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Background Chemotherapy-induced neutropenia (CIN) has been demonstrated to be a prognostic factor in several cancer conditions. We previously found a significant prognostic value of CIN on overall survival (OS), in a pooled dataset of patients with advanced non-small-cell lung cancer (NSCLC) receiving first line chemotherapy from 1996 to 2001. However, the prognostic role of CIN in NSCLC is still debated. Methods We performed a post hoc analysis pooling data prospectively collected in six randomized phase 3 trials in NSCLC conducted from 2002 to 2016. Patients who never started chemotherapy and those for whom toxicity data were missing were excluded. Neutropenia was categorized on the basis of worst grade during chemotherapy: absent (grade 0), mild (grade 1–2), or severe (grade 3–4). The primary endpoint was OS. Multivariable Cox model was applied for statistical analyses. In the primary analysis, a minimum time (landmark) at 180 days from randomization was applied in order to minimize the time-dependent bias. Results Overall, 1529 patients, who received chemotherapy, were eligible; 572 of them (who received 6 cycles of treatment) represented the landmark population. Severe CIN was reported in 143 (25.0%) patients and mild CIN in 135 (23.6%). At multivariable OS analysis, CIN was significantly predictive of prognosis although its prognostic value was entirely driven by severe CIN (hazard ratio [HR] of death 0.71; 95%CI: 0.53–0.95) while it was not evident with mild CIN (HR 1.21; 95%CI: 0.92–1.58). Consistent results were observed in the out-of-landmark group (including 957 patients), where both severe and mild CIN were significantly associated with a reduced risk of death. Conclusion The pooled analysis of six large trials of NSCLC treatment shows that CIN occurrence is significantly associated with a longer overall survival, particularly in patients developing severe CIN, confirming our previous findings.

Gut microbiota is involved in immune modulation and immune checkpoint inhibitors (ICIs) efficacy. Single-arm phase II CAVE-mCRC and CAVE-LUNG clinical trials investigated cetuximab + avelumab combination in RAS wild-type (WT) metastatic colorectal cancer (mCRC) and chemo-refractory nonsmall cell lung cancer (NSCLC) patients, respectively. A comprehensive gut microbiota genetic analysis was done in basal fecal samples of 14 patients from CAVE-mCRC trial with circulating tumor DNA (ctDNA) RAS/BRAF WT and microsatellite stable (MSS) disease. Results were validated in a cohort of 10 patients from CAVE-Lung trial. 16S rRNA sequencing revealed 23 027 bacteria species in basal fecal samples of 14 patients from CAVE-mCRC trial. In five long-term responding patients (progression-free survival [PFS], 9-24 months) significant increases in two butyrate-producing bacteria, Agathobacter M104/1 (P=.018) and Blautia SR1/5 (P=.023) were found compared to nine patients with shorter PFS (2-6months). A significantly better PFS was also observed according to the presence or absence of these species in basal fecal samples. For Agathobacter M104/1, median PFS (mPFS) was 13.5months (95% confidence interval [CI], 6.5-20.5months) vs 4.6months (95% CI, 1.8-7.4months); P=.006. For Blautia SR1/5, mPFS was 5.9months (95% CI, 2.2-9.7months) vs 3.6months (95% CI, 3.3-4.0months); P=.021. Similarly, in CAVE-Lung validation cohort, Agathobacter M104/1 and Blautia SR1/5 expression were associated with PFS according to their presence or absence in basal fecal samples. Agathobacter and Blautia species could be potential biomarkers of outcome in mCRC, and NSCLC patients treated with cetuximab + avelumab. These findings deserve further investigation.

Introduction: COVID-19 has disrupted the global health care system since March 2020. Lung cancer (LC) patients (pts) represent a vulnerable population highly affected by the pandemic. This multicenter Italian study aimed to evaluate whether the COVID-19 outbreak had an impact on access to cancer diagnosis and treatment of LC pts compared with pre-pandemic time. Methods: Consecutive newly diagnosed LC pts referred to 25 Italian Oncology Departments between March and December 2020 were included. Access rate and temporal intervals between date of symptoms onset and diagnostic and therapeutic services were compared with the same period in 2019. Differences between the 2 years were analyzed using the chi-square test for categorical variables and the Mann–Whitney U test for continuous variables. Results: A slight reduction (−6.9%) in newly diagnosed LC cases was observed in 2020 compared with 2019 (1523 versus 1637, P = 0.09). Newly diagnosed LC pts in 2020 were more likely to be diagnosed with stage IV disease (P < 0.01) and to be current smokers (someone who has smoked more than 100 cigarettes, including hand-rolled cigarettes, cigars, cigarillos, in their lifetime and has smoked in the last 28 days) (P < 0.01). The drop in terms of new diagnoses was greater in the lockdown period (percentage drop −12% versus −3.2%) compared with the other months included. More LC pts were referred to a low/medium volume hospital in 2020 compared with 2019 (P = 0.01). No differences emerged in terms of interval between symptoms onset and radiological diagnosis (P = 0.94), symptoms onset and cytohistological diagnosis (P = 0.92), symptoms onset and treatment start (P = 0.40), and treatment start and first radiological revaluation (P = 0.36). Conclusions: Our study pointed out a reduction of new diagnoses with a shift towards higher stage at diagnosis for LC pts in 2020. Despite this, the measures adopted by Italian Oncology Departments ensured the maintenance of the diagnostic-therapeutic pathways of LC pts.

CoronaVirus disease-2019 has changed the delivery of health care worldwide and the pandemic has challenged oncologists to reorganize cancer care. Recently, progress has been made in the field of precision medicine to provide to patients with cancer the best therapeutic choice for their individual needs. In this context, the Foundation Medicine (FMI)-Liquid@Home project has emerged as a key weapon to deal with the new pandemic situation. FoundationOne Liquid Assay (F1L) is a next-generation sequences-based liquid biopsy service, able to detect 324 molecular alterations and genomic signatures, from May 2020 available at patients’ home (FMI-Liquid@Home). We analyzed time and costs saving for patients with cancer, their caregivers and National Healthcare System (NHS) with FMI-Liquid@Home versus F1L performed at our Department. Different variables have been evaluated. Between May 2020 and August 2021, 218 FMI-Liquid@Home were performed for patients with cancer in Italy. Among these, our Department performed 153 FMI-Liquid@Home with the success rate of 98% (vs. 95% for F1L in the hospital). Time saving for patients and their caregivers was 494.86 and 427.36 hours, respectively, and costs saving was 13 548.70€. Moreover, for working people these savings were 1084.71 hours and 31 239.65€, respectively. In addition, the total gain for the hospital was 163.5 hours and 6785€, whereas for NHS was 1084.71 hours and 51 573.60€, respectively. FMI-Liquid@Home service appears to be useful and convenient allowing time and costs saving for patients, caregivers, and NHS. Born during the COVID-19 pandemic, it could be integrated in oncological daily routine in the future. Therefore, additional studies are needed to better understand the overall gain and how to integrate this service in different countries.

Simple Summary Multimodality treatment can improve outcome of malignant pleural mesothelioma (MPM) patients. However, the ideal scheme of combination of them is still unknown. We analyzed a case series of 17 patients treated at our institution with a sequence of induction chemotherapy, surgery, adjuvant radiotherapy and chemotherapy. Median overall survival was 32.1 months, median progression free survival was 23.7 months with a safe profile. These data according to our experience represent a great example of feasibility in clinical practice of three-modality four step approach and encourage further prospective studies to better define the details of treatment. Abstract Background: Multimodality treatment is considered the best treatment strategy for malignant pleural mesothelioma (MPM). However, the ideal combination of them is still a matter of controversy. Here, we report a case series of MPM treated with a trimodality approach: induction chemotherapy (CT), pleurectomy/decortication (P/D), postoperative radiotherapy (RT) and post-operative CT. Methods: A retrospective case series of 17 MPM patients treated between 2013 and 2020 is presented. Patients had epithelial or mixed MPM diagnosed by video-assisted thoracoscopy and pathologic IMIG stage I or II disease. Treatment details and radiological data were collected. Induction therapy consisted of combination of cisplatin and pemetrexed, every 21 days for two cycles. P/D was performed 4–6 weeks after induction CT, post-operative RT 3–6 weeks after surgery, while post-operative CT was given 4–6 weeks after RT, with the same schedule of induction. Results: All patients showed objective response or stability of disease at the restaging following induction CT and underwent surgery by posterolateral thoracotomy. There were two cases of cardiac arrest as major intraoperative complication, both resolved by manual cardiac massage. Minor complications included one hemidiaphragm elevation, 1 anemia requiring blood transfusion, one wound infection, and two persistent air leaks. Median overall survival was 32.1 months, median progression free survival was 23.7 months. Conclusions: These results suggest the feasibility of these trimodality treatment scheme for early stage MPM patients. Larger series and long-term prospective studies are needed to confirm the validity of this strategy.

