Sunitinib for the treatment of patients with advanced pheochromocytomas or paragangliomas: The phase 2 non-randomized SUTNET clinical trial.

BACKGROUND: Metastatic Pheochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumors characterized by high morbidity and limited systemic treatment options, mainly based on radiometabolic treatments or chemotherapy. Based on the preclinical rationale that PGGLs carcinogenesis relies on angiogenesis, treatment with tyrosine kinase inhibitors (TKI) may represent another viable therapeutic option. METHODS: We conducted a prospective phase II study in patients with metastatic or unresectable PGGLs. Patients received sunitinib (50 mg daily for 4 weeks, followed by a 2-week rest period) until progressive disease (PD), unacceptable toxicity or consent withdrawal. The primary endpoint was 12-month progression-free survival (PFS) rate; secondary endpoints were safety overall response rate (ORR) according to RECIST 1.1 criteria and overall survival (OS). EudraCT Number: 2011-002632-99. RESULTS: Fifty patients were included. At a median follow-up of 71.7 months (IQR 35.4-100.1), the 1 year-PFS rate was 53.4 % (95 %CI 41.1-69.3) and median PFS was 14.1 months (95 % CI 8.9-25.7). ORR was 15.6 %, the median OS was 49.4 months (95 %CI 21.2-NA), and grade 3 or higher treatment-related adverse events were reported in 34 % patients. No significant correlation was found between specific genetic alterations or genomic clusters and sunitinib efficacy. CONCLUSION: Sunitinib is an active drug in patients with advanced PGGLs, capable of inducing prolonged disease control with a manageable toxicity profile.

Colorectal carcinoma peritoneal metastases-derived organoids: results and perspective of a model for tailoring hyperthermic intraperitoneal chemotherapy from bench-to-bedside.

BACKGROUND: Peritoneal metastases from colorectal cancer (CRCPM) are related to poor prognosis. Cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC) have been reported to improve survival, but peritoneal recurrence rates are still high and there is no consensus on the drug of choice for HIPEC. The aim of this study was to use patient derived organoids (PDO) to build a relevant CRCPM model to improve HIPEC efficacy in a comprehensive bench-to-bedside strategy. METHODS: Oxaliplatin (L-OHP), cisplatin (CDDP), mitomycin-c (MMC) and doxorubicin (DOX) were used to mimic HIPEC on twelve PDO lines derived from twelve CRCPM patients, using clinically relevant concentrations. After chemotherapeutic interventions, cell viability was assessed with a luminescent assay, and the obtained dose-response curves were used to determine the half-maximal inhibitory concentrations. Also, induction of apoptosis by different HIPEC interventions on PDOs was studied by evaluating CASPASE3 cleavage. RESULTS: Response to drug treatments varied considerably among PDOs. The two schemes with better response at clinically relevant concentrations included MMC alone or combined with CDDP. L-OHP showed relative efficacy only when administered at low concentrations over a long perfusion period. PDOs showed that the short course/high dose L-OHP scheme did not appear to be an effective choice for HIPEC in CRCPM. HIPEC administered under hyperthermia conditions enhanced the effect of chemotherapy drugs against cancer cells, affecting PDO viability and apoptosis. Finally, PDO co-cultured with cancer-associated fibroblast impacted HIPEC treatments by increasing PDO viability and reducing CASPASES activity. CONCLUSIONS: Our study suggests that PDOs could be a reliable in vitro model to evaluate HIPEC schemes at individual-patient level and to develop more effective treatment strategies for CRCPM.

Comprehensive genomic and transcriptomic characterization of high-grade gastro-entero-pancreatic neoplasms.

