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PDF file - 5320K, Validation of antibodies for analysis of SphK1 expression on FFPE colorectal cancer tissues

PDF file - 503K, Effects of SphK1 modulation on cetuximab sensitivity in resistant colorectal cancer cell lines

PDF file - 1047K, Effects of the combination cetuximab plus fingolimod on apoptosis of GEO-CR tumor xenografts

PDF file - 2068K, Correlation of SphK1 expression with clinical data in colorectal cancer patients

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PDF file - 483K, Effects of SphK1 inhibition by DMS on cetuximab sensitivity in resistant colorectal cancer cell lines

PDF file - 357K, Sensitivity to cetuximab of a panel of human colorectal cancer cell lines

Purpose: Although the anti–EGF receptor (EGFR) monoclonal antibody cetuximab is an effective strategy in colorectal cancer therapy, its clinical use is limited by intrinsic or acquired resistance. Alterations in the “sphingolipid rheostat”—the balance between the proapoptotic molecule ceramide and the mitogenic factor sphingosine-1-phosphate (S1P)—due to sphingosine kinase 1 (SphK1) overactivation have been involved in resistance to anticancer-targeted agents. Moreover, cross-talks between SphK1 and EGFR-dependent signaling pathways have been described.Experimental design: We investigated SphK1 contribution to cetuximab resistance in colorectal cancer, in preclinical in vitro/in vivo models, and in tumor specimens from patients.Results: SphK1 was found overexpressed and overactivated in colorectal cancer cells with intrinsic or acquired resistance to cetuximab. SphK1 contribution to resistance was supported by the demonstration that SphK1 inhibition by N,N-dimethyl-sphingosine or silencing via siRNA in resistant cells restores sensitivity to cetuximab, whereas exogenous SphK1 overexpression in sensitive cells confers resistance to these agents. Moreover, treatment of resistant cells with fingolimod (FTY720), a S1P receptor (S1PR) antagonist, resulted in resensitization to cetuximab both in vitro and in vivo, with inhibition of tumor growth, interference with signal transduction, induction of cancer cells apoptosis, and prolongation of mice survival. Finally, a correlation between SphK1 expression and cetuximab response was found in colorectal cancer patients. Clin Cancer Res; 19(1); 138–47. ©2012 AACR.

PDF file - 908K, Multivariate analysis correlating the response to cetuximab-based regimens to different factors

Current therapies for chemo-refractory metastatic colorectal cancer (mCRC) have limited efficacy. Rechallenge with epidermal growth factor receptor (EGFR) inhibitors in RAS wild-type (WT) mCRC could be valuable in this setting. In VELO, a randomized two-arm phase 2 trial, anti-EGFR monoclonal antibody panitumumab plus standard-of-care trifluridine/tipiracil (31 patients, arm B) was compared to trifluridine/tipiracil (31 patients, arm A) as third-line therapy (ClinicalTrials.gov Identifier NCT05468892). Primary endpoint, progression-free survival (PFS), was met. Median PFS was 4.0 months in arm B versus 2.5 months in arm A [hazard ratio (HR): 0.48; 95% CI 0.28–0.82; P = 0.007]. Baseline plasma RAS/BRAF WT circulating tumor DNA identified patients obtaining prolonged clinical benefit with panitumumab plus trifluridine/tipiracil as compared to trifluridine/tipiracil with PFS rates at 6 months of 38.5% versus 13% and at 12 months of 15.4% versus 0%, respectively. These findings warrant further development for liquid biopsy-guided anti-EGFR rechallenge combination strategies in chemo-refractory RAS WT mCRC.

129 Background: Rechallenge therapy with epidermal growth factor receptor (EGFR) inhibitors in chemo-refractory metastatic colorectal cancer (mCRC) is an emerging therapeutic approach. Trifluridine-tipiracil is approved for treatment of chemo-refractory mCRC patients. Methods: Chemo-refractory RAS WT mCRC patients, that had a major response (partial or complete response) to first-line chemotherapy plus an anti-EGFR monoclonal antibody and had an-anti-EGFR drug-free interval during second-line therapy of four or more months were randomized in a phase II trial to assess the addition of the anti-EGFR monoclonal antibody panitumumab to trifluridine-tipiracil as third-line rechallenge therapy. The primary endpoint was progression free survival (PFS). Baseline plasma was analyzed for circulating free tumor (ct) DNA by using Biocartis Idylla platform to detect mutations in KRAS, NRAS, BRAF ( V600E) and EGFRextracellular domain ( S492R). In 24 patients with baseline RAS/BRAF wild type (WT) ctDNA, Foundation One liquid CDx (324 gene profiling) was also performed before treatment and at disease progression. Results: 62 patients were treated with trifluridine-tipiracil (31 patients, arm A) or with trifluridine-tipiracil plus panitumumab (31 patients, arm B). As of September 16, 2022, 1 patient in arm A and 2 patients in arm B were on treatment. The primary endpoint was met. Median PFS (mPFS) was 4 months in arm B versus 2.5 months in arm A [hazard ratio (HR): 0.48; 95% CI 0.28-0.82; P = 0.007]. Baseline plasma RAS/BRAF WT ctDNA was found in 23/31 patients in Arm A and in 26/31 patients in Arm B. In this group, mPFS was 4.5 months in Arm B versus 2.6 months in Arm A (HR: 0.48; 95% CI 0.26-0.89; P = 0.019). Disease control (major responses plus stable disease) was higher for patients in Arm B compared to Arm A (81% versus 48%), whereas disease progression was the best response in 19% versus 52% patients, respectively. PFS rates at 6 and 12 months were 38.5% and 15.4% in arm B versus 13% and 0% in arm A. At disease progression, Foundation One liquid CDx detected several mutations within the EGFR pathway, which could correlate with cancer cell resistance to panitumumab. Conclusions: This is the first prospective randomized trial which evaluated anti-EGFR monoclonal antibody (panitumumab) in addition to standard of care (trifluridine-tipiracil) as third-line rechallenge therapy in chemo-refractory RASWT mCRC patients. Baseline liquid biopsy allows selection of RAS/BRAF WT ctDNA patients who could have a relevant clinical benefit. Clinical trial information: NCT05468892 .