Non-small cell lung cancer (NSCLC), the leading cause of cancer death worldwide, is still an unmet medical problem due to the lack of both effective therapies against advanced stages and markers to allow a diagnosis of the disease at early stages before its progression. Immunotherapy targeting the PD-1/PD-L1 checkpoint is promising for many cancers, including NSCLC, but its success depends on the tumor expression of PD-L1. PATZ1 is an emerging cancer-related transcriptional regulator and diagnostic/prognostic biomarker in different malignant tumors, but its role in lung cancer is still obscure. Here we investigated expression and role of PATZ1 in NSCLC, in correlation with NSCLC subtypes and PD-L1 expression. A cohort of 104 NSCLCs, including lung squamous cell carcinomas (LUSCs) and adenocarcinomas (LUADs), was retrospectively analyzed by immunohistochemistry for the expression of PATZ1 and PD-L1. The results were correlated with each other and with the clinical characteristics, showing on the one hand a positive correlation between the high expression of PATZ1 and the LUSC subtype and, on the other hand, a negative correlation between PATZ1 and PD-L1, validated at the mRNA level in independent NSCLC datasets. Consistently, two NSCLC cell lines transfected with a PATZ1-overexpressing plasmid showed PD-L1 downregulation, suggesting a role for PATZ1 in the negative regulation of PD-L1. We also showed that PATZ1 overexpression inhibits NSCLC cell proliferation, migration, and invasion, and that Patz1-knockout mice develop LUAD. Overall, this suggests that PATZ1 may act as a tumor suppressor in NSCLC.

BackgroundThe coronavirus disease 2019 (COVID-19) pandemic has overwhelmed the health systems worldwide. Data regarding the impact of COVID-19 on cancer patients (CPs) undergoing or candidate for immune checkpoint inhibitors (ICIs) are lacking. We depicted the practice and adaptations in the management of patients with solid tumors eligible or receiving ICIs during the COVID-19 pandemic, with a special focus on Campania region.MethodsThis survey (25 questions), promoted by the young section of SCITO (Società Campana di ImmunoTerapia Oncologica) Group, was circulated among Italian young oncologists practicing in regions variously affected by the pandemic: high (group 1), medium (group 2) and low (group 3) prevalence of SARS-CoV-2–positive patients. For Campania region, the physician responders were split into those working in cancer centers (CC), university hospitals (UH) and general hospitals (GH). Percentages of agreement, among High (H) versus Medium (M) and versus Low (L) group for Italy and among CC, UH and GH for Campania region, were compared by using Fisher’s exact tests for dichotomous answers and χ2 test for trends relative to the questions with 3 or more options.ResultsThis is the first Italian study to investigate the COVID-19 impact on cancer immunotherapy, unique in its type and very clear in the results. The COVID-19 pandemic seemed not to affect the standard practice in the prescription and delivery of ICIs in Italy. Telemedicine was widely used. There was high consensus to interrupt immunotherapy in SARS-CoV-2–positive patients and to adopt ICIs with longer schedule interval. The majority of the responders tended not to delay the start of ICIs; there were no changes in supportive treatments, but some of the physicians opted for delaying surgeries (if part of patients’ planned treatment approach). The results from responders in Campania did not differ significantly from the national ones.ConclusionOur study highlights the efforts of Italian oncologists to maintain high standards of care for CPs treated with ICIs, regardless the regional prevalence of COVID-19, suggesting the adoption of similar solutions. Research on patients treated with ICIs and experiencing COVID-19 will clarify the safety profile to continue the treatments, thus informing on the most appropriate clinical conducts.

Background RAS mutations are the only validated biomarkers in metastatic colorectal cancer (mCRC) for anti-epidermal growth factor receptor (EGFR) therapy. Limited clinical information is available on AXL expression, marker of epithelial to mesenchymal transition, in mCRC. Methods AXL was retrospectively assessed by immunohistochemistry in 307 patients. RAS wild-type (WT) patients (N = 136) received first-line anti-EGFR–based therapy; RAS mutant patients (N = 171) received anti-angiogenic–based regimens. Preclinical experiments were performed using human RAS WT CRC cell lines and xenograft models. AXL RNA levels were assessed in a cohort of patients with available samples at baseline and at progression to anti-EGFR treatment and in the GSE5851 dataset. Results AXL was expressed in 55/307 tumour tissues, correlating with worse survival in the overall population (AXL-positive, 23.7 months; AXL-negative, 30.8 months; HR, 1.455, P = 0.032) and in RAS WT patients (AXL-positive, 23.0 months; AXL-negative, 35.8 months; HR,1.780, P = 0.032). Progression-free survival (PFS) in the RAS WT cohort was shorter in the AXL-positive cohort (6.2 months versus 12.1 months; HR, 1.796, P = 0.013). Three-dimensional cultures obtained from a patient following anti-EGFR therapy resulted AXL-positive, showing resistance to anti-EGFR drugs and sensitivity to AXL inhibition. AXL transfection in CRC cell lines induced AXL overexpression and resistance to the EGFR blockade. At progression to cetuximab, 2/10 SW48-tumour xenograft mice showed AXL expression. Consistently, AXL RNA levels increased in 5/7 patients following anti-EGFR therapy. Moreover, in the GSE5851 dataset higher AXL RNA levels correlated with worse PFS with cetuximab in KRAS-exon2 WT chemorefractory patients. Conclusions AXL is a marker of poor prognosis in mCRC with consistent clinical and preclinical evidences of involvement in primary and acquired resistance to anti-EGFR drugs in RAS WT patients.

Introduction: Cancer immunotherapies with monoclonal antibodies (mAbs) against immune checkpoints (i.e. CTLA-4 and PD-1/PD-L1) have revolutionized antineoplastic treatments. Immune checkpoint inhibitors (ICIs) approved for cancer immunotherapy are mAbs anti-CTLA-4 (ipilimumab), anti-PD-1 (nivolumab, pembrolizumab, and cemiplimab), and anti-PD-L1 (atezolizumab, avelumab, and durvalumab). Treatment with ICIs can be associated with immune-related adverse events (irAEs), including an increased risk of developing myocarditis. These findings are compatible with the observation that, CTLA-4, PD-1, and PD-L1 pathways play a central role in the modulation of autoimmunity.Areas covered: In this paper, we start from examining the pathogenesis of cardiovascular adverse events from ICIs, and then we focus on risk factors and strategies to prevent and manage this cardiotoxicity.Expert opinion: There is a growing need for a multidisciplinary approach of ICI-associated cardiotoxicity, involving oncologists, cardiologists, and immunologists. Prevention and effective management of ICIs cardiotoxicity starts with an in-depth screening and surveillance strategies of high-risk patients, in order to improve early detection and appropriate management in a personalized approach

Background Despite the advancements in new therapies for colorectal cancer (CRC), chemotherapy still constitutes the mainstay of the medical treatment. For this reason, new strategies to increase the efficacy of chemotherapy are desirable. Poly-ADP-Ribose Polymerase inhibitors (PARPi) have shown to increase the activity of DNA damaging chemotherapeutics used in the treatment of CRC, however previous clinical trials failed to validate these results and pointed out dose-limiting toxicities that hamper the use of such combinations in unselected CRC patients. Nevertheless, in these studies little attention was paid to the mutational status of homologous recombination repair (HRR) genes. Methods We tested the combination of the PARPi niraparib with either 5-fluorouracil, oxaliplatin or irinotecan (SN38) in a panel of 12 molecularly annotated CRC cell lines, encompassing the 4 consensus molecular subtypes (CMSs). Synergism was calculated using the Chou-Talalay method for drug interaction. A correlation between synergism and genetic alterations in genes involved in homologous recombination (HR) repair was performed. We used clonogenic assays, mice xenograft models and patient-derived 3D spheroids to validate the results. The induction of DNA damage was studied by immunofluorescence. Results We showed that human CRC cell lines, as well as patient-derived 3D spheroids, harboring pathogenic ATM mutations are significantly vulnerable to PARPi/chemotherapy combination at low doses, regardless of consensus molecular subtypes (CMS) and microsatellite status. The strongest synergism was shown for the combination of niraparib with irinotecan, and the presence of ATM mutations was associated to a delay in the resolution of double strand breaks (DSBs) through HRR and DNA damage persistence. Conclusions This work demonstrates that a numerically relevant subset of CRCs carrying heterozygous ATM mutations may benefit from the combination treatment with low doses of niraparib and irinotecan, suggesting a new potential approach in the treatment of ATM-mutated CRC, that deserves to be prospectively validated in clinical trials.