BACKGROUND: High-grade gastro-entero-pancreatic neoplasms (HG GEP-NENs) can be stratified according to their morphology and Ki-67 values into three prognostic classes: neuroendocrine tumors grade 3 (NETs G3), neuroendocrine carcinomas with Ki-67 < 55% (NECs <55) and NECs with Ki-67 ≥ 55% (NECs ≥55). METHODS: We analyzed a cohort of 49 HG GEP-NENs by targeted Next-Generation Sequencing (TrueSight Oncology 500), RNA-seq, and immunohistochemistry for p53, Rb1, SSTR-2A, and PD-L1. RESULTS: Frequent genomic alterations affected TP53 (26%), APC (20%), KRAS and MEN1 (both 11%) genes. NET G3 were enriched in MEN1 (p = 0.02) mutations, while both NECs groups were enriched in TP53 (p = 0.001), APC (p = 0.002) and KRAS (p = 0.02) mutations and tumors with TMB ≥ 10 muts/Mb (p = 0.01). No differentially expressed (DE) gene was found between NECs <55% and NECs ≥55%, while 1129 DE genes were identified between NET G3 and NECs. A slight enrichment of CD4+ and CD8+ T cells in NECs and of cancer-associated fibroblasts and macrophages (M2-like) in NET G3. Multivariate analysis identified histologic type and Rb1 loss as independent prognostic factors for overall survival. CONCLUSIONS: This study showed that GEP-NET G3 and GEP-NECs exhibit clear genomic and transcriptomic differences, differently from GEP-NECs <55% and GEP-NECs ≥55%, and provided molecular findings with prognostic and potentially predictive value.

Integrated mutational landscape analysis of poorly differentiated high-grade neuroendocrine carcinoma of the uterine cervix.

High-grade neuroendocrine cervical cancers (NETc) are exceedingly rare, highly aggressive tumors. We analyzed 64 NETc tumor samples by whole-exome sequencing (WES). Human papillomavirus DNA was detected in 65.6% (42/64) of the tumors. Recurrent mutations were identified in PIK3CA, KMT2D/MLL2, K-RAS, ARID1A, NOTCH2, and RPL10. The top mutated genes included RB1, ARID1A, PTEN, KMT2D/MLL2, and WDFY3, a gene not yet implicated in NETc. Somatic CNV analysis identified two copy number gains (3q27.1 and 19q13.12) and five copy number losses (1p36.21/5q31.3/6p22.2/9q21.11/11p15.5). Also, gene fusions affecting the ACLY-CRHR1 and PVT1-MYC genes were identified in one of the eight samples subjected to RNA sequencing. To resolve evolutionary history, multiregion WES in NETc admixed with adenocarcinoma cells was performed (i.e., mixed-NETc). Phylogenetic analysis of mixed-NETc demonstrated that adenocarcinoma and neuroendocrine elements derive from a common precursor with mutations typical of adenocarcinomas. Over one-third (22/64) of NETc demonstrated a mutator phenotype of C > T at CpG consistent with deficiencies in MBD4, a member of the base excision repair (BER) pathway. Mutations in the PI3K/AMPK pathways were identified in 49/64 samples. We used two patient-derived-xenografts (PDX) (i.e., NET19 and NET21) to evaluate the activity of pan-HER (afatinib), PIK3CA (copanlisib), and ATR (elimusertib) inhibitors, alone and in combination. PDXs harboring alterations in the ERBB2/PI3K/AKT/mTOR/ATR pathway were sensitive to afatinib, copanlisib, and elimusertib (P < 0.001 vs. controls). However, combinations of copanlisib/afatinib and copanlisib/elimusertib were significantly more effective in controlling NETc tumor growth. These findings define the genetic landscape of NETc and suggest that a large subset of these highly lethal malignancies might benefit from existing targeted therapies.

Hybrid epithelial-mesenchymal status of lung cancer dictates metastatic success through differential interaction with NK cells.