Purpose: To assess whether panitumumab is active in patients with cetuximab-refractory metastatic colorectal cancer (mCRC). Patients and Methods: Eligible patients had pretreated RAS (renin–angiotensin system) wild-type mCRC that progressed after cetuximab treatment, after having shown either objective response or stable disease. A minimax two-stage design was applied, with progression-free rate at 2 months as the primary end point. At least 12 of 28 and 21 of 41 successes at the first and second stage, respectively, were required for a positive result. Panitumumab 6 mg/kg was provided every 2 weeks, until progression or unacceptable toxicity. Results: Overall, 52 patients with KRAS (Kirsten rat sarcoma viral oncogene) wild-type disease were enrolled, but 11 were found to have mutated disease after all-RAS retesting. Among 41 eligible patients, median time since diagnosis was 38 months, and 71% experienced an objective response to previous cetuximab. First stage was passed with 12 of 28 patients alive without progression at 2 months. At the second stage, 17 of 41 patients were alive without progression at 2 months. At a median follow-up of 21.8 months, 35 patients experienced disease progression, and 26 died. Median progression-free survival was 2.1 months (95% confidence interval, 1.8-3.6) and median overall survival 6.8 months (95% confidence interval, 4.6-16.6). Most of the patients experienced no adverse reactions; 25% of patients had grade 3 rash. Conclusion: According to our study design, panitumumab was not effective in patients with cetuximab-refractory RAS wild-type mCRC. PACER is a phase 2 study aiming to assess whether panitumumab monotherapy is active in patients with cetuximab-refractory metastatic colorectal cancer. Primary end point was 2-month progression-free rate. According to a minimax two-stage design, 28 and 41 patients were required at the two stages, respectively. Panitumumab was shown to be not active at the second stage. The study failed to demonstrate the experimental hypothesis.

Although no definitive data exist in literature, adjuvant chemotherapy is usually recommended in patients with radically resected stage III rectal cancer treated with neo-adjuvant chemo-radiotherapy. We performed a systematic review of literature with direct and indirect comparisons to assess the role of adjuvant mono- or poli-chemotherapy in radically resected rectal cancer treated with neoadjuvant chemo-radiotherapy. Neither chemotherapy (mono-or poli-chemotherapy) nor polichemotherapy with oxaliplatin-containing regimens seems to improve Overall Survival and Disease-Free Survival in patients with radically resected rectal cancer treated with neoadjuvant chemo-adiotherapy. Neither the entire population of patients radically resected after neoadjuvant chemotherapy, nor high risk patients seem to benefit from adjuvant chemotherapy. Our data seem to suggest the need of review the actual international guidelines that suggest the need of adjuvant chemotherapy at least in high risk rectal cancer treated with surgery and neoadjuvant chemo-radiotherapy.

Background Combination of chemotherapies (fluoropirimidines, oxaliplatin and irinotecan) with biologic drugs (bevacizumab, panitumumab, cetuximab) have improved clinical responses and survival of metastatic colorectal cancer (mCRC). However, patients’ selection thorough the identification of predictive factors still represent a challange. Cetuximab (Erbitux®), a chimeric monoclonal antibody binding to the Epidermal Growth Factor Receptor (EGFR), belongs to the Immunoglobulins (Ig) grade 1 subclass able to elicite both in vitro and in vivo the Antibody-Dependent Cell-mediated Cytotoxicity (ADCC). ADCC is the cytotoxic killing of antibody-coated target cells by immunologic effectors. The effector cells express a receptor for the Fc portion of these antibodies (FcγR); genetic polymorphisms of FcγR modify the binding affinity with the Fc of IgG1. Interestingly, the high-affinity FcγRIIIa V/V is associated with increased ADCC in vitro and in vivo. Thus, ADCC could partially account for cetuximab activity. Methods/design CIFRA is a single arm, open-label, phase II study assessing the activity of cetuximab in combination with irinotecan and fluorouracile in FcγRIIIa V/V patients with KRAS, NRAS, BRAF wild type mCRC. The study is designed with a two-stage Simon model based on a hypothetical higher response rate (+ 10%) of FcγRIIIa V/V patients as compared to previous trials (about 60%) assuming ADCC as one of the possible mechanisms of cetuximab action. The test power is 95%, the alpha value of the I-type error is 5%. With these assumptions the sample for passing the first stage is 14 patients with > 6 responses and the final sample is 34 patients with > 18 responses to draw positive conclusions. Secondary objectives include toxicity, responses’ duration, progression-free and overall survival. Furthermore, an associated translational study will assess the patients’ cetuximab-mediated ADCC and characterize the tumor microenvironment. Discussion The CIFRA study will determine whether ADCC contributes to cetuximab activity in mCRC patients selected on an innovative immunological screening. Data from the translational study will support results’ interpretation as well as provide new insights in host-tumor interactions and cetuximab activity. Trial registration The CIFRA trial (version 0.0, June 21, 2018) has been registered into the NIH-US National Library of Medicine, ClinicalTrials.gov database with the identifier number (NCT03874062).