Background: The introduction of immunotherapy has improved the prognosis of patients with Non-Small Cell Lung Cancer (NSCLC). However, data in poor ECOG Performance Status (PS) patients remain scant due to their exclusion from randomized trials. Material and methods: We analyzed data of patients with advanced NSCLC treated with immunotherapy in two Italian Centers, to evaluate the impact of PS (0-1 vs 2) on disease control rate (DCR), progression free survival (PFS) and overall survival (OS). Chi-square test was used to compare clinical-pathological variables, their impact on survival was evaluated through Cox proportional hazard models. Results: Among 404 patients included, PS was 0 in 137 (33.9 %), 1 in 208 (51.5 %) and 2 in 59 (14.6 %) patients; 143 were female and 90 had squamous NSCLC. Clinical-pathological variables were uniformly distributed except for higher prevalence of liver metastases in patients with poor PS. We found that PS2 patients showed worse outcomes in terms of DCR (21.8 % vs 50.3 %, p = 0.001), PFS [2.0 (95 % CI 1.6–3.0) vs 3.0 (95 % CI 2.7–4.0) months, p < 0.0001] and OS [4.0 (95 % CI 2.8–5.7) vs 13.2 (95 % CI 11.0−15.8) months, p < 0.0001]. PS2 status, negative PDL1 expression and early corticosteroids exposure as well as higher Neutrophil to Lymphocyte Ratio and LDH at baseline were associated with worse outcomes at univariate and multivariable analysis. Subgroup analysis confirmed poor outcomes in PS2 patients with high LDH and concomitant corticosteroid therapies. The incidence of Grade 3/4 adverse events was 11.3 % in PS 0−1 and 10.2 % in PS 2 patients (p = 0.81). Conclusion: Our data confirm reduced efficacy of immunotherapy in patients with poor PS even though a good safety. Despite PS remains the most powerful independent prognostic factor for NSCLC, LDH levels and steroids exposure could support the decision making in PS2 patients.

Background The emerging role of next-generation sequencing (NGS) targeted panels is revolutionising our approach to cancer patients, providing information on gene alterations helpful for diagnosis and clinical decision, in a short time and with acceptable costs. Materials and methods In this work, we evaluated the clinical application of FoundationOne CDx test, a hybrid capture-based NGS. This test identifies alterations in 324 genes, tumour mutational burden and genomic signatures as microsatellite instability. The decision to obtain the NGS assay for a particular patient was done according to investigator's choice. Results Overall, 122 tumour specimens were analysed, of which 84 (68.85%) succeeded. The success rate was influenced by type of specimen formalin-fixed paraffin embedded (FFPE block vs FFPE slides), by origin of the sample (surgery vs biopsy) and by time of fixation (

Blocking angiogenesis represents a fundamental process in Colorectal Cancer (CRC) treatment. VEGF (vascular endothelial growth factor) pathway is implicated in various processes that regulate tumor vascularization and proliferation. In the last years, great efforts have been made thanks to the discovery of targeted drugs that block VEGF and its receptors conferring a benefit in a variety of tumors, including CRC. To date, four drugs have been approved for the treatment of metastatic CRC (mCRC): bevacizumab, aflibercept, ramucirumab and regorafenib. Unfortunately, patients relapse due to the appearance of resistance. The VEGF family, its role in the angiogenesis and complex heterogeneity of mechanisms that escape tumor blockade are not completely understood and there is a lack of biomarkers of response to anti-angiogenic drugs. We describe the principal mechanisms of resistance to anti-VEGF therapy and discuss potential biomarkers to be investigated in the near future.

Molecular heterogeneity is a frequent event in cancer responsible of several critical issues in diagnosis and treatment of oncologic patients. Lung tumours are characterized by high degree of molecular heterogeneity associated to different mechanisms of origin including genetic, epigenetic and non-genetic source. In this review, we provide an overview of recognized mechanisms underlying molecular heterogeneity in lung cancer, including epigenetic mechanisms, mutant allele specific imbalance, genomic instability, chromosomal aberrations, tumor mutational burden, somatic mutations. We focus on the role of spatial and temporal molecular heterogeneity involved in therapeutic implications in lung cancer patients.

Objectives: Never-smokers may be a distinct subgroup among patients with advanced non-small cell lung cancer, appearing to benefit less from immunotherapy than smokers. We report results from never-smokers enrolled in the Italian cohort of the nivolumab expanded access program in pre-treated patients with advanced squamous non-small cell lung cancer. Materials and methods: Nivolumab (3 mg/kg every 2 weeks for ≤24 months) was available on physician request. Efficacy data included objective tumor response, date of progression, and survival information. Safety was monitored. Results: Overall, 371 patients received at least one dose of nivolumab, including 31 never-smokers (8%). Objective response rate, disease-control rate, and median overall survival were 23%, 45%, and 12.1 months (95% confidence interval: 3.7–20.4), respectively, in never-smokers, and 18%, 47%, and 7.9 months (95% confidence interval: 6.2–9.6), respectively, in the overall expanded access program population. Any-grade and grade 3–4 treatment-related adverse events were reported in 12 (39%) and 3 (10%) never-smokers, respectively, and in 109 (29%) and 21 (6%) patients, respectively, in the overall expanded access program population. Grade 3–4 treatment-related adverse events in non-smokers were increased transaminases (n = 2; 6%) and diarrhea (n = 1; 3%). Treatment-related adverse events led to treatment discontinuation in 4 non-smokers (17%) and in 26 patients (9%) overall. Conclusion: Pre-treated never-smokers with advanced squamous non-small cell lung cancer in this Italian expanded access program demonstrated efficacy and safety that were consistent with those in the overall expanded access program population and clinical trials. These results suggest that a proportion of never-smoker patients with squamous non-small cell lung cancer may be responsive to immunotherapy. Other factors, such as the tumor mutational load and the status of programmed death-ligand 1, anaplastic lymphoma kinase, and epidermal growth factor receptor, might play a potential key role.

First-line pembrolizumab monotherapy improves overall and progression-free survival in patients with untreated metastatic non-small-cell lung cancer with a programmed death ligand 1 (PD-L1) tumour proportion score (TPS) of 50% or greater. We investigated overall survival after treatment with pembrolizumab monotherapy in patients with a PD-L1 TPS of 1% or greater.This randomised, open-label, phase 3 study was done in 213 medical centres in 32 countries. Eligible patients were adults (≥18 years) with previously untreated locally advanced or metastatic non-small-cell lung cancer without a sensitising EGFR mutation or ALK translocation and with an Eastern Cooperative Oncology Group (ECOG) performance status score of 0 or 1, life expectancy 3 months or longer, and a PD-L1 TPS of 1% or greater. Randomisation was computer generated, accessed via an interactive voice-response and integrated web-response system, and stratified by region of enrolment (east Asia vs rest of world), ECOG performance status score (0 vs 1), histology (squamous vs non-squamous), and PD-L1 TPS (≥50% vs 1-49%). Enrolled patients were randomly assigned 1:1 in blocks of four per stratum to receive pembrolizumab 200 mg every 3 weeks for up to 35 cycles or the investigator's choice of platinum-based chemotherapy for four to six cycles. Primary endpoints were overall survival in patients with a TPS of 50% or greater, 20% or greater, and 1% or greater (one-sided significance thresholds, p=0·0122, p=0·0120, and p=0·0124, respectively) in the intention-to-treat population, assessed sequentially if the previous findings were significant. This study is registered at ClinicalTrials.gov, number NCT02220894.From Dec 19, 2014, to March 6, 2017, 1274 patients (902 men, 372 women, median age 63 years [IQR 57-69]) with a PD-L1 TPS of 1% or greater were allocated to pembrolizumab (n=637) or chemotherapy (n=637) and included in the intention-to-treat population. 599 (47%) had a TPS of 50% or greater and 818 patients (64%) had a TPS of 20% or greater. As of Feb 26, 2018, median follow-up was 12·8 months. Overall survival was significantly longer in the pembrolizumab group than in the chemotherapy group in all three TPS populations (≥50% hazard ratio 0·69, 95% CI 0·56-0·85, p=0·0003; ≥20% 0·77, 0·64-0·92, p=0·0020, and ≥1% 0·81, 0·71-0·93, p=0·0018). The median surival values by TPS population were 20·0 months (95% CI 15·4-24·9) for pembrolizumab versus 12·2 months (10·4-14·2) for chemotherapy, 17·7 months (15·3-22·1) versus 13·0 months (11·6-15·3), and 16·7 months (13·9-19·7) versus 12·1 months (11·3-13·3), respectively. Treatment-related adverse events of grade 3 or worse occurred in 113 (18%) of 636 treated patients in the pembrolizumab group and in 252 (41%) of 615 in the chemotherapy group and led to death in 13 (2%) and 14 (2%) patients, respectively.The benefit-to-risk profile suggests that pembrolizumab monotherapy can be extended as first-line therapy to patients with locally advanced or metastatic non-small-cell lung cancer without sensitising EGFR or ALK alterations and with low PD-L1 TPS.Merck SharpDohme.