BACKGROUND: Epithelial to mesenchymal transition (EMT) endows cancer cells with pro-metastatic properties, which appear most effective when cells enter an intermediate hybrid (H) state, characterized by integrated mesenchymal (M) and epithelial (E) traits. The reasons for this advantage are poorly known and, especially, it is totally unexplored whether the interplay between H-cells and NK cells could have a role. Here we characterize the pro-metastatic mechanics of non-small cell lung cancer (NSCLC) H-cells and their subset of cancer-initiating cells (CICs), dissecting crucial interactions with NK cells. METHODS: Human lung cancer cell lines and sublines representative of E, M, or H states, assessed by proteomics, were analyzed in vivo for their tumor-forming and disseminating capabilities. Interactions with NK cells were investigated in vitro using migration assays, cytotoxic degranulation assays, and evaluation of CD133+ CICs modulation after coculture, and validated in vivo through NK cell neutralization assays. Correlation between EMT status, NK cell infiltration, and survival data, was evaluated in a cohort of surgically resected NSCLC cases (n=79). RESULTS: We demonstrated that H-cells, have limited dissemination capability but show the highest potential to initiate metastases in vivo. This property was related to their ability to escape NK cell surveillance. Mechanistically, H-cells expressed low levels of NK-attracting chemokines (CXCL1 and CXCL8), generating poorly infiltrated metastases. Accordingly, proteomics and GO enrichment analysis of E, H, M cell lines showed that the related secretory processes could change during EMT.Furthermore, H-CICs uniquely expressed high levels of the inhibitory ligand B7-H3, which protected H-CIC from NK cell-mediated clearance. In vivo neutralization assays confirmed that, indeed, the pro-metastatic properties of H-cells are poorly controlled by NK cells.Finally, the analysis of patients revealed that detection of hybrid phenotypes associated with low NK infiltration in NSCLC clinical specimens could identify a subset of patients with poor prognosis. CONCLUSIONS: Our study demonstrates that H-cells play a central role in the metastatic spread in NSCLC. Such pro-metastatic advantage of H-cells is supported by their altered interaction with NK cells and by the critical role of B7-H3 in preserving their H-CIC component, indicating B7-H3 as a potential target in combined NK-based therapies.

Antibody dependent cellular cytotoxicity-inducing anti-EGFR antibodies as effective therapeutic option for cutaneous melanoma resistant to BRAF inhibitors.

Introduction: About 50% of cutaneous melanoma (CM) patients present activating BRAF mutations that can be effectively targeted by BRAF inhibitors (BRAFi). However, 20% of CM patients exhibit intrinsic drug resistance to BRAFi, while most of the others develop adaptive resistance over time. The mechanisms involved in BRAFi resistance are disparate and globally seem to rewire the cellular signaling profile by up-regulating different receptor tyrosine kinases (RTKs), such as the epidermal growth factor receptor (EGFR). RTKs inhibitors have not clearly demonstrated anti-tumor activity in BRAFi resistant models. To overcome this issue, we wondered whether the shared up-regulated RTK phenotype associated with BRAFi resistance could be exploited by using immune weapons as the antibody-dependent cell cytotoxicity (ADCC)-mediated effect of anti-RTKs antibodies, and kill tumor cells independently from the mechanistic roots. Methods and results: By using an in vitro model of BRAFi resistance, we detected increased membrane expression of EGFR, both at mRNA and protein level in 4 out of 9 BRAFi-resistant (VR) CM cultures as compared to their parental sensitive cells. Increased EGFR phosphorylation and AKT activation were observed in the VR CM cultures. EGFR signaling appeared dispensable for maintaining resistance, since small molecule-, antibody- and CRISPR-targeting of EGFR did not restore sensitivity of VR cells to BRAFi. Importantly, immune-targeting of EGFR by the anti-EGFR antibody cetuximab efficiently and specifically killed EGFR-expressing VR CM cells, both in vitro and in humanized mouse models in vivo, triggering ADCC by healthy donors' and patients' peripheral blood cells. Conclusion: Our data demonstrate the efficacy of immune targeting of RTKs expressed by CM relapsing on BRAFi, providing the proof-of-concept supporting the assessment of anti-RTK antibodies in combination therapies in this setting. This strategy might be expected to concomitantly trigger the crosstalk of adaptive immune response leading to a complementing T cell immune rejection of tumors.

An Italian real-world multicenter study of patients with advanced mixed neuroendocrine non-neuroendocrine neoplasms (MiNENs) of the gastro-entero-pancreatic system treated with chemotherapy.