There is an unmet need for predictive biomarkers of the clinical benefit of antiangiogenic drugs. The aim of the present study was to prospectively evaluate the value of (18)F-FDG PET/CT performed during and after preoperative chemoradiotherapy with bevacizumab for the prediction of complete pathologic tumor regression and survival in patients with MRI-defined high-risk locally advanced rectal cancer. Methods: Sixty-one patients treated in a nonrandomized phase II study (BRANCH) with concomitant or sequential (4 d before chemoradiotherapy) administration of bevacizumab with preoperative chemoradiotherapy were included. (18)F-FDG PET/CT was performed at baseline, 11 d after the beginning of chemoradiotherapy (early), and before surgery (late). Metabolic changes were compared with pathologic complete tumor regression (TRG1) versus incomplete tumor regression (TRG2–TRG5), progression-free survival, cancer-specific survival, and overall survival. Receiver-operating-characteristic curves were calculated for those (18)F-FDG PET/CT parameters that significantly correlated with TRG1. Results: Early total-lesion glycolysis and its percentage change compared with baseline (ΔTLG-early) could discriminate TRG1 from TRG2–TRG5. Only receiver-operating-characteristic analysis of ΔTLG-early showed an area under the curve greater than 0.7 (0.76), with an optimal cutoff at 59.5% (80% sensitivity, 71.4% specificity), for identifying TRG1. Late metabolic assessment could not discriminate between the 2 groups. After a median follow-up of 98 mo (range, 77–132 mo), metabolic responders (ΔTLG-early ≥ 59.5%) demonstrated a significantly higher 10-y progression-free survival (89.3% vs. 63.6%, P = 0.02) and cancer-specific survival (92.9% vs. 72.6%, P = 0.04) than incomplete metabolic responders. Conclusion: Our results suggest that early metabolic response can act as a surrogate marker of the benefit of antiangiogenic therapy. The findings provide further support for the use of early (18)F-FDG PET/CT evaluation to predict pathologic response and survival in the preoperative treatment of patients with locally advanced rectal cancer. ΔTLG-early showed the best accuracy in predicting tumor regression and may be particularly useful in guiding treatment-modifying decisions during preoperative chemoradiotherapy based on expected response.

3503 Background: Continuous anti-EGFR-based FOLFIRI is a first-line standard of care in pts with RAS/BRAF wt mCRC. The emergence of resistance and treatment-related toxicity limit the efficacy of continuous treatment. Thus, an intermittent strategy could reduce both toxicity and resistance. Methods: This is a prospective, randomized, non-comparative, open-label, multicenter phase II study. Unresectable, previously untreated RAS/BRAF wt mCRC pts, were randomized to a control arm (A) receiving FOLFIRI/PANI continuously until progression or to the experimental arm (B), receiving 8 cycles of the same regimen followed by a treatment free interval. This lasted untill progressive disease, when another treatment period of up to 8 cycles was restarted. This intermittent strategy was continued until progression occurred on treatment. Tumor assessment was always done every 8 weeks in both arms. Pts were stratified for center, ECOG PS (0-1 vs 2), previous adjuvant therapy (yes or no), sidedness (right vs left) and metastatic sites (1 vs ≥ 2). The primary endpoint was the progression-free survival on treatment (PFSOT) at 1 year. Assuming p1=43% PFSOT at 1 year, corresponding to an expected median PFSOT time ≥ 10 months in the experimental arm, and a 5% drop-out rate, a sample size of 68 pts in each arm granted the study a power of 80%, with a type I error of 10% (binomial test) for rejecting the null hypothesis, p0=30%, corresponding to a median PFSOT time of ≤ 7 months. Secondary endpoints were safety, quality of life, OS and response rate (ORR); ctDNA samples were also collected. No formal comparison between the two arms was planned. Results: From May 2018 to June 2021, 137 pts were randomized (69 arm A/68 arm B). Main pts’ characteristics were (arm A/B): males 59/61%; median age 62/66yrs; PS 0 84/72%; right colon 17/15%; previous adjuvant therapy 22/29%; single metastatic site 33/26%. At a median follow-up of 18 months (IQR: 10-26), median PFS OT was 12.6 months (95% CI: 9.0-16.1) in arm A and 17.6 months (95% CI: 7.5-27.8) in arm B with a 1 year PFSOT rate of 51.7% and 61.3%, respectively. ORR (arm A/B) was 64/56%. Median number of FOLFIRI/PANI cycles administered per patient were (arm A/B) 13/12. Main grade 3-4 toxicities were (arm A/B): skin 27/13%, neutropenia 23/22%; diarrhea 13/15%. Conclusions: The primary endpoint of the study was met with the intermittent FOLFIRI-PANI strategy producing a long PFS with a reduced skin toxicity. These data deserve further investigations in a phase III trial. Clinical trial information: NCT04425239.

Rat sarcoma (RAS) oncogenes have intensively been investigated during the last decades. Taking into account all human tumours, Kirsten Rat Sarcoma Viral Oncogene Homolog (KRAS) gene is the most frequently mutated (about 22%) among the three isoforms, followed by Neuroblastoma RAS Viral Oncogene Homolog (NRAS) (8%) and Harvey Rat Sarcoma Viral Oncogene Homolog (HRAS) (3%). In the last years, careful attention has been paid on KRAS and NRAS gene mutations in non-small-cell lung cancer (NSCLC) and colorectal cancer (CRC) patients because of their prognostic and predictive roles. In particular, a large body of literature data has been generated investigating clinical outcomes of targeted treatments in NSCLC and CRC KRAS- and NRAS-mutated patients. The latest evidences are here reviewed, providing also an overview of the real-world RAS mutation testing practice across different Italian laboratories. On this basis, we propose a knowledge-based system, www.rasatlas.com, to support the healthcare personnel in the management of patients featuring RAS gene mutations in the landscape of precision oncology.