Malignant pleural mesothelioma is a rare and aggressive malignancy mostly associated with occupational asbestos exposure. Prognosis is poor and only highly selected patients may benefit from aggressive surgical management, also as part of a multimodal approach. In advanced disease, the combination of pemetrexed and platinum remains the only established treatment, while efficacy evidence of second line chemotherapy is lacking. Thus, a deeper knowledge of biology of the disease and more effective treatments are urgently needed. Refer to specialised centres with multidisciplinary expertise is mandatory, as well as inclusion of patients in clinical trials is advisable whenever possible. In all stages of disease focus on symptoms control is paramount.

The lung cancer is the principle cause of the worldwide deaths and its prognosis is poor with a 5-year overall survival rate. Computed tomography (CT) gives many information about the prognosis, but the problem is the subject interpretation of the findings. Thanks to the computer-aided diagnosis/detection (CAD), it is possible to reduce the second opinion. "Radiomics" is an extension of CAD and overlaps the quantitative imaging data of the CT texture analysis (CTTA) with the clinical information, increasing the power and precision of the decision going through the personalized medicine. The aim of this study is to describe the role of the radiomics in the characterization of the pulmonary nodule. For this study, we retrospectively analyzed the images of the 87 NSCLC patients with a waiver of informed consent from the Institutional Review Board (IRB) at the Campania University "Luigi Vanvitelli" of Naples. All tumors were semiautomatically segmented by a radiologist with 10years of experience using three diameters (AW Server 3.2). The examinations were acquired using 128 MDCT (GSI CT, GE) with a peak tube voltage of 120 kVp, tube current of 100 or 200mA, and rotation times of 0.5 or 0.8s. To confirm the imaging results, the FNAC was performed and for every nodule the following parameters were extracted: the presence of the solid component (named = 1), papillary component (named = 2), and mixed component (named = 3). Feature calculation was performed using the HealthMyne software and Integrated Platform That Enables Better Patient Management Decisions For Oncology. The radiologist uses the Rapid Precise Metrics (RPM)™ functionality to identify a lesion with the algorithm and these methods are put to work. The correlation between each feature and the tumor volume was calculated using a two-step cluster statistical analysis. In this retrospective study, in one year from 2018 to 2019 20 patients with lung adenocarcinoma confirmed with FNAC were enrolled. The pathologic results were subdivided into three categories: the solid architecture (group 1), papillary architecture (group 2), and mixed architecture (group 3). Nine lesions resulted with component 1, seven patients with component 2, and 3 patients with component 3. Eight females and 12 males with a median age 61 and 15years (mean ± SD = 67.4 ± 9.7years, range 39-73years) were enrolled. The two results suggest, with p

Molecular profiling of a tumor allows the opportunity to design specific therapies which are able to interact only with cancer cells characterized by the accumulation of several genomic aberrations. This study investigates the usefulness of next-generation sequencing (NGS) and mutation-specific analysis methods for the detection of target genes for current therapies in non-small-cell lung cancer (NSCLC), metastatic colorectal cancer (mCRC), and melanoma patients. We focused our attention on EGFR, BRAF, KRAS, and BRAF genes for NSCLC, melanoma, and mCRC samples, respectively. Our study demonstrated that in about 2% of analyzed cases, the two techniques did not show the same or overlapping results. Two patients affected by mCRC resulted in wild-type (WT) for BRAF and two cases with NSCLC were WT for EGFR according to PGM analysis. In contrast, these samples were mutated for the evaluated genes using the therascreen test on Rotor-Gene Q. In conclusion, our experience suggests that it would be appropriate to confirm the WT status of the genes of interest with a more sensitive analysis method to avoid the presence of a small neoplastic clone and drive the clinician to correct patient monitoring.

Tumor heterogeneity represents a possible cause of error in detecting predictive genetic alterations on tumor tissue and can be overcome by testing alterations in circulating tumor DNA (ctDNA) using liquid biopsy. We assessed 72 consecutive patients with a diagnosis of metastatic colorectal cancer (mCRC) using Idylla&trade, Biocartis, a fully automated platform that evaluates the most frequent mutations of KRAS, NRAS and BRAF genes. We correlated the results of liquid biopsy and standard tissue-based next generation sequencing (NGS) analyses to patient clinical features. The overall agreement was 81.94%. Concordance was 85.71% and 96.15% in treatment-na&iuml, ve patients and in the patient subgroup with liver metastases, respectively. In liver metastases positive, treatment-na&iuml, ve patients, sensitivity, specificity and positive predictive value (PPV) were 92.31%, 100% and 100%, respectively. Circulating mutational fraction (CMF) was significantly higher in patients with liver metastases and high carcinoembryonic antigen (CEA) levels. In a subgroup of patients pre-treated with anti-Epidermal Growth Factor Receptor (EGFR) agents, emerging KRAS mutations were evidenced in 33% of cases. Testing RAS/BRAF mutations on plasma using the Idylla&trade, Biocartis platform is feasible and reliable in mCRC patients in clinical practice.

Background Targeting the epidermal growth factor receptor (EGFR) either alone or in combination with chemotherapy is an effective treatment for patients with RAS wild-type metastatic colorectal cancer (mCRC). However, only a small percentage of mCRC patients receive clinical benefits from anti-EGFR therapies, due to the development of resistance mechanisms. In this regard, HER2 has emerged as an actionable target in the treatment of mCRC patients with resistance to anti-EGFR therapy. Methods We have used SW48 and LIM1215 human colon cancer cell lines, quadruple wild-type for KRAS, NRAS, BRAF and PI3KCA genes, and their HER2–amplified (LIM1215-HER2 and SW48-HER2) derived cells to perform in vitro and in vivo studies in order to identify novel therapeutic strategies in HER2 gene amplified human colorectal cancer. Results LIM1215-HER2 and SW48-HER2 cells showed over-expression and activation of the HER family receptors and concomitant intracellular downstream signaling including the pro-survival PI3KCA/AKT and the mitogenic RAS/RAF/MEK/MAPK pathways. HER2-amplified cells were treated with several agents including anti-EGFR antibodies (cetuximab, SYM004 and MM151); anti-HER2 (trastuzumab, pertuzumab and lapatinib) inhibitors; anti-HER3 (duligotuzumab) inhibitors; and MEK and PI3KCA inhibitors, such as refametinib and pictilisib, as single agents and in combination. Subsequently, different in vivo experiments have been performed. MEK plus PI3KCA inhibitors treatment determined the best antitumor activity. These results were validated in vivo in HER2-amplified patient derived tumor xenografts from three metastatic colorectal cancer patients. Conclusions These results suggest that combined therapy with MEK and PI3KCA inhibitors could represent a novel and effective treatment option for HER2-amplified colorectal cancer. Electronic supplementary material The online version of this article (10.1186/s13046-019-1230-z) contains supplementary material, which is available to authorized users.

Introduction In the era of precision medicine, research studies are aiming to design patient-tailored treatment strategies. In this work, we present a clinical case of a patient with non-small cell lung cancer (NSCLC) accompanied by a translational study with the intent to assess the correspondence of drug sensitivity in ex vivo spheroidal tumour cultures and peripheral blood biomarkers with clinical outcome. Methods Primary tumour tissue, patient-derived tumour spheroids, peripheral blood mononuclear cells and circulating DNA were analysed to assess drug sensitivity and immunological profiling, and all these data were correlated with clinical and radiological evaluations. Results Immunohistochemistry, immunofluorescence, next generation sequencing analysis and T-lymphocyte receptor repertoire assay results showed elevated concordance among primary tumour tissue, ex vivo three-dimensional tumour spheroid specimen and circulating DNA. Cisplatin-based chemotherapy and anti-programmed death 1 drug sensitivity assessed in spheroidal cultures were strictly consistent with patient clinical response to adjuvant chemotherapy and first-line immune therapy. Conclusion These results revealed that ex vivo drug sensitivity testing in three-dimensional spheroidal culture can reproduce clinical response to chemotherapy and immunotherapy, with the potential to use those culture models to predict patients‘ outcome from anticancer treatments and, therefore, the feasibility to select individualised therapy.

