Proteomic analysis of small intestinal neuroendocrine tumors and mesenteric fibrosis.

Mesenteric metastases in small intestinal neuroendocrine tumors (SI-NETs) are associated with mesenteric fibrosis (MF) in a proportion of patients. MF can induce severe abdominal complications, and an effective preventive treatment is lacking. To elucidate possible novel therapeutic targets, we performed a proteomics-based analysis of MF. The tumor cell and stromal compartment of primary tumors and paired mesenteric metastases of SI-NET patients with MF (n = 6) and without MF (n = 6) was analyzed by liquid chromatography-mass spectrometry-based proteomics. Analysis of differential protein abundance was performed. Collagen alpha-1(XII) (COL12A1) and complement component C9 (C9) expression was evaluated by immunohistochemistry (IHC) in mesenteric metastases. A total of 2988 proteins were identified. Unsupervised hierarchical clustering showed close clustering of paired primary and mesenteric tumor cell samples. Comparing MF to non-MF samples, we detected differentially protein abundance solely in the mesenteric metastasis stroma group. There was no differential abundance of proteins in tumor cell samples or primary tumor stroma samples. Analysis of the differentially abundant proteins (n = 36) revealed higher abundance in MF samples of C9, various collagens and proteoglycans associated with profibrotic extracellular matrix dysregulation and signaling pathways. Proteins involved in fatty acid oxidation showed a lower abundance. COL12A1 and C9 were confirmed by IHC to have significantly higher expression in MF mesenteric metastases compared to non-MF. In conclusion, proteome profiles of SI-NETs with and without MF differ primarily in the stromal compartment of mesenteric metastases. Analysis of differentially abundant proteins revealed possible new signaling pathways involved in MF development. In conclusion, proteome profiles of SI-NETs with and without MF differ primarily in the stromal compartment of mesenteric metastases. Analysis of differentially abundant proteins revealed possible new signaling pathways involved in MF development.

Aberrant tryptophan metabolism in stromal cells is associated with mesenteric fibrosis in small intestinal neuroendocrine tumors.

Background: Increased levels of serotonin secretion are associated with mesenteric fibrosis (MF) in small intestinal neuroendocrine tumors (SI-NETs). However, the profibrotic potential of serotonin differs between patients, and in this study, we aimed to gain an understanding of the mechanisms underlying this variability. To this end, we analyzed the proteins involved in tryptophan metabolism in SI-NETs. Methods: Proteomes of tumor and stroma from primary SI-NETs and paired mesenteric metastases of patients with MF (n = 6) and without MF (n = 6) were identified by liquid chromatography-mass spectrometry (LC-MS). The differential expression of proteins involved in tryptophan metabolism between patients with and without MF was analyzed. Concurrently, monoamine oxidase A (MAO-A) expression was analyzed in the tumor and stromal compartment by immunohistochemistry (IHC) and reported as intensity over area (I/A). Results: Of the 42 proteins involved in tryptophan metabolism, 20 were detected by LC-MS. Lower abundance of ten proteins was found in mesenteric metastases stroma in patients with MF. No differential expression was found in primary SI-NETs. In patients with MF, IHC showed lower MAO-A expression in the stroma of the primary SI-NETs (median 4.2 I/A vs 6.5 I/A in patients without MF, P = 0.003) and mesenteric metastases (median 2.1 I/A vs 2.8 I/A in patients without MF, P= 0.019). Conclusion: We found a decreased expression of tryptophan and serotonin-metabolizing enzymes in the stroma in patients with MF, most notably in the mesenteric stroma. This might account for the increased profibrotic potential of serotonin and explain the variability in the development of SI-NET-associated fibrotic complications.

Sexual Dimorphism in Small-intestinal Neuroendocrine Tumors: Lower Prevalence of Mesenteric Disease in Premenopausal Women.