Importance Although bevacizumab is a standard of care in combination treatments for metastatic colorectal cancer (mCRC), its clinical benefit has been limited. Objective To determine whether sequential scheduling of bevacizumab administration in combination with chemotherapy improves treatment efficacy in patients with mCRC, in keeping with the tumor vascular normalization hypothesis. Design, Setting, and Participants This open-label, randomized clinical phase 3 trial was conducted from May 8, 2012, to December 9, 2015, at 3 Italian centers. Patients aged 18 to 75 years with unresectable, previously untreated, or single line–treated mCRC were recruited. Follow-up was completed December 31, 2019, and data were analyzed from February 26 to July 24, 2020. Interventions Patients received 12 biweekly cycles of standard oxaliplatin-based regimens (modified FOLFOX-6 [levo–folinic acid, fluorouracil, and oxaliplatin]/modified CAPOX [capecitabine and oxaliplatin]) plus bevacizumab administered either on the same day as chemotherapy (standard arm) or 4 days before chemotherapy (experimental arm). Main Outcomes and Measures The primary end point was the objective response rate (ORR) measured with Response Evaluation Criteria in Solid Tumors, version 1.1. Secondary end points included progression-free survival, overall survival, safety, and quality of life (QOL). Results Overall, 230 patients (136 men [59.1%]; median age, 62.3 [interquartile range, 53.3-67.6] years) were randomly assigned to the standard arm (n = 115) or the experimental arm (n = 115). The median duration of follow-up was 68.3 (95% CI, 61.0-70.0) months. No difference in ORR (57.4% [95% CI, 47.8%-66.6%] in the standard arm and 56.5% [95% CI, 47.0-65.7] in the experimental arm;P = .89) or progression-free survival (10.5 [95% CI, 9.1-12.3] months in the standard arm and 11.7 [95% CI, 9.9-12.9] months in the experimental arm;P = .15) was observed. However, the median overall survival was 29.8 (95% CI, 22.5-41.1) months in the experimental arm compared with 24.1 (95% CI, 18.6-29.8) months in the standard arm (adjusted hazard ratio, 0.73; 95% CI, 0.54-0.99;P = .04). Moreover, the experimental arm was associated with a significant reduction in the rate of severe diarrhea (6 [5.3%] vs 19 [16.5%];P = .006) and nausea (2 [1.8%] vs 8 [7.0%];P = .05) and improved physical functioning (mean [SD] change from baseline, 0.65 [1.96] vs −7.41 [2.95] at 24 weeks;P= .02), and constipation scores (mean [SD] change from baseline, −17.2 [3.73] vs −0.62 [4.44];P= .003). Conclusions and Relevance In this randomized clinical trial, sequential administration of bevacizumab plus chemotherapy did not improve ORR, the primary end point. However, the overall survival advantage, fewer adverse effects, and better health-related QOL associated with sequential bevacizumab administration might provide the basis for exploring antiangiogenic combination treatments with innovative perspectives. Trial Registration EudraCT Identifier:2011-004997-27; ClinicalTrials.gov Identifier:NCT01718873

Purpose: To assess the association of RAS mutation status and radiomics-derived data by Contrast Enhanced-Magnetic Resonance Imaging (CE-MRI) in liver metastases. Materials and Methods: 76 patients (36 women and 40 men, 59 years of mean age and 36&ndash, 80 years as range) were included in this retrospective study. Texture metrics and parameters based on lesion morphology were calculated. Per-patient univariate and multivariate analysis were made. Wilcoxon-Mann-Whitney U test, receiver operating characteristic (ROC) analysis, pattern recognition approaches with features selection approaches were considered. Results: Significant results were obtained for texture features while morphological parameters had not significant results to classify RAS mutation. The results showed that using a univariate analysis was not possible to discriminate accurately the RAS mutation status. Instead, considering a multivariate analysis and classification approaches, a KNN exclusively with texture parameters as predictors reached the best results (AUC of 0.84 and an accuracy of 76.9% with 90.0% of sensitivity and 67.8% of specificity on training set and an accuracy of 87.5% with 91.7% of sensitivity and 83.3% of specificity on external validation cohort). Conclusions: Texture parameters derived by CE-MRI and combined using multivariate analysis and patter recognition approaches could allow stratifying the patients according to RAS mutation status.

Background: Previous studies on oxaliplatin and fluoropyrimidines as adjuvant therapy in older patients with stage III colon cancer (CC) produced conflicting results. Patients and methods: We assessed the impact of age on time to tumour recurrence (TTR), disease-free survival (DFS), cancer-specific survival (CSS), and overall survival (OS) in 2360 patients with stage III CC (1667 aged

Background: Colorectal cancer (CRC) is the second most common cause of cancer-specific death in both sexes in Western countries. KRAS mutations occur in about 50% of metastatic CRCs (mCRCs). The prognostic value of specific KRAS mutations still remains unexplored and unclear. Methods: Two hundred and forty KRAS wild-type and 206 KRAS/NRAS mutant consecutive unresectable mCRC patients with PS Eastern Cooperative Oncology Group (ECOG) 0 or 1, aged &lt, 80 years, and with a life expectancy &gt, 3 months entered into this study. DNA was extracted from paraffin-embedded formalin-fixed tumour tissues, and it was sequenced with the Oncomine Solid Tumour DNA kit (Thermo Fisher Scientific, Waltham, MA, USA). Data were analysed using the Torrent Suite Software v5.0 (Thermo Fisher Scientific). The primary outcome was the analysis of the prognostic role of different KRAS mutations in terms of overall survival (OS). Results: There were no significant differences among the most prevalent mutations (p.G12D, p.G12V, p.G13D, p.G12A, p.G12C, and p.G12S) in terms of age (&lt, 65 vs. &ge, 65 years), gender (male vs. female), grading (G1/G2 vs. G3), side of primary tumour (left vs. right), pT, and pN. At the median follow-up of 25.6 months, there were 77 deaths in KRAS-mutated patients and 94 in wild-type patients. Three homogeneous prognostic groups were identified: wild-type patients (group A, median survival: 27.5 months), p.G13D/p.G12A/p.G12V/p.G12D mutants (group B, median survival: 17.3 months), and p.G12C/p.G12S mutants (group C, median survival: 5.0 months, p &lt, 0.0001 according to Log Rank test). Upon multivariate analysis, metastatic involvement and p.G12C/p.G12S KRAS mutation group C (vs. other mutations) emerged as independent prognostic variables for survival. Conclusions: We show that mutant KRAS is a negative prognostic factor and that p.G12C/p.G12S variants present the worst clinical courses. This information suggests a clear difference among KRAS mutations, and it will be useful to test potentiated and/or innovative therapeutic strategies in p.G12C/p.G12S metastatic CRC patients.