Background Pioglitazone, a synthetic peroxisome proliferator activated receptor (PPAR-γ) ligand, is known as an antidiabetic drug included in the thiazolidinediones (TZDs) class. It regulates the lipid and glucose cell metabolism and recently a role in the inhibition of numerous cancer cell processes has been described. Methods In our work we investigate the anti-tumor effects of pioglitazone in in vitro models of non small cell lung cancer (NSCLC) and also, we generated ex-vivo three-dimensional (3D) cultures from human lung adenocarcinoma (ADK) as a model to test drug efficacy observed in vitro. The inhibitory effect of pioglitazone on cell proliferation, apoptosis and cell invasion in a panel of human NSCLC cell lines was evaluated by multiple assays. Results Pioglitazone reduced proliferative and invasive abilities with an IC50 ranging between 5 and 10 μM and induced apoptosis of NSCLC cells. mRNA microarray expression profiling showed a down regulation of MAPK, Myc and Ras genes after treatment with pioglitazone; altered gene expression was confirmed by protein analysis in a dose-related reduction of survivin and phosphorylated proteins levels of MAPK pathway. Interestingly mRNA microarray analysis showed also that pioglitazone affects TGFβ pathway, which is important in the epithelial-to-mesenchimal transition (EMT) process, by down-regulating TGFβR1 and SMAD3 mRNA expression. In addition, extracellular acidification rate (ECAR) and a proportional reduction of markers of altered glucose metabolism in treated cells demonstrated also cell bioenergetics modulation by pioglitazone. Conclusions Data indicate that PPAR-γ agonists represent an attractive treatment tool and by suppression of cell growth (in vitro and ex vivo models) and of invasion via blockade of MAPK cascade and TGFβ/SMADs signaling, respectively, and its role in cancer bioenergetics and metabolism indicate that PPAR-γ agonists represent an attractive treatment tool for NSCLC.

Non-small cell lung cancer (NSCLC) is the most common type of lung cancer and the ineffectiveness of the current therapies seriously limits the survival rate of NSCLC patients. In the search for new antitumor agents, nature has played a pivotal role providing a variety of molecules, which are likely to exert selective anti-tumour properties. Herein, we investigated the antiproliferative potential of Urtica dioica L. extract (UD) against NSCLC cell models with low sensitivity to cisplatin, a cytotoxic agent largely employed to cure NSCLCs. UD inhibited cell proliferation in the selected cells, while no toxic effects were observed in normal lung cells. Furthermore, the co-treatment of UD and cisplatin notably sensitised NSCLC cells to cisplatin. Mechanistically, we discovered that UD-promoted endoplasmic reticulum (ER) stress via activation of the growth arrest and DNA damage-inducible gene 153 (GADD153) triggering apoptosis. We also performed an extensive NMR analysis of UD, identifying rutin and oxylipins as the main secondary metabolites present in the mixture. Additionally, we discovered that an oxylipins’ enriched fraction contributes to the antiproliferative activity of the plant extract. In the future, this study may provide new chemical scaffolds for the design of anti-cancer agents that target NSCLCs with low sensitivity to cisplatinum.

Background: Autoimmune hypophysitis is a frequent immune-related adverse event (irAE) in cancer patients treated with immunecheckpoint inhibitors. Studies seeking anti-pituitary (APA) and anti-hypothalamus (AHA) antibodies in patients treated with anti-PD-1 and anti-PD-L1 are scarce. The aim of this study is to search for APA and AHA and related pituitary dysfunction in patients treated with these agents. Methods:Cross-sectional and preliminary longitudinal studies were conducted at the Medical Oncology Unit and Endocrinology and Metabolic Diseases Unit of the University of Campania “Luigi Vanvitelli”. Fifty-four cancer patients on treatments with anti-PD-1 or anti-PD-L1 (Group 1) and 50 healthy controls were enrolled for a cross-sectional study, 13 cancer patients (Group 2) were enrolled for our preliminary longitudinal study. APA/AHA titers and changes in biochemical and hormonal profile were evaluated in Group 1, in Group 2, they were evaluated before and after nine weeks from the start of immunotherapy. Results: Patients of Group 1 showed a higher prevalence of APA and AHA than controls: 21 of them had APA, 16 had AHA, and 11 had both autoantibodies. In total, 7 of 13 patients in Group 2 became APA-positive and 3 became AHA-positive after nine weeks of immunotherapy, showing an increase in prolactin and a decrease in ACTH and IGF-1 levels compared with basal values. Conclusions:Anti-pituitary and anti-hypothalamus antibodies seem to play a pivotal role in hypothalamic–pituitary autoimmunity and secondary endocrine-related alterations evoked by anti-PD-1 and PD-L1 antibodies.

// Carminia Maria Della Corte 1 , Umberto Malapelle 2 , Elena Vigliar 2 , Francesco Pepe 2 , Giancarlo Troncone 2 , Vincenza Ciaramella 1 , Teresa Troiani 1 , Erika Martinelli 1 , Valentina Belli 1 , Fortunato Ciardiello 1 , Floriana Morgillo 1 1 Oncologia Medica, Dipartimento Medico-Chirurgico di Internistica Clinica e Sperimentale “F. Magrassi e A. Lanzara,” Universita degli studi della Campania “Luigi Vanvitelli”, Naples, Italy 2 Dipartimento di Sanita Pubblica, Universita degli Studi di Napoli Federico II, Naples, Italy Correspondence to: Floriana Morgillo, email: florianamorgillo@yahoo.com Keywords: EGFR inhibitors, lung cancer, cell signalling, hedgehog, EMT Received: January 13, 2017 Accepted: February 08, 2017 Published: February 18, 2017 ABSTRACT Purpose: The aim of this work was to investigate the efficacy of sequential treatment with first-, second- and third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors and the mechanisms of acquired resistance occurring during the sequential use of these inhibitors. Experimental design: We developed an in vivo model of acquired resistance to EGFR-inhibitors by treating nude mice xenografted with HCC827, a human non-small-cell lung cancer (NSCLC) cell line harboring EGFR activating mutation, with a sequence of first-generation EGFR tyrosine kinase inhibitors (EGFR-TKIs) (erlotinib and gefitinib), of second-generation EGFR-TKI (afatinib) plus/minus the anti-EGFR monoclonal antibody cetuximab, and of third-generation EGFR-TKI (osimertinib). Results: HCC827-derived xenografts and with acquired resistance to EGFR-inhibitors were sensitive to the sequential use of first-, second- and third-generation EGFR-TKIs. Continuous EGFR inhibition of first-generation resistant tumors by sequential treatment with afatinib plus/minus cetuximab, followed by osimertinib, represented an effective therapeutic strategy in this model. Whereas T790M resistance mutation was not detected, a major mechanism of acquired resistance was the activation of components of the Hedgehog (Hh) pathway. This phenomenon was accompanied by epithelial-to-mesenchymal transition. Cell lines established in vitro from gefitinib-, or afatinib- or osimertinib-resistant tumors showed metastatic properties and maintained EGFR-TKIs resistance in vitro , that was reverted by the combined blockade of Hh, with the selective SMO inhibitor sonidegib, and EGFR. Conclusions: EGFR -mutant NSCLC can benefit from continuous treatment with EGFR-inhibitors, indepenently from mechanisms of resistance. In a complex and heterogenous scenario, Hh showed an important role in mediating resistance to EGFR-inhibitors through the induction of mesenchymal properties.

BACKGROUND: About 20% of advanced non-small-cell lung cancer (NSCLC) cases harbor somatic mutations in the tyrosine kinase domain of the epidermal growth factor receptor (EGFR) gene. In these patients, the standard first-line treatments are the EGFR-tyrosine kinase inhibitors, such as gefitinib, erlotinib, or afatinib. Most of these patients develop resistance and relapse within about 1 year of initiation of an EGFR-tyrosine kinase inhibitor. Consequently, it is important to develop new combination strategies to delay this resistance. Preclinical data have showed that EGFR and vascular endothelial growth factor (VEGF) share a common downstream pathway, suggesting the important role of VEGF in the resistance to EGFR blockade. The combination of erlotinib and bevacizumab, an anti-VEGF agent, showed very interesting clinical results. PATIENTS AND METHODS: The bevacizumab plus erlotinib study (BEVERLY) is a randomized, open-label, phase III trial investigating first-line erlotinib plus bevacizumab versus erlotinib in patients with advanced NSCLC harboring activating EGFR mutations. The co-primary endpoints are investigator-assessed progression-free survival (PFS) and blinded, independent centrally reviewed PFS. The secondary endpoints include overall survival, quality of life, objective response rate, and safety. A total of 200 patients will be randomized 1:1 to receive oral erlotinib (150 mg daily) plus bevacizumab (15 mg/kg, intravenously, on day 1 of every 21-day cycle) or erlotinib alone, until objective disease progression or unacceptable toxicity or the patient's or physician's motivated decision to stop the treatment. CONCLUSION: If the primary endpoint of PFS is met, the erlotinib plus bevacizumab combination will be confirmed as the best first-line treatment for patients with advanced NSCLC harboring activating EGFR mutations. Background About 20% of advanced non–small-cell lung cancer (NSCLC) cases harbor somatic mutations in the tyrosine kinase domain of the epidermal growth factor receptor (EGFR) gene. In these patients, the standard first-line treatments are the EGFR-tyrosine kinase inhibitors, such as gefitinib, erlotinib, or afatinib. Most of these patients develop resistance and relapse within about 1 year of initiation of an EGFR-tyrosine kinase inhibitor. Consequently, it is important to develop new combination strategies to delay this resistance. Preclinical data have showed that EGFR and vascular endothelial growth factor (VEGF) share a common downstream pathway, suggesting the important role of VEGF in the resistance to EGFR blockade. The combination of erlotinib and bevacizumab, an anti-VEGF agent, showed very interesting clinical results. Patients and Methods The bevacizumab plus erlotinib study (BEVERLY) is a randomized, open-label, phase III trial investigating first-line erlotinib plus bevacizumab versus erlotinib in patients with advanced NSCLC harboring activating EGFR mutations. The co-primary endpoints are investigator-assessed progression-free survival (PFS) and blinded, independent centrally reviewed PFS. The secondary endpoints include overall survival, quality of life, objective response rate, and safety. A total of 200 patients will be randomized 1:1 to receive oral erlotinib (150 mg daily) plus bevacizumab (15 mg/kg, intravenously, on day 1 of every 21-day cycle) or erlotinib alone, until objective disease progression or unacceptable toxicity or the patient's or physician's motivated decision to stop the treatment. Conclusion If the primary endpoint of PFS is met, the erlotinib plus bevacizumab combination will be confirmed as the best first-line treatment for patients with advanced NSCLC harboring activating EGFR mutations.