PURPOSE: The aim of this study is to describe the clinical management of an Italian series of patients with advanced gastro-entero-pancreatic (GEP) MiNENs treated in clinical practice. METHODS: Clinical records of patients from four Italian referral Centers were retrospectively analyzed to correlate clinical/biological data with clinical outcomes. All the surgical specimens were centrally reviewed. RESULTS: Clinical data and surgical samples of 51 patients during 1995-2015 were analyzed. Sites of origin were: 32 colorectal, 14 gastro-esophageal, and 5 pancreatobiliary. Twenty-one out of fifty-one (42.2%) developed metachronous distant metastases. Only 5/51 (9.8%) patients received peri-operative therapy, and 23/51 (45.1%) first-line chemotherapy, mostly fluoropyrimidines/oxaliplatin. The NEN component was poorly differentiated in the whole population. Patients with Ki67 index < 55% in the NEC component had a significantly longer median overall survival (OS) (35.3 months; 95% CI 27.1-41.0) than those with Ki67 ≥ 55% (11.9 months; 95% CI 9.1-14.0) P = 0.0005. The median OS was 14 months (95% CI 10.1-19.1) in the whole cohort, with 11.4 months (95% CI 6.2-20.2) in patients who received a first-line therapy. CONCLUSION: This study confirms that GEP-MiNENs represent a complex disease and that over the past years the clinical management has been predominantly guided by the subjective judgment of the clinicians. Although, in this series, the NEC component appeared mostly responsible for the systemic spread and prognosis on the whole neoplasm, the lack of strong prognostic and predictive factors universally recognized seems to condition their management so far. Future prospective clinical and biomolecular studies could help clinicians to improve clinical management of GEP-MiNENs.

Modulation of faecal miRNAs highlights the preventive effects of a Mediterranean low-inflammatory dietary intervention.

BACKGROUND: Dietary interventions have been proposed as therapeutic approaches for several diseases, including cancer. A low-inflammatory Mediterranean dietary intervention, conducted as a pilot study in subjects with Familial Adenomatous Polyposis (FAP), reduced markers of local and systemic inflammation. We aim to determine whether this diet may modulate faecal microRNA (miRNA) and gene expression in the gut. METHODS: Changes in the faecal miRNome were evaluated by small RNA sequencing at baseline (T0), after the three-month intervention (T1), and after an additional three months (T2). Changes in the transcriptome of healthy rectal mucosa and adenomas were evaluated by RNA sequencing at T0 and T2. The identification of validated miRNA-gene interactions and functional analysis of miRNA targets were performed using in silico approaches. RESULTS: Twenty-seven subjects were included in this study. It was observed that the diet modulated 29 faecal miRNAs (p < 0.01; |log2 Fold Change|>1), and this modulation persisted for three months after the intervention. Levels of miR-3612-3p and miR-941 correlated with the adherence to the diet, miR-3670 and miR-4252-5p with faecal calprotectin, and miR-3670 and miR-6867 with serum calprotectin. Seventy genes were differentially expressed between adenoma and normal tissue, and most were different before the dietary intervention but reached similar levels after the diet. Functional enrichment analysis identified the proinflammatory ERK1/2, cell cycle regulation, and nutrient response pathways as commonly regulated by the modulated miRNAs and genes. CONCLUSIONS: Faecal miRNAs modulated by the dietary intervention target genes that participate in inflammation. Changes in levels of miRNAs and genes with oncogenic and tumour suppressor functions further support the potential cancer-preventive effect of the low-inflammatory Mediterranean diet. CLINICAL TRIAL NUMBER REGISTRATION: NCT04552405, Registered in ClinicalTrials.gov.

Characterization of two transcriptomic subtypes of marker-null large cell carcinoma of the lung suggests different origin and potential new therapeutic perspectives.