CONTEXT: Small-intestinal neuroendocrine tumors (SI-NETs) have a modest but significantly higher prevalence and worse prognosis in male patients. OBJECTIVE: This work aims to increase understanding of this sexual dimorphism in SI-NETs. PATIENTS AND METHODS: Retrospectively, SI-NET patients treated in a single tertiary center were included and analyzed for disease characteristics. Estrogen receptor 1 (ESR1) and 2 (ESR2), progesterone receptor (PGR), and androgen receptor (AR) messenger RNA (mRNA) expression was assessed in primary tumors and healthy intestine. Estrogen receptor alpha (ERα) and AR protein expression were analyzed by immunohistochemistry in primary tumors and mesenteric metastases. RESULTS: Of the 559 patients, 47% were female. Mesenteric metastasis/fibrosis was more prevalent in men (71% / 46%) than women (58% / 37%; P = 0.001 and P = 0.027, respectively). In women, prevalence of mesenteric metastases increased gradually with age from 41.1% in women <50 years to 71.7% in women >70 years. Increased expression of ESR1 and AR mRNA was observed in primary tumors compared to healthy intestine (both P < 0.001). ERα staining was observed in tumor cells and stroma with a strong correlation between tumor cells of primary tumors and mesenteric metastases (rho = 0.831, P = 0.02), but not in stroma (rho = -0.037, P = 0.91). AR expression was only found in stroma. CONCLUSION: Sexual dimorphism in SI-NETs was most pronounced in mesenteric disease, and the risk of mesenteric metastasis in women increased around menopause. The combination of increased ERα and AR expression in the SI-NET microenvironment suggests a modulating role of sex steroids in the development of the characteristic SI-NET mesenteric metastasis and associated fibrosis.

Peptide Receptor Radionuclide Therapy With 177Lu-DOTATATE for Symptomatic Control of Refractory Carcinoid Syndrome.

CONTEXT: Peptide receptor radionuclide therapy (PRRT) with [Lutetium-177-DOTA0-Tyr3]octreotate (177Lu-DOTATATE) results in an increase of progression-free survival and quality of life in patients with progressive, well-differentiated neuroendocrine neoplasms (NENs). OBJECTIVE: To study the effect of 177Lu-DOTATATE in patients with carcinoid syndrome and radiologically stable or newly diagnosed disease treated solely for the purpose of symptom reduction. DESIGN: Retrospective cohort study. SETTING: Tertiary care hospital. PATIENTS: Twenty-two patients with a metastatic midgut NEN, elevated urinary 5-hydroxyindolacetic acid excretion, and flushing and/or diarrhea despite treatment with a somatostatin analog, without documented disease progression. INTERVENTION: PRRT with 177Lu-DOTATATE (intended cumulative dose: 29.6 GBq) with a primary aim to reduce symptoms. RESULTS: After PRRT, mean bowel movement frequency (BMF) decreased from 6.1 ± 3.4 to 4.6 ± 3.6 per day (P = 0.009). Flushes decreased from 4.3 ± 2.9 to 2.4 ± 2.7 flushes per day (P = 0.002). A decrease of BMF of more than 30% occurred in 47% of patients with baseline BMF of 4 or more (n = 17). In patients with ≥2 episodes of flushing a day (n = 15), 67% of patients had more than 50% decrease of daily flushing. A decrease in urinary 5-hydroxyindolacetic acid excretion of more than 30% was seen in 56% of patients. The European Organization for Research and Treatment of Cancer-Core Module diarrhea subscale score showed a trend toward improvement by an average of 16.7 ± 33.3 points (P = 0.11). CONCLUSION: PRRT with 177Lu-DOTATATE effectively reduced diarrhea and flushing in patients with carcinoid syndrome and can be considered for symptomatic treatment of carcinoid syndrome insufficiently controlled with somatostatin analogs.

Predicting symptomatic mesenteric mass in small intestinal neuroendocrine tumors using radiomics.

