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In preclinical experiments, cyclic fasting-mimicking diets (FMDs) showed broad anticancer effects in combination with chemotherapy. Among different tumor types, triple-negative breast cancer (TNBC) is exquisitely sensitive to FMD. However, the antitumor activity and efficacy of cyclic FMD in TNBC patients remain unclear. Here, we show that a severely calorie-restricted, triweekly, 5-day FMD regimen results in excellent pathologic complete response (pCR) rates (primary endpoint) and long-term clinical outcomes (secondary endpoints) when combined with preoperative chemotherapy in 30 patients with early-stage TNBC enrolled in the phase 2 trial BREAKFAST. Bulk and single-cell RNA sequencing analysis revealed that highly glycolytic cancer cells, myeloid cells, and pericytes from tumors achieving pCR undergo a significant, early downmodulation of pathways related to glycolysis and pyruvate metabolism. Our findings pave the wave for conducting larger clinical trials to investigate the efficacy of cyclic FMD in early-stage TNBC patients and to validate early changes of intratumor glycolysis as a predictor of clinical benefit from nutrient restriction. This study was registered at Clinicaltrials.gov (NCT04248998)., Competing Interests: Declaration of interests The authors declare the following competing interests: G.V.B., speaker/advisory board: Novartis, Eli Lilly, Daichi/Astrazeneca, Roche, MSD, and Seagen; G.P., consulting fees: Roche, Bayer, and Astra Zeneca; C.V., consulting fees/advisory board: Novartis, Pfizer, Eli Lilly, Daiichi Sankyo, and Menarini Stemline; C.V., honoraria as speaker: Novartis, Eli Lilly, Istituto Gentili, Accademia Nazionale di Medicina, MSD, Research grant: Roche; F.d.B., consulting fees/advisory board: Roche, EMD Serono, NMS Nerviano Medical Science, Sanofi, MSD, Novartis, Incyte, BMS, Menarini, Astra Zeneca, Pierre Fabre, Mattioli 1885, MCCann Health, Taiho; F.d.B., IQVIA Speaker Bureau: BMS, Healthcare Research & Pharmacoepidemiology, Merck Group, MSD, Pfizer, Servier, Sanofi, Roche, AMGEN, Incyte, Dephaforum, Seagen, Nadirex, BMS, Ambrosetti, and Itanet; and F.d.B., research grant/funding: Novartis, F.Hoffmann-LaRoche Ltd, BMS, Ignyta Inc., Merck Sharp & Dohme Spa, Kymab, Pfizer, Tesaro, MSD, MedImmune LCC, Exelixis Inc., LOXO Oncology Inc., DAICHI SANKIO Dev. Ltd, Basilea Pharmaceutica International AG, Janssen-Cilag International NV, and Merck KGAA. C.V. and F.d.B. are co-inventors of the FMD regimen used in this study. The Italian patent (no. 102019000009954) has been released and was first deposited on June 24(th), 2019, with an extension granted under PCT WO 2020/261131 on June 24(th), 2020. The European patent (no. 207403399), Canadian patent (no. 2144217), and USA patent are pending., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Purpose: The multi-kinase inhibitor (mKi) regorafenib has demonstrated efficacy in chemorefractory patients with metastatic colorectal cancer (mCRC). However, lack of predictive biomarkers and concerns over significant toxicities hamper the use of regorafenib in clinical practice., Experimental Design: Serial liquid biopsies were obtained at baseline and monthly until disease progression in chemorefractory patients with mCRC treated with regorafenib in a phase II clinical trial (PROSPECT-R n = 40; NCT03010722) and in a multicentric validation cohort (n = 241). Tissue biopsies collected at baseline, after 2 months and at progression in the PROSPECT-R trial were used to establish patient-derived organoids (PDO) and for molecular analyses. MicroRNA profiling was performed on baseline bloods using the NanoString nCounter platform and results were validated by digital-droplet PCR and/or ISH in paired liquid and tissue biopsies. PDOs co-cultures and PDO-xenotransplants were generated for functional analyses., Results: Large-scale microRNA expression analysis in longitudinal matched liquid and tissue biopsies from the PROSPECT-R trial identified MIR652-3p as a biomarker of clinical benefit to regorafenib. These findings were confirmed in an independent validation cohort and in a "control" group of 100 patients treated with lonsurf. Using ex vivo co-culture assays paired with single-cell RNA-sequencing of PDO established pre- and post-treatment, we modeled regorafenib response observed in vivo and in patients, and showed that MIR652-3p controls resistance to regorafenib by impairing regorafenib-induced lethal autophagy and by orchestrating the switch from neo-angiogenesis to vessel co-option., Conclusions: Our results identify MIR652-3p as a potential biomarker and as a driver of cell and non-cell-autonomous mechanisms of resistance to regorafenib., (©2024 American Association for Cancer Research.)