Background:Despite effective treatments, metastatic colorectal cancer (mCRC) prognosis is still poor, mostly in RAS-mutated tumors, thus suggesting the need for novel combinatorial therapies. Epigenetic alterations play an important role in initiation and progression of cancers, including CRC. Histone-deacetylase inhibitors (HDACi) have shown activity in combination with chemotherapy in the treatment of solid tumors. Owing to its HDACi activity and its safe use for epileptic disorders, valproic acid (VPA) is a good candidate for anticancer therapy that we have largely explored preclinically translating our findings in currently ongoing clinical studies. We have shown in CRC models that HDACi, including VPA, induces synergistic antitumor effects in combination with fluoropyrimidines. Furthermore, unpublished results from our group demonstrated that VPA induces differentiation and sensitization of CRC stem cells to oxaliplatin. Moreover, preclinical and clinical data suggest that HDACi may prevent/reverse anti-angiogenic resistance.Methods/Design:A randomized, open-label, two-arm, multicenter phase-II study will be performed to explore whether the addition of VPA to first line bevacizumab/oxaliplatin/fluoropyrimidine regimens (mFOLFOX-6/mOXXEL) might improve progression-free survival (PFS) in RAS-mutated mCRC patients. A sample size of 200 patients was calculated under the hypothesis that the addition of VPA to chemotherapy/bevacizumab can improve PFS from 9 to 12 months, with one-sided alpha of 0.20 and a power of 0.80. Secondary endpoints are overall survival, objective response rate, metastases resection rate, toxicity, and quality of life. Moreover, the study will explore several prognostic and predictive biomarkers on blood samples, primary tumors, and on resected metastases.Discussion:The “Revolution” study aims to improve the treatment efficacy of RAS-mutated mCRC through an attractive strategy evaluating the combination of VPA with standard cancer treatment. Correlative studies could identify novel biomarkers and could add new insight in the mechanism of interaction between VPA, fluoropyrimidine, oxaliplatin, and bevacizumab.Trial Registration:EudraCT: 2018-001414-15; ClinicalTrials.gov identifier: NCT04310176

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.

Objectives: To detect RAS mutation in colorectal liver metastasis by Diffusion-Weighted Magnetic Resonance Imaging (DWI-MRI) - and Diffusion Kurtosis imaging (DKI)-derived parameters. Methods: In total, 106 liver metastasis (60 metastases with RAS mutation) in 52 patients were included in this retrospective study. Diffusion and perfusion parameters were derived by DWI (apparent diffusion coefficient (ADC), basal signal (S0), pseudo-diffusion coefficient (DP), perfusion fraction (FP) and tissue diffusivity (DT)) and DKI data (mean of diffusion coefficient (MD) and mean of diffusional Kurtosis (MK)). Wilcoxon&ndash, Mann&ndash, Whitney U tests for non-parametric variables and receiver operating characteristic (ROC) analyses were calculated with area under ROC curve (AUC). Moreover, pattern recognition approaches (linear classifier, support vector machine, k-nearest neighbours, decision tree), with features selection methods and a leave-one-out cross validation approach, were considered. Results: A significant discrimination between the group with RAS mutation and the group without RAS mutation was obtained by the standard deviation value of MK (MK STD), by the mean value of MD, and by that of FP. The best results were reached by MK STD with an AUC of 0.80 (sensitivity of 72%, specificity of 85%, accuracy of 79%) using a cut-off of 203.90 &times, 10&minus, 3, and by the mean value of MD with AUC of 0.80 (sensitivity of 84%, specificity of 73%, accuracy of 77%) using a cut-off of 1694.30 mm2/s &times, 6. Considering all extracted features or the predictors obtained by the features selection method (the mean value of S0, the standard deviation value of MK, FP and of DT), the tested pattern recognition approaches did not determine an increase in diagnostic accuracy to detect RAS mutation (AUC of 0.73 and 0.69, respectively). Conclusions: Diffusion-Weighted imaging and Diffusion Kurtosis imaging could be used to detect the RAS mutation in liver metastasis. The standard deviation value of MK and the mean value of MD were the more accurate parameters in the RAS mutation detection, with an AUC of 0.80.

Objective: In this case report, we describe a huge response of pulmonary lung lesions that resulted in a pneumothorax in a metastatic colorectal cancer patient treated with Regorafenib. Case Report: A 65-year-old woman, diagnosed with lung metastases from colon cancer began in the assumption of Regorafenib after failure of a prior line of chemotherapy. The disease was re-evaluated every two months by CT scan and a responding disease was reported, through the cavitation of all pulmonary nodules. Suddenly, after 9 months of a well-tolerated treatment, the patient showed cough, dyspnea and asthenia. She was admitted in our ward and a sub-massive pneumothorax was diagnosed and treated by thoracic surgeons by positioning a chest tube to get a normal expansion of the collapsed lung. Pneumothorax was solved, but patient discontinued Regorafenib as its positive activity on lung metastases, showing a deep response to treatment, was considered an adverse event that caused such a threatening side effect. Results: The pneumothorax was very probably caused by an excessive cavitation of lung sub-pleural nodules responding to chemotherapy extraordinarily. Conclusions: This case highlights the need to monitor constantly patients with lung metastases receiving regorafenib especially with subpleural localization for the consistent risk of such sequelae.