Metastatic mesenteric masses of small intestinal neuroendocrine tumors (SI-NETs) are known to often cause intestinal complications. The aim of this study was to identify patients at risk to develop these complications based on routinely acquired CT scans using a standardized set of clinical criteria and radiomics. Retrospectively, CT scans of SI-NET patients with a mesenteric mass were included and systematically evaluated by five clinicians. For the radiomics approach, 1128 features were extracted from segmentations of the mesenteric mass and mesentery, after which radiomics models were created using a combination of machine learning approaches. The performances were compared to a multidisciplinary tumor board (MTB). The dataset included 68 patients (32 asymptomatic, 36 symptomatic). The clinicians had AUCs between 0.62 and 0.85 and showed poor agreement. The best radiomics model had a mean AUC of 0.77. The MTB had a sensitivity of 0.64 and specificity of 0.68. We conclude that systematic clinical evaluation of SI-NETs to predict intestinal complications had a similar performance than an expert MTB, but poor inter-observer agreement. Radiomics showed a similar performance and is objective, and thus is a promising tool to correctly identify these patients. However, further validation is needed before the transition to clinical practice.

Evolution of the Mesenteric Mass in Small Intestinal Neuroendocrine Tumours.

BACKGROUND: A metastatic mesenteric mass is a hallmark of small intestinal neuroendocrine tumours (SI-NETs). However, little is known on its development over time. Therefore, we conducted a study to assess the evolution of a SI-NET-associated mesenteric mass over time. METHODS: Retrospectively, 530 patients with proven SI-NET were included. The presence and growth of a mesenteric mass was assessed using RECIST 1.1 criteria on every consecutive CT-scan until the end of follow-up or resection. RESULTS: At baseline, a mesenteric mass was present in 64% of the patients, of whom 13.5% showed growth of the mesenteric mass with a median time to growth of 40 months. Male gender was the only independent predictor of growth (OR 2.67). Of the patients without a mesenteric mass at the first evaluation, 2.6% developed a pathological mesenteric mass. Treatment with peptide receptor radionuclide therapy (PRRT; N = 132) resulted in an objective size reduction of the mesenteric mass in 3.8%. CONCLUSION: The metastatic mesenteric mass in SI-NETs has a static behavior over time. Therefore, site-specific growth behavior should be taken into account when selecting target lesions and assessing disease progression and therapeutic response. PRRT appears not to be effective for size reduction of the mesenteric mass.

Critical appraisal of MGMT in digestive NET treated with alkylating agents.

The efficacy of alkylating agents (temozolomide, dacarbazine, streptozotocin) in patients with advanced neuroendocrine tumors (NETs) has been well documented, especially in pancreatic NETs. Alkylating agents transfer methyl adducts on DNA bases. Among them, O6-methylguanine accounts for many of their cytotoxic effects and can be repaired by the O6-methylguanine-methyltransferase (MGMT). However, whether the tumor MGMT status could be a reliable biomarker of efficacy of alkylating agents in NETs is still a matter of debate. Herein, we sought to provide a critical appraisal of the role of the MGMT status in NETs. After reviewing the molecular mechanisms of repair of DNA damage induced by alkylating agents, we aimed to comprehensively review the methods of determination of the MGMT status and its impact on prognosis, prediction of objective response and progression-free survival in patients with advanced digestive NETs treated by alkylating agents. About half of pancreatic NETs are MGMT-deficient, as determined by impaired tumor MGMT expression or by MGMT promoter methylation. Overall, while published studies are heterogeneous and mostly limited in size, they advocate that MGMT deficiency may be a relevant biomarker for increased objective response rate, prolonged progression-fee survival and overall survival in patients with advanced NETs treated by alkylating agents. While these data require confirmation in prospective controlled studies, future research should focus on the standardization of MGMT status assessment. Additional mechanisms of repair of DNA damages induced by alkylating agents should be explored in order to identify biomarkers complementary to MGMT and targets for potential antitumor synergy, such as PARP.

Symptomatic and Radiological Response to 177Lu-DOTATATE for the Treatment of Functioning Pancreatic Neuroendocrine Tumors.