Background: Biliary tract cancers (BTCs) are rare and lethal cancers, with a 5-year survival inferior to 20%(1-3). The only potential curative treatment is surgical resection. However, despite complex surgical procedures that have a remarkable risk of postoperative morbidity and mortality, the 5-year survival rate after radical surgery (R0) is 20-40% and recurrence rates are up to ~ 75%(4-6). Up to ~ 40% of patients relapse within 12 months after resection, and half of these patient will recur systemically(4-6). There is no standard of care for neoadjuvant chemotherapy (NAC) in resectable BTC, but retrospective reports suggest its potential benefit (7, 8)., Methods: PURITY is a no-profit, multicentre, randomized phase II/III trial aimed at evaluating the efficacy of the combination of gemcitabine, cisplatin and nabpaclitaxel (GAP) as neoadjuvant treatment in patients with resectable BTC at high risk for recurrence. Primary objective of this study is to evaluate the efficacy of neoadjuvant GAP followed by surgery as compared to upfront surgery, in terms of 12-month progression-free survival for the phase II part and of progression free survival (PFS) for the phase III study. Key Secondary objectives are event free survival (EFS), relapse-free survival, (RFS), overall survival (OS), R0/R1/R2 resection rate, quality of life (QoL), overall response rate (ORR), resectability. Safety analyses will include toxicity rate and perioperative morbidity and mortality rate. Exploratory studies including Next-Generation Sequencing (NGS) in archival tumor tissues and longitudinal ctDNA analysis are planned to identify potential biomarkers of primary resistance and prognosis., Discussion: Considering the poor prognosis of resected BTC experiencing early tumor recurrence and the negative prognostic impact of R1/R2 resections, PURITY study is based on the rationale that NAC may improve R0 resection rates and ultimately patients' outcomes. Furthermore, NAC should allow early eradication of microscopic distant metastases, undetectable by imaging but already present at the time of diagnosis and avoid mortality and morbidity associated with resection for patients with rapid progression or worsening general condition during neoadjuvant therapy. The randomized PURITY study will evaluate whether patients affected by BTC at high risk from recurrence benefit from a neoadjuvant therapy with GAP regimen as compared to immediate surgery., Trial Registration: PURITY is registered at ClinicalTrials.gov (NCT06037980) and EuCT(2023-503295-25-00)., (© 2024. The Author(s).)