// Alessandro Ottaiano 1, * , Valerio Scotti 2, * , Chiara De Divitiis 3 , Monica Capozzi 3 , Carmen Romano 3 , Antonino Cassata 3 , Rossana Casaretti 3 , Lucrezia Silvestro 3 , Anna Nappi 3 , Valeria Vicario 3 , Alfonso De Stefano 3 , Salvatore Tafuto 3 , Massimiliano Berretta 4 , Guglielmo Nasti 1 and Antonio Avallone 3 1 Department of Abdominal Oncology, SSD–Innovative Therapies for Abdominal Metastases, Istituto Nazionale Tumori di Napoli G. Pascale IRCCS, National Cancer Institute, 80131, Naples, Italy 2 San Rossore Clinic, Viale delle Cascine, 56122, Pisa, Italy 3 Department of Abdominal Oncology, Experimental Clinical Oncology, Istituto Nazionale Tumori di Napoli G. Pascale IRCCS, National Cancer Institute, 80131, Naples, Italy 4 Department of Medical Oncology, CRO Aviano, National Cancer Institute, 33081, Aviano, Italy * These authors have contributed equally to this work Correspondence to: Guglielmo Nasti, email: g.nasti@istitutotumori.na.it , ale.otto@libero.it Keywords: colorectal cancer; radiation therapy; chemotherapy; metastatic colorectal cancer Received: April 26, 2018 Accepted: June 23, 2018 Published: October 16, 2018 ABSTRACT Background: There are very few clinical or prognostic studies on the role of SRT (Stereotactic Radiation Therapy) in the continuum of care of metastatic colorectal cancer (mCRC) patients. Patients and methods: Patients affected by oligo-mCRC were treated with SRT before or after front-line standard treatments. SRT was delivered according to a risk-adapted protocol. Total body CT (Computed Tomography) scan was done before therapy and every three months thereafter. The radiologic responses to therapy were evaluated by RECIST (Response Evaluation Criteria In Solid Tumors). FDG-PET (FluoroDeoxyGlucose - Positron Emission Tomography) was done before and after SRT; metabolic responses were evaluated by using the EORTC (European Organization for Research and Treatment of Cancer) criteria. The Kaplan-Meier product limit method was applied to graph Overall Survival (OS) and Progression-Free Survival (PFS). Results: Forty-seven patients were included. Twenty-one patients had disease limited to lungs, 9 to lung and liver, 7 only to liver, 10 to multiple sites. The median prescription SRT dose was 60 Gy per organ in 3 fractions (median biological effective dose of 180 Gy). The reduction of delta SUVmax (maximum Standardized Uptake Value) correlated with the local control (p

Prognosis of advanced pancreatic cancer is dismal and the novel targeted therapies, albeit successfully used to treat many advanced tumors, have shown modest results. We performed a meta-analysis in order to quantify the effect size on survival of adding targeted therapy to single agent gemcitabine.Randomized phase III trials comparing gemcitabine mono-therapy versus gemcitabine plus a targeted agent in first-line treatment of advanced pancreatic cancer designed on survival as primary outcome were selected. Search was done through Medline and the registry of the NIH. Keywords used for searching were 'pancreas', 'pancreatic', 'gemcitabine'. Study quality was assessed with MERGE criteria. Findings were depicted in classical Forest plots. Publication bias was evaluated by the construction of funnel plot.Nine studies met the meta-analysis inclusion criteria including 4564 patients. The target therapies were: erlotinib, cetuximab, rigosertib, elpamotide, bevacizumab, aflibercept, axitinib, masitinib and ganitumab. There was no statistically significant heterogeneity among the nine trials (p = 0.77). The hazard ratio (HR) of the pooled analysis was 0.998 (CI 95%: 0.932-1.068). Subgroup meta-analysis was also performed in anti-EGFR and anti-angiogenesis trials: the pooled HR were 0.94 (CI 95%: 0.705-1.175) and 1.055 (CI 95%: 0.913-1.197), respectively.The present meta-analysis does not show significant improvements in survival for targeted drugs in advanced pancreatic cancer. The possible reason of these results could be linked to the biology of pancreatic cancer as well as to the absence of predictive factors.

The proportion of patients with metastatic colorectal cancer (mCRC) receiving second or further lines of treatment has not been widely studied. To shed light on this issue, we retrospectively analysed the treatments administered for metastatic disease, and investigated prognostic factors after a diagnosis of metastases, in a consecutive cohort of mCRC patients. Three hundred forty-six mCRC patients were enrolled: 173 were stage II or III (metachronous group), and 173 stage IV (synchronous group) at diagnosis. Survival was calculated between the date of metastatic disease and the date of death or last follow-up. Patients with synchronous lesions more frequently had multiple disease sites, peritoneal carcinomatosis and massive liver deposits, whereas significantly more patients with metachronous lesions developed lung metastases as the sole disease site. 97.4% patients received at least one, 62.4% two, 41.9% three and 23.7% four treatment lines. Patients with metachronous metastases more frequently underwent surgery of metastases in first-line treatment (48.5 versus 24.8%), and more of them were progression-free at the time of the analysis (44 versus 34.9%). At univariate analysis, age > 70 years, multiple disease sites and peritoneal carcinomatosis were associated with significantly decreased survival, whereas surgery of metastases and isolated lung metastases predicted better survival. At multivariate analysis, only peritoneal carcinomatosis and surgery of metastases independently affected survival. The percentage of patients who received an active treatment decreased going from first- to fourth-line treatment. However, the proportion of patients who received efficacious treatment in advanced line remained high. Surgery of metastases was the most important prognostic factors.