Background Anti-PD-1/PD-L1 drugs are effective as monotherapy in a proportion of NSCLC patients and there is a strong rationale for combining them with targeted therapy. Inhibition of MAPK pathway may have pleiotropic effects on the microenvironment. This work investigates the efficacy of combining MEK and PD-L1 inhibition in pre-clinical and ex-vivo NSCLC models. Methods We studied the effects of MEK inhibitors (MEK-I) on PD-L1 and MCH-I protein expression and cytokine production in vitro in NSCLC cell lines and in PBMCs from healthy donors and NSCLC patients, the efficacy of combining MEK-I with anti-PD-L1 antibody in ex-vivo human spheroid cultures obtained from fresh biopsies from NSCLC patients in terms of cell growth arrest, cytokine production and T-cell activation by flow cytometry. Results MEK-I modulates in–vitro the immune micro-environment through a transcriptionally decrease of PD-L1 expression, enhance of MHC-I expression on tumor cells, increase of the production of several cytokines, like IFNγ, IL-6, IL-1β and TNFα. These effects trigger a more permissive anti-tumor immune reaction, recruiting immune cells to the tumor sites. We confirmed these data on ex-vivo human spheroids, showing a synergism of MEK and PD-L1 inhibition as result of both direct cancer cell toxicity of MEK-I and its immune-stimulatory effect on cytokine secretion profile of cancer cells and PBMCs with the induction of the ones that sustain an immune-reactive and inflammatory micro-environment. Conclusions Our work shows the biological rationale for combining immunotherapy with MEK-I in a reproducible ex-vivo 3D-culture model, useful to predict sensitivity of patients to such therapies. Electronic supplementary material The online version of this article (10.1186/s13046-019-1257-1) contains supplementary material, which is available to authorized users.

Colorectal cancer; Epidermal growth factor receptor (EGFR); MAPK pathway Càncer colorectal; Receptor epidèrmic de factor del creixement (EGFR); Via MAPK Cáncer colorrectal; Receptor epidérmico de factor del crecimiento (EGFR); Vía MAPK BACKGROUND: Previous studies showed that the combination of an anti-Epidermal growth factor (EGFR) and a MEK-inhibitor is able to prevent the onset of resistance to anti-EGFR monoclonal antibodies in KRAS-wild type colorectal cancer (CRC), while the same combination reverts anti-EGFR primary resistance in KRAS mutated CRC cell lines. However, rapid onset of resistance is a limit to combination therapies in KRAS mutated CRC. METHODS: We generated four different KRAS mutated CRC cell lines resistant to a combination of cetuximab (an anti-EGFR antibody) and refametinib (a selective MEK-inhibitor) after continuous exposure to increasing concentration of the drugs. We characterized these resistant cell lines by evaluating the expression and activation status of a panel of receptor tyrosine kinases (RTKs) and intracellular transducers by immunoblot and qRT-PCR. Oncomine comprehensive assay and microarray analysis were carried out to investigate new acquired mutations or transcriptomic adaptation, respectively, in the resistant cell lines. Immunofluorescence assay was used to show the localization of RTKs in resistant and parental clones. RESULTS: We found that PI3K-AKT pathway activation acts as an escape mechanism in cell lines with acquired resistance to combined inhibition of EGFR and MEK. AKT pathway activation is coupled to the activation of multiple RTKs such as HER2, HER3 and IGF1R, though its pharmacological inhibition is not sufficient to revert the resistant phenotype. PI3K pathway activation is mediated by autocrine loops and by heterodimerization of multiple receptors. CONCLUSIONS: PI3K activation plays a central role in the acquired resistance to the combination of anti-EGFR and MEK-inhibitor in KRAS mutated colorectal cancer cell lines. PI3K activation is cooperatively achieved through the activation of multiple RTKs such as HER2, HER3 and IGF1R.

Background Molecular mechanisms driving acquired resistance to anti-EGFR therapies in metastatic colorectal cancer (mCRC) are complex but generally involve the activation of the downstream RAS-RAF-MEK-MAPK pathway. Nevertheless, even if inhibition of EGFR and MEK could be a strategy for overcoming anti-EGFR resistance, its use is limited by the development of MEK inhibitor (MEKi) resistance. Methods We have generated in vitro and in vivo different CRC models in order to underline the mechanisms of MEKi resistance. Results The three different in vitro MEKi resistant models, two generated by human CRC cells quadruple wild type for KRAS, NRAS, BRAF, PI3KCA genes (SW48-MR and LIM1215-MR) and one by human CRC cells harboring KRAS mutation (HCT116-MR) showed features related to the gene signature of colorectal cancer CMS4 with up-regulation of immune pathway as confirmed by microarray and western blot analysis. In particular, the MEKi phenotype was associated with the loss of epithelial features and acquisition of mesenchymal markers and morphology. The change in morphology was accompanied by up-regulation of PD-L1 expression and activation of EGFR and its downstream pathway, independently to RAS mutation status. To extend these in vitro findings, we have obtained mouse colon cancer MC38- and CT26-MEKi resistant syngeneic models (MC38-MR and CT26-MR). Combined treatment with MEKi, EGFR inhibitor (EGFRi) and PD-L1 inhibitor (PD-L1i) resulted in a marked inhibition of tumor growth in both models. Conclusions These results suggest a strategy to potentially improve the efficacy of MEK inhibition by co-treatment with EGFR and PD-L1 inhibitors via modulation of host immune responses.

Background The activation of multiple signaling pathways jeopardizes the clinical efficacy of EGFR tyrosine kinase inhibitors (TKIs) in EGFR-mutation positive non-small cell lung cancer (NSCLC). Integrin-linked kinase (ILK) regulates the interactions between tumor cells and extracellular environment to activate signaling pathways and promote cell proliferation, migration, and epithelial-mesenchymal transition. Src homology 2 domain-containing phosphatase 2 (SHP2) is essential for receptor tyrosine kinase signaling and mitogen-activated protein kinase (MAPK) pathway activation. Methods We analyzed tumor ILK, β-receptor subunit glycoprotein 130 (gp130), SHP2, and stromal hepatocyte growth factor (HGF) and interleukin-6 (IL-6) mRNA expression in baseline tumor specimens of advanced EGFR-mutation positive NSCLC patients treated with EGFR TKIs. Results ILK, when highly expressed, was an independent poor prognostic factor for the progression-free survival of the patients, both in the univariate (hazard ratio [HR for disease progression, 2.49; 95% CI, 1.37–4.52; P = .0020]) and in the multivariate (HR 3.74; 95% CI, 1.33–10.56; P = .0126) Cox regression model. Patients with high SHP2 expression had an almost 13-month shorter progression-free survival (P = .0094) and an 18-month shorter overall survival (P = .0182) in comparison to those with low SHP2 mRNA expression. Interpretation The levels of ILK and SHP2 could be predictive for upfront combinatory therapy of EGFR TKIs plus SHP2 or ILK inhibitors. Fund A grant from La Caixa Foundation, an Instituto de Salud Carlos III grant (RESPONSE, PIE16/00011), an Instituto de Salud Carlos III grant (PI14/01678), a Marie Skłodowska-Curie Innovative Training Networks European Grant (ELBA No 765492) and a Spanish Association Against Cancer (AECC) grant (PROYE18012ROSE).