Severe calorie restriction, in the form of cyclic fasting or fasting-mimicking diets (FMDs), boosts the antitumor activity of cytotoxic chemotherapy in mouse models of triple-negative breast cancer (TNBC). This effect is mostly mediated by fasting/FMD-induced reduction of plasma glucose concentration and by a boost in antitumor immunity. However, clinical evidence that cyclic FMD may impact on the outcomes of advanced TNBC (aTNBC) patients is lacking. We compared the overall survival (OS) of 14 aTNBC patients receiving first-line carboplatin-gemcitabine plus cyclic FMD in the context of the NCT03340935 trial with the OS of 76 consecutive aTNBC patients treated with carboplatin-based chemotherapy alone at Fondazione IRCCS Istituto Nazionale dei Tumori. Multivariable Cox regression models were used to adjust the prognostic impact of FMD for other prognostic variables. Patients undergoing cyclic FMD in combination with carboplatin-gemcitabine had better OS when compared to patients receiving chemotherapy alone (median OS 30.3 months, 95% CI 18-NR, vs 17.2 months, 95% CI 15.3-25.1, log-rank P value .041). Multivariable analysis confirmed an association between FMD use and better OS (HR: 0.40; 95% CI: 0.19-0.86; P = .019) also after propensity score-based matching according to patient ECOG PS and the presence of de novo metastatic disease (HR: 0.41; 95% CI: 0.21-0.83; P = .013). Cyclic FMD in combination with first-line chemotherapy may improve clinical outcomes in aTNBC patients. Our study paves the way for conducting phase II trials to investigate if cyclic FMD can increase the antitumor activity/efficacy of chemotherapy or chemoimmunotherapy in patients with early-stage TNBC or aTNBC., (© 2023 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Background: Bone-targeted agents (BTA), such as denosumab (DN) and zoledronic acid (ZA), have historically reduced the risk of skeletal related events in cancer patients with bone metastases (BM), with no improvement in survival outcomes. In the immunotherapy era, BM have been associated with poor prognosis upon immune-checkpoint inhibitors (ICI). Currently, the impact of bone tumor burden on survival upon BTAs in advanced non-small cell lung cancer (aNSCLC) patients treated with ICI remains unknown., Methods: Data from ICI-treated aNSCLC patients with BM (4/2013-5/2022) in one institution were retrospectively collected. BTA-ICI concurrent treatment was defined as BTA administration at any time before or within 90 days from ICI start. High bone tumor burden (HBTB) was defined as ≥ 3 sites of BM. Median OS (mOS) was estimated with Kaplan-Meier. Aikaike's information criterion (AIC) was used to select the best model for data analysis adjusted for clinical variables., Results: Of 134 patients included, 51 (38 %) received BTA. At a mFU of 39.6 months (m), BTA-ICIs concurrent treatment did not significantly impact on mOS [8.3 m (95% CI 3.9-12.8) versus (vs) 6.8 m (95% CI 4.0-9.6) p = 0.36]; these results were confirmed after adjustment for clinical variables selected by AIC. A multivariate model showed a significant interaction between BTA use and HBTB or radiation therapy to BM. In subgroup analyses, only HBTB confirmed to be associated with significantly longer mOS [8.3 m (95% CI 2.4-14.2) vs 3.5 m (95% CI 2.9-4.1), p = 0.003] and mPFS [3.0 m (95% CI 1.6-4.4) vs 1.8 m (95% CI 1.6-2.0) p = 0.001] upon BTA-ICI concurrent treatment, with the most pronounced OS benefit observed for DN-ICI concurrent regimen [15.2 m (95% CI 0.1-30.7) vs 3.5 m (95% CI 2.9-4.1) p = 0.002]., Conclusions: In the immunotherapy era, HBTB can identify patients experiencing survival benefit with BTA, especially with DN-ICI combination. HBTB should be included as a stratification factor in the upcoming trials assessing BTA and ICI combinations in patients with aNSCLC and BM., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: SM declares personal fees from Italfarmaco, Sanofi and MSD outside the submitted work. EZ declares personal fees from AMGEN, outside the submitted work CP declares personal fees from BMS and MSD, outside the submitted work. AP declares personal fees from Astrazeneca, Roche, BMS. GG declares travel accommodation from Roche and personal fees from Italfarmaco, Astra Zeneca, BMS, MSD outside the submitted work. MCG declares personal financial interests with the following organizations: AstraZeneca, MSD International GmbH, BMS, Boehringer Ingelheim Italia S.p.A, Celgene, Eli Lilly, Ignyta, Incyte, Inivata, MedImmune, Novartis, Pfizer, Roche, Takeda; she also declares Institutional financial interests with