Pulmonary large cell carcinoma (LCC) is an undifferentiated neoplasm lacking morphological, histochemical, and immunohistochemical features of small cell lung cancer, adenocarcinoma (ADC), or squamous cell carcinoma (SCC). The available molecular information on this rare disease is limited. This study aimed to provide an integrated molecular overview of 16 cases evaluating the mutational asset of 409 genes and the transcriptomic profiles of 20,815 genes. Our data showed that TP53 was the most frequently inactivated gene (15/16; 93.7%) followed by RB1 (5/16; 31.3%) and KEAP1 (4/16; 25%), while CRKL and MYB genes were each amplified in 4/16 (25%) cases and MYC in 3/16 (18.8%) cases; transcriptomic analysis identified two molecular subtypes including a Pure-LCC and an adenocarcinoma like-LCC (ADLike-LCC) characterized by different activated pathways and cell of origin. In the Pure-LCC group, POU2F3 and FOXI1 were distinctive overexpressed markers. A tuft cell-like profile and the enrichment of a replication stress signature, particularly involving ATR, was related to this profile. Differently, the ADLike-LCC were characterized by an alveolar-cell transcriptomic profile and association with AIM2 inflammasome complex signature. In conclusion, our study split the histological marker-null LCC into two different transcriptomic entities, with POU2F3, FOXI1, and AIM2 genes as differential expression markers that might be probed by immunohistochemistry for the differential diagnosis between Pure-LCC and ADLike-LCC. Finally, the identification of several signatures linked to replication stress in Pure-LCC and inflammasome complex in ADLike-LCC could be useful for designing new potential therapeutic approaches for these subtypes.

Endoscopic management of gastric, duodenal and rectal NETs: Position paper from the Italian Association for Neuroendocrine Tumors (Itanet), Italian Society of Gastroenterology (SIGE), Italian Society of Digestive Endoscopy (SIED).

The present paper reflects the position of the Italian Association for Neuroendocrine Tumors (Itanet), the Italian Society of Gastroenterology (SIGE), and the Italian Society of Digestive Endoscopy (SIED) regarding the management of patients affected by gastric, duodenal, and rectal neuroendocrine neoplasms (NENs) amenable to endoscopic treatment. The key questions discussed in this paper are summarized in Table 1. Data were extracted from the MEDLINE database through searches; expert opinions and recommendations are provided in accordance with the available scientific evidence and the authors' expertise. Recommendations are presented alongside a level of evidence and grade of recommendation based on the GRADE system. This paper specifically focuses on subgroups of NENs considered suitable for endoscopic management according to current international guidelines: i. well-differentiated gastric neuroendocrine tumors (gNET) type 1 < 2 cm and selected cases of type 3; ii. well-differentiated duodenal, non-functioning, non-ampullary NET with size < 2 cm; and well-differentiated rectal NET with size < 2 cm.

Clinical impact of mixed pulmonary carcinoma and carcinoid: the driver from their mono-clonal origin.

The combination of neuroendocrine/non neuroendocrine lung tumors (CNNELT) mentioned in the last edition of the World Health Organization (WHO) of Thoracic Tumors refers to small cell carcinoma (SCLC) or large cell neuroendocrine carcinoma (LCNEC) mixed with any other non-small cell lung carcinoma (NSCLC). Typical Carcinoid (TC)/Atypical Carcinoid (AC) combined with NSCLC is not included among this category. However, case reports of TC/AC combined with NSCLC have been described. We previously reported 2 cases of lung adenocarcinoma (LUA) mixed with carcinoid sharing mutations in both components supporting the hypothesis of a clonal origin. We extended our analysis to other four cases of mixed NSCLC-carcinoid by performing targeted-DNA and RNA-based NGS analysis in both primary and their paired lymph nodes metastasis. In all cases, LUA and AC components shared at least 1 common mutation (KRAS driver mutation p.Gly12Val in cases 1 and 3, AKAP13-RET fusion in case 2, and missense KRAS driver mutation p.Gly12Ala in case 4, reinforcing the hypothesis of a clonal origin. Moreover, the same mutation was detected in the metastasis constituted only by AC (cases 2 and 4). Although it is a rare malignancy in the lung, mixed LUA and TC/AC could be included among the histotypes for which a deep molecular characterization of both components is needed to identify the presence of potential druggable genetic alterations.

Optimising Radioligand Therapy for Patients with Gastro-Entero-Pancreatic Neuroendocrine Tumours: Expert Opinion from an Italian Multidisciplinary Group.