An overactivation of hepatocyte growth factor (HGF)/mesenchymal-epithelial transition factor (MET) axis promotes tumorigenesis and tumor progression in various cancer types. Research data recently evidenced that HGF/MET signaling is also involved also in the immune response, mainly modulating dendritic cells functions. In general, the pathway seems to play an immunosuppressive role, thus hypothesizing that it could constitute a mechanism of primary and acquired resistance to cancer immunotherapy. Recently, some approaches are being developed, including drug design and cell therapy to combine MET and programmed cell death receptor-1 (PD-1)/programmed cell death receptor-ligand 1 (PD-L1) inhibition. This approach could represent a new weapon in cancer therapy in the future.

The EPHA2 tyrosine kinase receptor is implicated in tumor progression and targeted therapies resistance. We evaluated EPHA2 as a potential resistance marker to the antiepidermal growth factor receptor (EGFR) monoclonal antibody cetuximab in colorectal cancer. We studied activation of EPHA2 in a panel of human colorectal cancer cell lines sensitive or resistant to anti-EGFR drugs. The in vitro and in vivo effects of ALW-II-41-27 (an EPHA2 inhibitor) and/or cetuximab treatment were tested. Formalin-fixed paraffin-embedded tumor specimens from 82 RAS wild-type (WT) metastatic colorectal cancer patients treated with FOLFIRI + cetuximab as first-line therapy in the CAPRI-GOIM trial were assessed for EPHA2 expression by immunohistochemistry and correlated with treatment efficacy. EPHA2 was differentially activated in colorectal cancer cell lines. Combined treatment with ALW-II-41-27 plus cetuximab reverted primary and acquired resistance to cetuximab, causing cell growth inhibition, inducing apoptosis and cell-cycle G1–G2 arrest. In tumor xenograft models, upon progression to cetuximab, ALW-II-41-27 addition significantly inhibited tumor growth. EPHA2 protein expression was detected in 55 of 82 tumor samples, frequently expressed in less-differentiated and left-sided tumors. High levels of EPHA2 significantly correlated with worse progression-free survival [8.6 months; confidence interval (CI) 95%, 6.4–10.8; vs. 12.3 months; CI 95%, 10.4–14.2; P = 0.03] and with increased progression rate (29% vs. 9%, P = 0.02). A specific EPHA2 inhibitor reverts in vitro and in vivo primary and acquired resistance to anti-EGFR therapy. EPHA2 levels are significantly associated with worse outcome in patients treated with FOLFIRI + cetuximab. These results highlight EPHA2 as a potential therapeutic target in metastatic colorectal cancer.

Two inhibitors of phosphatidylinositol 3-kinase (PI3K) pathway taselisib, targeting the mutant PI3K-subunit-alpha (PI3KA) and ipatasertib, AKT-inhibitor, are currently under clinical investigation in breast cancer (BC) patients. We have previously demonstrated the anti-tumor efficacy of these anti-PI3K/AKT-inibitors in combination with anti-microtubule drugs in human BC cell lines, through a complete cytoskeleton disorganization. In this work, we generated ex-vivo three-dimensional (3D) cultures from human BC as a model to test drug efficacy and to identify new molecular biomarkers for selection of BC patients suitable for anti-PI3K/AKT-inibitors treatment. We have established 3D cultures from 25/27 human BC samples, in which the ability of growth in vitro replicates the clinical and biological aggressiveness of the original tumors. According to the results of next generation sequencing analysis, a direct correlation was found between PI3KA mutations and the sensitivity in 3D models in vitro to taselisib and ipatasertib alone and combined with anti-microtubule agents. Moreover, mutations in HER and MAPK families related genes, including EGFR, KRAS and BRAF, were found in resistant samples, suggesting their potential role as negative predictive factors of response to these agents. Thus, we demonstrated that ex vivo 3D cultures from human BC patients allow a rapid and efficient drug screening for chemotherapies and targeted agents in genetically selected patients and represent an innovative model to identify new biomarkers of drug resistance.

Cancer disease is the second leading cause of death in the world and one of the main fields of medical research. Although there is now a greater understanding of biological mechanisms of uncontrolled cell growth, invasiveness and metastasization, the multi-step process of cancer development and evolution is still incompletely understood. The inhibition of molecules activated in cancer metastasization is an hot topic in cancer research. Among the known antimetastatic genes, KiSS-1 is involved in the metastatic cascade by preventing growth of metastasis. Moreover, loss of KiSS-1 protein expression by tumor cells has been associated with a more aggressive phenotype. KiSS-1 gene encodes a 145-amino acid protein, which following proteolytic cleavage, generates a family of kisspeptins (Kp-10, -13, and -14), that are endogenous agonists for the G-protein-coupled receptor (GPR54). The antitumor effect of KiSS-1 was primarily associated with the inhibition of proliferation, migration and cell invasion and, consequently, the reduced formation of metastasis and intratumoral microvessels. In this review, we highlight the latest data on the role of kisspeptin signaling in the suppression of metastasis in various cancer types and the use modulators of KiSS/GPR54 signaling as potential novel therapeutic agents for the treatment of cancer.

Purpose: The use of cetuximab in the treatment of metastatic colorectal cancer is limited by development of resistance. Experimental Design: We have investigated in three models of highly epidermal growth factor receptor (EGFR)–dependent colorectal cancer xenografts, the effect of maintenance therapy with different kinase inhibitors alone or in combination with cetuximab, after cytotoxic treatment induction with irinotecan plus cetuximab. Results: SW48, LIM 1215, and GEO colorectal cancer cell lines were engrafted into nude mice and treated for 3 weeks with irinotecan and/or cetuximab. The combined treatment induced a significant reduction of tumor size. A subsequent experiment was performed in all three xenograft models in which after an induction treatment with irinotecan plus cetuximab, mice were randomly assigned to one of the following treatments: control, cetuximab, regorafenib, a selective PIK3CA inhibitor (PIK3CAi), a selective MEK inhibitor (MEKi), and/or the combination of each inhibitor with cetuximab. The cetuximab plus MEKi treatment determined the best antitumor activity with suppression of tumor growth. This effect was prolonged for 13 to 15 weeks after cessation of therapy and was accompanied by prolonged survival. Antitumor activity was accompanied by inhibition of the MAPK and MEK pathways. Moreover, in the cetuximab plus MEKi-treated SW48 xenograft group, KRAS mutations as a mechanism of acquired resistance were detected in 25% of cases compared with 75% KRAS mutations in the MEKi-treated group. Conclusions: A possible strategy to prevent and/or overcome resistance to anti-EGFR inhibitors in metastatic colorectal cancer is a maintenance therapy with cetuximab plus MEKi after an initial treatment with irinotecan plus cetuximab. Clin Cancer Res; 21(18); 4153–64. ©2015 AACR.

AXL is a tyrosine kinase receptor activated by GAS6 and regulates cancer cell proliferation migration and angiogenesis. We studied AXL as new therapeutic target in colorectal cancer (CRC). Expression and activation of AXL and GAS6 were evaluated in a panel of human CRC cell lines. AXL gene silencing or pharmacologic inhibition with foretinib suppressed proliferation, migration and survival in CRC cells. In an orthotopic colon model of human HCT116 CRC cells overexpressing AXL, foretinib treatment caused significant inhibition of tumour growth and peritoneal metastatic spreading. AXL and GAS6 overexpression by immunohistochemistry (IHC) were found in 76,7% and 73.5%, respectively, of 223 human CRC specimens, correlating with less differentiated histological grading. GAS6 overexpression was associated with nodes involvement and tumour stage. AXL gene was found amplified by Fluorescence in situ hybridization (FISH) in 8/146 cases (5,4%) of CRC samples. Taken together, AXL inhibition could represent a novel therapeutic approach in CRC.

Anaplastic lymphoma kinase (ALK) gene activation is involved in the carcinogenesis process of several human cancers such as anaplastic large cell lymphoma, lung cancer, inflammatory myofibroblastic tumors and neuroblastoma, as a consequence of fusion with other oncogenes (NPM, EML4, TIM, etc) or gene amplification, mutation or protein overexpression. ALK is a transmembrane tyrosine kinase receptor that, upon ligand binding to its extracellular domain, undergoes dimerization and subsequent autophosphorylation of the intracellular kinase domain. When activated in cancer it represents a target for specific inhibitors, such as crizotinib, ceritinib, alectinib etc. which use has demonstrated significant effectiveness in ALK-positive patients, in particular ALK-positive non- small cell lung cancer. Several mechanisms of resistance to these inhibitors have been described and new strategies are underway to overcome the limitations of current ALK inhibitors.