PURPOSE: Peptide receptor radionuclide therapy (PRRT) with the radiolabeled somatostatin analogue [Lutetium-177-DOTA0-Tyr3]octreotate (177Lu-DOTATATE) is widely applied for inoperable metastatic small intestinal and nonfunctioning pancreatic neuroendocrine tumors (pNETs). The aim of this study is to describe the safety and efficacy of the treatment of functioning pNETs. METHODS: Patients were treated with up to four cycles of 177Lu-DOTATATE with an intended dose of 7.4 Gbq per cycle. Radiological (Response Evaluation Criteria in Solid Tumors 1.1), symptomatic, and biochemical response were analyzed retrospectively for all patients with a functioning pNET (insulinoma, gastrinoma, VIPoma, and glucagonoma) treated with 177Lu-DOTATATE. Quality of life (QOL) was assessed with the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire-Core Module questionnaire. RESULTS: Thirty-four patients with a metastatic functioning pNET (European Neuroendocrine Tumor Society grade 1 or 2) were included: 14 insulinomas, 5 VIPomas, 7 gastrinomas, and 8 glucagonomas. Subacute hematological toxicity, grade 3 or 4 occurred in 4 patients (12%) and a hormonal crisis in 3 patients (9%). PRRT resulted in partial or complete response in 59% of patients and the disease control rate was 78% in patients with baseline progression. 71% of patients with uncontrolled symptoms had a reduction of symptoms and a more than 80% decrease of circulating hormone levels was measured during follow-up. After PRRT, median progression-free survival was 18.1 months (interquartile range: 3.3 to 35.7) with a concurrent increase in QOL. CONCLUSION: Treatment with 177Lu-DOTATATE is a safe and effective therapy resulting in radiological, symptomatic and biochemical response in a high percentage of patients with metastatic functioning pNETs. Hormonal crises occur relatively frequent and preventive therapy should be considered before and/or during PRRT.

Small intestinal neuroendocrine tumours and fibrosis: an entangled conundrum.

Small intestinal neuroendocrine tumours (SI-NETs) are neoplasms characterized by their ability to secrete biogenic amines and peptides. These cause distinct clinical pathology including carcinoid syndrome, marked by diarrhoea and flushing, as well as fibrosis, notably mesenteric fibrosis. Mesenteric fibrosis often results in significant morbidity by causing intestinal obstruction, oedema and ischaemia. Although advancements have been made to alleviate symptoms of carcinoid syndrome and prolong the survival of patients with SI-NETs, therapeutic options for patients with mesenteric fibrosis are still limited. As improved insight in the complex pathogenesis of mesenteric fibrosis is key to the development of new therapies, we evaluated the literature for known and putative mediators of fibrosis in SI-NETs. In this review, we discuss the tumour microenvironment, growth factors and signalling pathways involved in the complex process of fibrosis development and tumour progression in SI-NETs, in order to elucidate potential new avenues for scientific research and therapies to improve the management of patients suffering from the complications of mesenteric fibrosis.

Mesenteric fibrosis and palliative surgery in small intestinal neuroendocrine tumours.

Mesenteric fibrosis (MF) surrounding a mesenteric mass is a hallmark feature of small intestinal neuroendocrine tumours (SI-NETs). Since this can induce intestinal obstruction, oedema and ischaemia, prophylactic resection of the primary tumour and mesenteric mass is often recommended. This study assessed the predictors for mesenteric metastasis and fibrosis and the effect of MF and palliative surgery on survival. A retrospective analysis of 559 patients with pathologically proven SI-NET and available CT-imaging data was performed. Clinical characteristics, presence of mesenteric mass and fibrosis on CT imaging and the effect of palliative abdominal surgery on overall survival were assessed. We found that MF was present in 41.4%. Older age, 5-HIAA excretion ≥67 µmol/24 h, serum CgA ≥121.5 µg/L and a mesenteric mass ≥27.5 mm were independent predictors of MF. In patients ≤52 years, mesenteric mass was less often found in women than in men (39% vs 64%, P = 0.002). Corrected for age, tumour grade, CgA and liver metastasis, MF was not a prognostic factor for overall survival. In patients undergoing palliative surgery, metastasectomy of mesenteric mass or prophylactic surgery did not result in survival benefit. In conclusion, we confirmed known predictors of MF and mesenteric mass and suggest a role for sex hormones as women ≤52 years have less often a mesenteric mass. Furthermore, the presence of MF has no effect on survival in a multivariate analysis, and we found no benefit of metastasectomy of mesenteric mass or prophylactic surgery on overall survival.