Radioligand therapy (RLT) with lutetium (177Lu) oxodotreotide is an approved therapy in combination with somatostatin analogues (SSAs) for patients with advanced, well-differentiated G1-G2, gastro-entero-pancreatic neuroendocrine tumours (GEP-NETs) that progress on SSAs. We conducted a series of round table meetings throughout Italy to identify issues related to RLT delivery to patients with GEP-NETs. Four key issues were identified: (1) the proper definition of tumour progression prior to RLT initiation; (2) the impact of RLT in patients with bone metastases and/or high hepatic tumour burden; (3) the optimal follow-up protocol after RLT; and (4) organisational issues related to RLT use and managerial implications. This article reviews the literature relating to the aforementioned issues and makes recommendations based on available evidence and Italian NET experts' opinions. In particular, the group recommends the development of a diagnostic-therapeutic care pathway (DTCP) for patients undergoing RLT which provides systematic guidance but can still be individualised for each patient's clinical and psychosocial needs. A DTCP may clarify the diagnostic, therapeutic and post-treatment monitoring process, and improve communication and the coordination of care between hub and spoke centres. The DTCP may also contribute to changes in the care process related to the 2013/59/EURATOM Directive and to the definition of costs when planning for future or updated reimbursement of RLT in Italy.

Safety and antitumor activity of metformin plus lanreotide in patients with advanced gastro-intestinal or lung neuroendocrine tumors: the phase Ib trial MetNET2.

In retrospective studies, metformin use has been associated with better clinical outcomes in diabetic patients with advanced, well-differentiated neuroendocrine tumors (WDNETs). However, prospective evidence of metformin safety and activity is lacking. Here, we conducted the first-in-human phase Ib MetNET2 trial to investigate the safety and antitumor activity of metformin in combination with the somatostatin analog lanreotide autogel (ATG) in both diabetic and non-diabetic patients with advanced WDNETs of the gastrointestinal (GI) or thoracic tract. Enrolled patients received lanreotide ATG 120 mg plus oral metformin, up to a maximum dosage of 2550 mg/day. We enrolled 20 patients, of whom 18 (90%) and 2 (10%) had WDNETs of the GI and thoracic tract, respectively. Fourteen patients (70%) were non-diabetic. With a 5% incidence of SAEs, the study met its primary objective of demonstrating treatment safety. With a median follow-up of 39 months (95% CI 28-NE), median PFS was 24 months (95% CI 16-NE), with 12-month and 24-month PFS probability of 75% (95% CI 58-97) and 49% (95% CI 31-77), respectively. We found no statistically significant PFS differences between diabetic and non-diabetic patients. Among exploratory analyses, the presence of tumor genomic alterations in DNA damage pathways was associated with trend towards worse PFS, whereas a precocious reduction of HOMA-IR index and plasma cholesterol concentration showed a trend towards an association with better PFS. In conclusion, metformin plus lanreotide ATG is a safe and well tolerated combination treatment that is associated with promising antitumor activity in both non-diabetic and diabetic patients with WDNETs, and that warrants further investigation in larger clinical trials.

Multidisciplinary Approach to the Diagnosis of Occult Primary Neuroendocrine Neoplasm: A Clinical Challenge.

Approximately 11% to 14% of subjects with neuroendocrine neoplasms (NENs) have metastatic lesions with unknown primary origin (UPO), with the majority of UPO-NENs found in the small bowel. Herein, we assessed the available literature on UPO-NENs, focusing on clinical presentation and diagnostic techniques to identify the primary site. The identification of the primary tumor is important as it affects the prognosis; however, the clinical presentation can be non-specific in non-functioning forms. In the presence of metastatic disease, the histological sample is fundamental to obtain immunohistochemical markers that might orientate the clinician in the search for the primary tumor through radiology, functional imaging and endoscopic techniques. In summary, multidisciplinary management plays a key role in UPO-NENs, even more than in other NENs. Molecular biology and gene-expression profiling represent areas of great interest which might be developed in the near future for both the diagnosis and the treatment of these neoplasms.

Mytomicin-C, Metronomic Capecitabine, and Bevacizumab in Patients With Unresectable or Relapsed Pseudomyxoma Peritonei of Appendiceal Origin.