// Vincenza Ciaramella 1 , Carminia Maria Della Corte 1 , Concetta Di Mauro 2 , Stefano Tomassi 3 , Salvatore Di Maro 3 , Teresa Troiani 1 , Erika Martinelli 1 , Roberto Bianco 2 , Sandro Cosconati 3 , Riccardo Pierantoni 4 , Rosaria Meccariello 5 , Rosanna Chianese 4 , Fortunato Ciardiello 1 and Floriana Morgillo 1 1 Oncologia Medica, Dipartimento di Internistica Clinica e Sperimentale “F. Magrassi”, Universita degli Studi della Campania Luigi Vanvitelli, Naples, Italy 2 Oncologia Medica, Dipartimento di Medicina Clinica e Chirurgia, Universita degli Studi di Napoli “Federico II”, Naples, Italy 3 DISTABIF, Universita degli Studi della Campania Luigi Vanvitelli, Caserta, Italy 4 Dipartimento di Medicina Sperimentale sez ‘F. Bottazzi’, Universita degli Studi della Campania Luigi Vanvitelli, Naples, Italy 5 Dipartimento di Scienze Motorie e del Benessere, Universita di Napoli Parthenope, Napoli, Italy Correspondence to: Floriana Morgillo, email: florianamorgillo@yahoo.com Keywords: mesothelioma; KiSS1; biomarker; metastasis; EMT Received: February 03, 2018 Accepted: February 27, 2018 Published: April 10, 2018 ABSTRACT Purpose: Kisspeptin signaling, via its receptors GPR54, could be an essential players in the inhibition of mesothelioma progression, invasion and metastasis formation. The loss of KiSS1 by tumor cells has been associated with a metastatic phenotype but the mechanistic insights of this process are still unknown. Experimental design: The blockade of the metastatic process at early stage is a hot topic in cancer research. We studied the role of KiSS1 on proliferation, invasiveness, migration abilities of mesothelioma cell lines focusing on the effect on epithelial-to-mesenchymal transition (EMT). Results: Treatment with the KiSS1 peptide or with a synthesis peptide with longer half-life, the FTM080, significantly inhibited cell proliferation, migration and invasion of mesothelioma cell lines; the same treatment reduced the activity of MMP-2 and MMP-9 determining consequently a marked reduction in the invasiveness of primary tumors and metastases. Thespecificexpression of EMT markers, as E-caderin, Vimentin, Slug and Snail, suggested the inhibition of EMT after treatment with KiSS1 as well as the preservation of epithelial components. Conclusion: Our results support anti-proliferative effect of KiSS1 in cancer cells and suggest that targeting the KiSS1/GPR54 system may represent a novel therapeutic approach for mesothelioma.

// Stefania Napolitano 1 , Giulia Martini 1 , Erika Martinelli 1 , Carminia Maria Della Corte 1 , Floriana Morgillo 1 , Valentina Belli 1 , Claudia Cardone 1 , Nunzia Matrone 1 , Fortunato Ciardiello 1 and Teresa Troiani 1 1 Oncologia Medica, Dipartimento di Internistica Clinica e Sperimentale “F. Magrassi”, Universita degli Studi della Campania Luigi Vanvitelli, 80131 Naples, Italy Correspondence to: Teresa Troiani, email: teresa.troiani@unicampania.it Keywords: MM151, cetuximab, acquired resistance, epidermal growth factor receptor, metastatic colorectal cancer Received: April 13, 2017 Accepted: June 17, 2017 Published: August 02, 2017 ABSTRACT Purpose: We investigated the effect of triple monoclonal antibody inhibition of EGFR to overcome acquired resistance to first generation of anti-EGFR inhibitors. Experimental design: MM151 is a mixture of three different monoclonal IgG1 antibodies directed toward three different, non-overlapping, epitopes of the EGFR. We performed an in vivo study by using human CRC cell lines (SW48, LIM 1215 and CACO2) which are sensitive to EGFR inhibitors, in order to evaluate the activity of MM151 as compared to standard anti-EGFR mAbs, such as cetuximab, as single agent or in a sequential strategy of combination MM151 with irinotecan (induction therapy) followed by MM151 with a selective MEK1/2 inhibitor (MEKi) (maintenance therapy). Furthermore, the ability of MM151 to overcome acquired resistance to cetuximab has been also evaluated in cetuximab-refractory CRC models. Results: MM151 shown stronger antitumor activity as compared to cetuximab. The maintenance treatment with MM151 plus MEKi resulted the most effective therapeutic modality. In fact, this combination caused an almost complete suppression of tumor growth in SW48, LIM 1215 and CACO2 xenografts model at 30 week. Moreover, in this treatment group, mice with no evidence of tumor were more than double as compared to single agent treated mice. Its superior activity has also been demonstrated, in cetuximab-refractory CRC models. Conclusions: These results provide experimental evidence that more efficient and complete EGFR blockade may determine better antitumor activity and could contribute to prevent and/or overcome acquired resistance to EGFR inhibitors.

Background Antibody-dependent cell-mediated cytotoxicity (ADCC) may mediate antitumour activity of IgG1-isotype monoclonal antibody (mAb), suggesting as potential treatment combination of IgG1-mAbs, anti-epidermal growth factor receptor cetuximab and anti-programmed death-ligand-1 avelumab. Methods We evaluated ADCC induction in lung cancer cells by lactate dehydrogenase (LDH) release assay. Antitumour activity and safety of cetuximab plus avelumab were explored in a single-arm proof-of-concept study in pre-treated non-small cell lung cancer (NSCLC) patients (pt) (Cetuximab-AVElumab-lung, CAVE-Lung). Search for predictive biomarkers of response was done. Results Avelumab plus cetuximab induced ADCC in NSCLC cells in vitro in presence of natural killers (NK) from healthy donors (HD) or NSCLC pt, as effectors. Sixteen relapsed NSCLC pt were treated with avelumab plus cetuximab. Antitumour activity was observed in 6/16 pt, defined by progression free survival (PFS) ≥8 months, with 4 of them still on treatment at data lock time (range, 14–19 months). Of note, 3/6 responders had received as previous line anti-programmed death-1 therapy. In responders, clinical benefit was accompanied by significant increase in LDH release over baseline at the first radiological evaluation (8 weeks) (p=0.01) and by early skin toxicity; while in the 10 non-responders, that had PFS ≤5 months, LDH release tends to reduce. Baseline circulating DNA levels were higher in non-responders compared with responders and HD (p=0.026) and decrease in responders during therapy. Mutations in DNA damage responsive family genes were found in responders. Conclusion Cetuximab and avelumab activates NSCLC pt NK cells. Ex vivo evaluation of ADCC, circulating DNA levels and early skin toxicity may predict response to cetuximab plus avelumab in NSCLC. EUDRACT 2017-004195-58

The availability of targeted drugs has made the assessment of the EGFR mutation and ALK rearrangement critical in choosing the optimal treatment for patients with advanced non-small-cell lung cancer (NSCLC). In May 2013, the Italian Association of Thoracic Oncology (AIOT) organized an International Experts Panel Meeting to review strengths and limitations of the available evidence for the diagnosis and treatment of advanced NSCLC with EGFR or anaplastic lymphoma kinase (ALK) alterations and to discuss implications for clinical practice and future clinical research. All patients with advanced NSCLC, with the exclusion of pure squamous cell carcinoma in former or current smokers, should be tested for EGFR mutations and ALK rearrangements before decisions are made on first-line treatment. First-line treatment of EGFR-mutated cases should be with an EGFR tyrosine kinase inhibitor (TKI). Any available agent (gefitinib, erlotinib, or afatinib) can be used, until further data from comparative studies may better guide TKI selection. As general rule, and when clinically feasible, results of EGFR mutational status should be awaited before starting first-line treatment. Panelists agreed that the use of crizotinib is justified in any line of treatment. Although solid evidence supporting the continuation of EGFR TKIs or crizotinib beyond progression is lacking, in some cases (minimal, asymptomatic progression, or oligoprogression manageable by local therapy), treatment continuation beyond progression could be justified. Experimental strategies to target tumor heterogeneity and to treat patients after failure of EGFR TKIs or crizotinib are considered high-priority areas of research. A number of relevant research priorities were identified to optimize available treatment options.

Mouse models of cancer have consistently been used to qualify new anti-cancer drugs for development of human clinical trials. The most used models are xenografts of human tumors grown subcutaneously in immunodeficient mice such as athymic (nude) or severe combined immune deficient (SCID) mice. However, the number of anti-cancer agents that fail in the clinic far outweighs those considered effective, suggesting that the selection procedure for progression of molecules into the clinic requires improvement. This has provoked considerable skepticism about the value of using such preclinical models. As a result, a shift has occurred towards developing and using spontaneous mouse tumor arising in transgenic and/or knockout mice engineered to recapitulate various genetic alterations thought to be causative of specific types of human cancers. Alternatively, the option has been to improve human tumor xenograft models by using orthotopic transplantation and, therefore, promotion of metastatic spread of the resultant ‘primary’ tumors. Here we review the value and the limitations of xenograft models and their role in developing new anti-cancer treatments.