Background: The intensive study of predictive factors has strongly ameliorated the therapeutic flow-chart of metastatic colorectal cancer (mCRC) by allowing the selection of patients who benefit from specific therapies. For instance, in mRAS (mutated RAS) mCRC patients, anti-EGFR drugs (cetuximab and panitumumab) are not recommended; in this group of patients, the use of anti-angiogenic drugs (bevacizumab and aflibercept) is predominant. However, at progression to standard bevacizumab-based first-line chemotherapy, still to date, there are no studies to guide oncologists in the choice of the best second-line anti-angiogenic drug (bevacizumab beyond progression versus aflibercept). Methods: ARBITRATION is a prospective, observational study assessing efficacy differences between second-line fluorouracil/irinotecan (FOLFIRI)/bevacizumab versus FOLFIRI/aflibercept at progression to fluoropyrimidines, oxaliplatin and bevacizumab in mRAS mCRC patients. A test power of 80%, a median survival of 9 months from second-line treatment start and a hazard ratio of 0.67 between the two schedules were the basis for statistical design. The final sample will be 220 patients (110 per treatment). The significance will be verified with a two-tailed log-rank test with an alpha value of the I-type error of 5%. Time-to-outcome will be described by Kaplan–Meier curves and prognostic factors studied through multivariable analyses based on the Cox model. Secondary objectives include safety, responses’ duration and progression-free survival. A translational research will be conducted to measure several angiogenic proteins in patients’ serum before starting the therapy in order to evidence any angiogenic factor patterns related to outcome. Discussion: We present a large, prospective, observational study aiming to answer two scientific questions: (1) outcome differences between second-line treatments with FOLFIRI/bevacizumab beyond progression versus FOLFIRI/aflibercept in mRAS mCRC patients, (2) angiogenic factors’ patterns that could associate with efficacy and help oncologists to apply best the therapeutic anti-angiogenic strategies. Trial registration: The ARBITRATION trial (version 0.0, 13 April 2020) has been registered into the clinicaltrials.gov registry on 20 May 2020 with identifier NCT04397601.

In 2005, we performed the largest survey on clinical trials of biotherapies for all solid tumors and found indirect evidence of a publication bias: editors of medical journals were more prone to publish positive results independently from the quality of the studies. We collected data from 2003 to 2015 in 487 studies, and the publication bias previously described was not found in the years between 2010 and 2015: this could be related to changes and/or innovations in the guidelines and editorial policies of oncology journals occurred over the last years.

Colorectal cancer (CRC) is the third leading cause of cancer-related deaths worldwide. It has been estimated that more than one-third of patients are diagnosed when CRC has already spread to the lymph nodes. One out of five patients is diagnosed with metastatic CRC. The stage of diagnosis influences treatment outcome and survival. Notwithstanding the recent advances in multidisciplinary management and treatment of CRC, patients are still reluctant to undergo screening tests because of the associated invasiveness and discomfort (e.g., colonoscopy with biopsies). Moreover, the serological markers currently used for diagnosis are not reliable and, even if they were useful to detect disease recurrence after treatment, they are not always detected in patients with CRC (e.g., CEA). Recently, translational research in CRC has produced a wide spectrum of potential biomarkers that could be useful for diagnosis, treatment, and follow-up of these patients. The aim of this review is to provide an overview of the newer noninvasive or minimally invasive biomarkers of CRC. Here, we discuss imaging and biomolecular diagnostics ranging from their potential usefulness to obtain early and less-invasive diagnosis to their potential implementation in the development of a bespoke treatment of CRC.

Colorectal cancer (CRC) is one of the most frequent and deadly malignancies worldwide. Despite the progresses made in diagnosis and treatment, the identification of tumor markers is still a strong clinical need, because current treatments are efficacious only in a subgroup of patients. UbcH10 represents a potential candidate biomarker, whose expression levels could be employed to predict response or resistance to chemotherapy or targeted agents. UbcH10 mRNA and protein expression levels have been evaluated in a large group of CRC patients and correlated with clinico-pathological characteristics, including KRAS mutations. Moreover, the endogenous levels of UbcH10 and its role on cell growth have been evaluated in CRC cells. Finally, to investigate the impact of UbcH10 protein expression on the response to irinotecan, its active metabolite SN-38 and cetuximab treatment, UbcH10 silencing experiments were carried-out on two colon carcinoma cell lines, Caco-2, and DLD1. Overexpression of UbcH10 mRNA and protein was observed in the vast majority of patients analyzed. UbcH10 suppression decreased CRC cell growth rate (at least in part through deregulation of Cyclin B and ERK1) and sensitized them to pharmacological treatments with irinotecan, SN-38 and cetuximab (at least in part through a down-regulation of AKT). Taken together, these findings indicate that UbcH10 expression regulates CRC growth and could play an important role in the personalization of the therapy of CRC patients.

BackgroundAnalysis of K- and N-RAS mutations is mandatory before planning treatment of metastatic colorectal cancer, because only RAS wild-type (WT) patients can benefit from treatment with anti-EGFR monoclonal antibodies (cetuximab and panitumumab).Case reportHere we report the case of a 69-year-old male patient affected by metastatic sigmoid cancer. He underwent left hemicolectomy, and histology diagnosed a well-differentiated, pT4, node-positive adenocarcinoma; KRAS analysis performed with direct sequencing identified a mutation in exon 2 of the KRAS gene (GGT->GTT). After first-line chemotherapy with FOLFOX6 plus bevacizumab, the patient underwent surgical resection of residual liver metastases. Histology showed metastatic deposits from colic adenocarcinoma with extensive coagulative necrosis. Mutational analysis of the KRAS gene was also performed on liver metastases by pyrosequencing assay, and no mutation was identified. Due to the discordant results (GGT->GTT exon 2 KRAS mutation in the primary tumor, and KRAS-WT in the liver metastases), mutational analysis on liver metastasis was repeated using next-generation sequencing and enriching the sample in tumor cells by manual microdissection; the same type of mutation of the primary tumor (GGT->GTT exon 2 KRAS gene) was confirmed.ConclusionsAccurate tissue sampling and adequately sensitive assays are essential to correctly identify colorectal cancer patients who can be treated with an anti-EGFR monoclonal antibody.