INTRODUCTION: Pseudomyxoma peritonei (PMP) is a rare, slow growing tumor, traditionally considered chemoresistant. The only curative approach is cytoreductive surgery (CRS) followed by hyperthermic intraperitoneal chemotherapy (HIPEC). At disease relapse, or in patients with inoperable disease at diagnosis, no standard treatment has been defined, though nonrandomized series showed promising results with fluoropyrimidine-based regimens. PATIENTS AND METHODS: We conducted a prospective study in patients with relapsed or unresectable PMP and confirmed disease progression at baseline. Patients received MMC (7 mg/m2 every 6 weeks, up to a maximum of 4 cycles) plus metronomic capecitabine (625 mg/sqm/day b.i.d.) and bevacizumab (7.5 mg/kg every 3 weeks) until disease progression, unacceptable toxicity, or consent withdrawal. Primary endpoint was progression-free survival (PFS); secondary endpoints were overall survival (OS), overall response rate according to RECIST v1.1 criteria, serum markers response and safety. RESULTS: Fifteen patients were included. At a median follow-up of 26.1 months (IQR, 17.7-49.6), median PFS was 17.9 months (95% CI, 11.0-NE), with 1-year PFS and OS rates of 73% and 87%. Safety profile was manageable, with only 13% G3/G4 treatment-related adverse events. CONCLUSION: Metronomic capecitabine, bevacizumab, and MMC are an active regimen in advanced and progressive PMP and favorably compares with historical series.

Assessing the safety and activity of cabozantinib combined with lanreotide in gastroenteropancreatic and thoracic neuroendocrine tumors: rationale and protocol of the phase II LOLA trial.

BACKGROUND: Well-differentiated (WD) neuroendocrine tumors (NETs) are a group of rare neoplasms with limited therapeutic options. Cabozantinib is an inhibitor of multiple tyrosine kinases with a pivotal role in NET pathogenesis, including c-MET and Vascular Endothelial Growth Factor Receptor 2 (VEGFR2). LOLA is the first prospective phase II trial aiming to assess the safety and activity of cabozantinib combined with lanreotide in WD NETs of gastroenteropancreatic (GEP), thoracic and of unknown origin. METHODS: This is a multicenter, open-label, double-cohort, non comparative, non-randomized, three-stage phase II trial. Eligible patients have to meet the following inclusion criteria: diagnosis of advanced or metastatic, progressive, non-functioning WD thoracic NETs, GEP-NETs or NETs of unknown origin with Ki67 ≥ 10%; positive 68 Ga-PET uptake or somatostatin receptor 2 immunohistochemical (IHC) stain; maximum 1 prior systemic regimen for metastatic disease. Two cohorts will be considered: pNETs and carcinoids (typical or atypical lung and thymus NETs, gastro-intestinal NETs or NETs of unknown origin). In stage I, the primary objective is to find the optimal dose of cabozantinib in combination with lanreotide and to evaluate the safety of the combination (percentage of patients experiencing grade 3-5 toxicities according to NCI-CTCAE version 5.0). Starting dose of cabozantinib is 60 mg/day continuously, plus lanreotide 120 mg every 28 days. In stage II and III, co-primary endpoints are safety and overall response rate (ORR) according to RECIST version 1.1. The uninteresting antitumor activity is fixed in ORR ≤ 5%. Secondary endpoints are progression-free survival and overall survival. Exploratory objectives include the assessment of c-MET, AXL and VEGFR2 IHC expression, to identify predictive or prognostic tissue biomarkers. Enrolment started in July 2020, with an expected trial duration of 42 months comprehensive of accrual, treatment and follow-up. Considering a drop-out rate of 5%, the maximum number of enrolled patients will be 69. DISCUSSION: Supported by a solid rationale, the trial has the potential to generate milestone data about the synergistic effects of cabozantinib plus lanreotide in a group of NET patients with relatively aggressive disease and limited therapeutic options. TRIAL REGISTRATION: LOLA is registered at ClinicalTrials.gov (NCT04427787) and EudraCT (2019-004506-10).

