focused on targeting TSP-1:CD47 interaction, aiming for up-regulation of physiological angiogenesis to enhance post-ischemia recovery or to facilitate engraftment. Thus, we sought to identify an innovative selective antagonist for TSP-1:CD47 interaction. Protein-protein docking and molecular dynamics simulations were conducted to design a novel CD47-derived peptide, called TAX2. TAX2 binds TSP-1 to prevent TSP-1:CD47 interaction, as revealed by ELISA and co-immunoprecipitation experiments. Unexpectedly, TAX2 inhibits in vitro and ex vivo angiogenesis features in a TSP-1-dependent manner. Consistently, our data highlighted that TAX2 promotes TSP-1 binding to CD36-containing complexes, leading to disruption of VEGFR2 activation and downstream NO signaling. Such unpredicted results prompted us to investigate TAX2 potential in tumor pathology. A multimodal imaging approach was conducted combining histopathological staining, MVD, MRI analysis and µCT monitoring for tumor angiography longitudinal follow-up and 3D quantification. TAX2 in vivo administrations highly disturb syngeneic melanoma tumor vascularization inducing extensive tumor necrosis and strongly inhibit growth rate and vascularization of human pancreatic carcinoma xenografts in nude mice.

Metachronous hormonal syndromes in patients with pancreatic neuroendocrine tumors: a case-series study.

BACKGROUND: Pancreatic neuroendocrine tumors (PNETs) may evolve and cause hormonal hypersecretion-related symptoms that were not present at the initial diagnosis, termed metachronous hormonal syndromes (MHSs). Their setting, characteristics, and outcomes are not well-described. OBJECTIVE: To describe MHSs in patients with sporadic PNETs. DESIGN: Retrospective, multicenter study. SETTING: 4 French referral centers. PATIENTS: Patients with PNETs who developed MHSs related to hypersecretion of insulin, gastrin, vasoactive intestinal peptide, or glucagon between January 2009 and January 2014. MEASUREMENTS: Tumor extension, biological markers, and treatments at initial PNET diagnosis and MHS onset. Pathologic specimens were evaluated centrally, including Ki-67 index and hormone immunolabeling. RESULTS: Of 435 patients with PNETs, 15 (3.4%) were identified as having MHSs involving the hypersecretion of insulin (5 patients), vasoactive intestinal peptide (5 patients), gastrin (2 patients), or glucagon (4 patients). Metachronous hormonal syndromes developed after a median of 55 months (range, 7 to 219) and in the context of PNET progression, stability, and tumor response in 8, 6, and 1 patients, respectively. The median Ki-67 index was 7% (range, 1% to 19%) at PNET diagnosis and 17.5% (range, 2.0% to 70.0%) at MHS onset. Immunolabeling of MHS-related peptides was retrospectively found in 8 of 14 of pathologic PNET specimens obtained before MHS diagnosis. Median survival after MHS onset was 28 months (range, 3 to 56). Seven patients with MHSs died during follow-up, all due to PNETs, including 4 patients with insulin-related MHSs. LIMITATION: Retrospective data collection and heterogeneity of pathologic specimen size and origin. CONCLUSION: Metachronous hormonal syndromes were identified more often in the context of PNET progression and increased Ki-67 indices. Patients with insulin-related MHSs may have decreased survival rates. PRIMARY FUNDING SOURCE: None.