In metastatic colorectal cancer (mCRC) patients (pts), treatment strategies integrating liver resection with more effective therapies offer better 5-year survival rates than palliative chemotherapy alone. However, resectability is established only on clinical-morphovolumetric criteria, liver resection is a complex and costly procedure and relapse occurs in almost 2/3 of pts after potentially curative resection. Therefore, prompt identification of pts at higher risk of recurrence is critical to avoid not-beneficial, expensive procedures. Aberrant metabolism is an emerging hallmark of cancer and recent observations suggest that specific metabolic changes can be used to stratify pts for prognosis and drug-response. We evaluated by 600MHz NMR spectroscopy the metabolomics profiling on sera from 30 mCRC pts, enrolled in the Obelics trial (NCT01718873), which investigated different schedules of bevacizumab in combination with oxaliplatin plus fluoropyrimidines regimens, and subdivided on the basis of outcome, in good (R) vs bad responders (NR) according to PFS: 12 months or longer (R, n = 12) and shorter than 12 months (NR, n = 18). We compared the samples of the two mCRC pts groups, collected at response evaluation when resectability was established in case of appropriate tumor reduction and PCA, sPLS-DA and loading plots evidenced metabolites with statistically different levels between the two sub-groups. ROC curves were performed to identify the cutoff levels of these significant metabolites to be correlated with patient survival. In this way we demonstrated that low levels of 3-hydroxybutyrate and of hydroxyproline as well as high levels of histidine correlated with both poor progression free survival (PFS) and overall survival (OS). Notably, either 3-hydroxybutyrate or histidine are better predictor of both PFS and OS compared to pathologic response on resected metatastases. Lipidomics analysis confirmed clear differences between R and NR pts indicating statistically significant increase of lipids in NR pts, with both higher triglycerides and phospholipids correlating with poor PFS and OS. This latter effects, may reflect, at least in part a non specific inflammatory response; indeed a significant increase of pro-inflammatory cytokines was also demonstrated in NR pts sera by cytokinomics using multiplex ELISA approach. Finally, basal serum metabolomics analysis in both NR and R pts demonstrated that on-treatment evaluation is more informative than pre-treatment evaluation to stratify patients for outcome. Overall, these data suggest that NMR-based metabolomics is a potent and affordable method that could play a role in the prediction of mCRC outcome. Citation Format: Alfredo Budillon, Susan Costantini, Angela Sorice, Francesca Capone, Silvia Marchese, Elena Di Gennaro, Carlo Vitagliano, Fabiana Tatangelo, Alfonso De Stefano, Franco Bianco, Paolo Delrio, Francesco Izzo, Antonio Avallone. Outcome prediction of metastatic colorectal cancer patients undergoing liver resection by analyzing serum metabolomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5268.

Pancreatic and biliary tract carcinomas are very chemoresistant. After a first-line treatment with a gemcitabine-based regimen, no second-line scheme is consolidated in clinical practice. The aim of this study was to evaluate the toxicity and the activity of the FOLFIRI regimen as first-line or second-line chemotherapy in patients with pancreatic or biliary tract tumors. Fifty-four patients (30 with pancreatic tumor, nine with gallbladder tumor, and 15 with biliary tract tumor) were treated with FOLFIRI (irinotecan 180 mg/m², day 1; leucovorin 100 mg/m² intravenously, days 1 and 2; 5-fluorouracil 400 mg/m² intravenous bolus, days 1 and 2; and 600 mg/m² in 22 h intravenously, continuous infusion days 1 and 2; every 14 days). Toxicity was recorded at each cycle according to the NCI-CTC V3.0 criteria, the response rate was verified each four cycles according to the RECIST criteria, and the progression-free survival rates as well as the overall survival rates were calculated according to the Kaplan-Meier method. Overall, the toxicity was mild. Grade 3-4 neutropenia occurred in 42.6% of patients. Grade 3-4 gastrointestinal toxicity was rare. FOLFIRI as a first-line treatment produced a response rate of 25%. In the second-line group, 9/21 patients (42.9%) obtained a stable disease as best response. In the entire population, the median progression-free survival rates were 3.1 months [95% confidence interval (CI), 1.9-4.4] and 3.5 months (95% CI, 2.6-4.4), respectively, in the first-line and the second-line cohort of patients. The median overall survival rates were 14.5 months (95% CI, 7.0-22.1) and 6.2 months (95% CI, 5.4-7.0), respectively, in the first-line and the second-line cohort of patients. FOLFIRI is feasible and well tolerated in patients with pancreatic or biliary tract tumors; it has a good activity in first line and mostly in patients with pancreatic cancer.

BACKGROUND Epidermal growth factor receptor (EGFR) mutations are reliably detected by referral laboratories, even if most lung cancer cytology specimens sent to such laboratories contain very few cells. However, EGFR mutations may be distributed heterogeneously within tumors, thereby raising concerns that mutations detected on cytology are not representative of the entire tumor and, thus, are less reliable in predicting response to tyrosine kinase inhibitor (TKI) treatment than mutations detected on histology. To address this issue, the authors reviewed their clinical practice archives and compared the outcome of TKI treatment among patients who were selected by cytology versus patients who were selected by histology. METHODS From July 2010 to July 2012, 364 cytology samples and 318 histology samples were received. Exon 19 deletions and the L858R point mutation in exon 21, detected by fragment assay and TaqMan assay, respectively, were confirmed by direct sequencing; discrepancies were resolved by cloning polymerase chain reaction products. The response rate (RR) and progression-free survival (PFS) at 12 months (range, 3-34 months) were evaluable in 13 EGFR-mutated patients who were selected for treatment by cytology and 13 patients who were selected by histology. RESULTS The mutation rate was similar in histology samples (8.5%) and cytology samples (8.8%). The RR (54%) and PFS (9.2 months) were similar in histologically selected patients and cytologically selected patients (RR, 62%; PFS, 8.6 months; P = .88). The disease control rate (responsive plus stable disease) was 92% in histologically selected patients and 100% in cytologically selected patients. CONCLUSIONS EGFR mutations detected on cytology specimens by a centralized laboratory can predict TKI treatment response equally well as mutations identified on histology samples. Cancer (Cancer Cytopathol) 2013;121:552–560. © 2013 American Cancer Society.