Transcriptome analysis of primary sporadic neuroendocrine tumours of the intestine identified three different molecular subgroups.

BACKGROUND: Intestinal neuroendocrine tumours (I-NETs) represent a non-negligible entity among intestinal neoplasms, with metastatic spreading usually present at the time of diagnosis. In this context, effective molecular actionable targets are still lacking. Through transcriptome analysis, we aim at refining the molecular taxonomy of I-NETs, also providing insights towards the identification of new therapeutic vulnerabilities. MATERIALS AND METHODS: A retrospective series of 38 primary sporadic, surgically-resected I-NETs were assessed for transcriptome profiling of 20,815 genes. RESULTS: Transcriptome analysis detected 643 highly expressed genes. Unsupervised hierarchical clustering, differential expression analysis and gene set enriched analysis identified three different tumour clusters (CL): CL-A, CL-B, CL-C. CL-A showed the overexpression of ARGFX, BIRC8, NANOS2, and SSTR4 genes. Its most characterizing signatures were those related to cell-junctions, and activation of mTOR and WNT pathway. CL-A was also enriched in T CD8 + lymphocytes. CL-B showed the overexpression of PCSK1, QPCT, ST18, and TPH1 genes. Its most characterizing signatures were those related to adipogenesis, neuroendocrine metabolism, and splice site machinery-related processes. CL-B was also enriched in T CD4 + lymphocytes. CL-C showed the overexpression of ALB, ANG, ARG1, and HP genes. Its most characterizing signatures were complement/coagulation and xenobiotic metabolism. CL-C was also enriched in M1/2 macrophages. These CL-based differences may have therapeutic implications in refining the management of I-NET patients. At last, we described a specific gene-set for differentiating I-NET from pancreatic NET. DISCUSSION: Our data represent an additional step for refining the molecular taxonomy of I-NET, identifying novel transcriptome subgroups with different biology and therapeutic opportunities.

A2AR Expression and Immunosuppressive Environment Independent of KRAS and GNAS Mutations in Pseudomyxoma Peritonei.

In pseudomyxoma peritonei (PMP), KRAS and GNAS mutations are frequent. We hypothesized that these mutations may contribute to the suppression of antitumor immunity: KRAS may induce GMCSF expression, while GNAS may enhance the expression of cyclic adenosine monophosphate and A2AR signaling. This study aimed to explore possible mechanisms facilitated by KRAS and GNAS mutations for escaping immune surveillance. Additionally, we looked for new potential therapeutic and prognostic targets in this rare disease which is poorly characterized at the molecular level. GM-CSF, A2AR, CD73, CD39, and PD-L1 expression was investigated by immunohistochemistry in 40 PMPs characterized for GNAS and KRAS mutational status. Immune cell populations were studied by immunohistochemistry and nanostring nCounter®. Following the criteria of a prognostic nomogram reported for PMP, we stratified the patients into two different risk groups, with 28 "low-risk" and 12 "high-risk" patients. We observed the expression of GM-CSF (74%); CD39 (37%); CD73 (53%); A2AR (74%); and PD-L1 (16%) which was unrelated to GNAS or KRAS status. The tumor microenvironment showed the presence of CD4+ T cells (86%); CD8+ T cells (27%); CD20+ B (67%); CD15+ cells (86%); and CD163+ M2 macrophages (67%), while CD56+ NK cells were absent. CD163 expression (27%) in PMP tumor cells was associated with poor prognosis. GNAS mutation and A2AR expression were not associated with a specific immune transcriptional signature. However, the expression assay revealed 21 genes associated with prognosis. The "high-risk" patients exhibited worse progression-free survival (HR = 2.3, CI 95%: 1.1-5.1, p = 0.034) and significant downregulation of MET, IL8, PPARG, DTX4, HMGA1, ZIC2, WNT5B, and CCRL2. In conclusion, we documented the presence of immunosuppressive factors such as GM-CSF, A2AR, and PD-L1 in PMP. These factors were not associated with GNAS and KRAS status and could be explored as therapeutic molecular targets. Additionally, a set of potential prognostic biomarkers, including CD163 expression in tumor cells, deserve further investigation.