the following organizations: Eli Lilly, MSD, Pfizer (MISP), AstraZeneca, MSD International GmbH, BMS, Boehringer Ingelheim Italia S.p.A, Celgene, Ignyta, Incyte, Inivata, MedImmune, Novartis, Pfizer, Roche, Takeda, Tiziana, Foundation Medicine; at the end, she has received research funding from the following organizations: AIRC, AIFA, Italian Moh, TRANSCAN. FdB provided consultation, attended advisory boards and/or provided lectures for the following organizations, from whom received honoraria or education grants: Amgen, AstraZeneca, Boehringer-Ingelheim, BMS, Eli Lilly, F. Hoffmann-La Roche, Ignyta, Merck Sharp and Dohme, Merck Serono, Novartis, Pfizer. GLR declares personal fees from MSD, TAKEDA; GSK; AMGEN; PFIZER; SANOFI; JANSEN; Eli Lilly, BMS, Roche, Italfarmaco, Novartis and AstraZeneca, outside the submitted work. DS declares personal fees from AstraZeneca, MSD, Boehringer Ingelheim and BMS, outside the submitted work. RF declares personal fees from MSD and Beigene outside the submitted work. The other authors report no conflict of interest. ., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Purpose: Prognostic tools to estimate the risk of relapse for patients with liver-limited metastatic colorectal cancer (LL-mCRC) undergoing resection with curative intent are needed. Circulating tumor DNA (ctDNA) as a surrogate of postsurgical minimal residual disease is a promising marker in localized CRC. We explored the role of postoperative ctDNA as a marker of minimal residual disease in patients with radically resected LL-mCRC., Materials and Methods: Seventy-six patients with LL-mCRC were retrospectively included. DNA from tumor tissue was sequenced, and one somatic mutation was then assessed by digital droplet polymerase chain reaction in plasma samples collected after surgery to identify the persistence of ctDNA. Relapse-free survival and postresection overall survival were compared between patients with positive vs negative postoperative ctDNA., Results: ctDNA was found in 39 (51%) of 76 patients with LL-mCRC. At a median follow-up of 77 months, 33 of 39 ctDNA-positive patients and 20 of 37 ctDNA-negative patients experienced disease relapse ( P = .008). ctDNA-positive patients reported significantly shorter RFS than ctDNA-negative ones (median RFS 12.7 v 27.4 months hazard ratio, 2.09, P = .008). In the multivariable model including other prognostic covariates, this association was still significant ( P = .046) and a trend toward shorter overall survival among ctDNA-positive patients was reported (hazard ratio, 1.65, P = .183)., Conclusion: The detection of postsurgical ctDNA is an independent negative prognostic marker and identifies patients at high risk of relapse after liver metastases resection., Competing Interests: Paolo MancaPatents, Royalties, Other Intellectual Property: I have a patent for a method for the identification of gene panels optimal for TMB estimation (Inst) Romano DanesiHonoraria: Lilly, EUSA Pharma, Novartis Italy, Seattle Genetics, GENTILI, AstraZeneca, GlaxoSmithKline Gianluca MasiConsulting or Advisory Role: AstraZeneca, Eisai, MSD OncologyPatents, Royalties, Other Intellectual Property: Terumo (Inst) Marzia Del ReConsulting or Advisory Role: Sanofi, Celgene, Janssen-Cilag, Ipsen, Roche Molecular DiagnosticsSpeakers' Bureau: Sanofi, Pfizer, Novartis, Janssen-Cilag, Ipsen, AstraZeneca, Astellas Pharma, Roche, MSD Oncology Filippo PietrantonioHonoraria: Servier, Bayer, AstraZeneca/MedImmune, Lilly, Sanofi, MSD Oncology, AmgenConsulting or Advisory Role: Amgen, Servier, MSD Oncology, organonResearch Funding: Bristol Myers Squibb (Inst), AstraZeneca (Inst) Chiara CremoliniHonoraria: Roche, Amgen, Bayer, Servier, MSD, Merck, Pierre Fabre, OrganonConsulting or Advisory Role: Roche, Bayer, Amgen, MSD, Pierre Fabre, Nordic PharmaSpeakers' Bureau: Servier, Merck, Pierre FabreResearch Funding: Merck, Bayer, Roche, ServierNo other potential conflicts of interest were reported.

Background: Systemic immunosuppression characterizing cancer patients represents a concern regarding the efficacy of anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination, and real-world evidence is needed to define the efficacy and the dynamics of humoral immune response to mRNA-based anti-SARS-CoV-2 vaccines., Methods: We conducted an observational study that included patients with solid tumors who were candidates for mRNA anti-SARS-CoV-2 vaccination at the Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy. The primary objective was to monitor the immunologic response to the mRNA anti-SARS-CoV-2 vaccination in terms of anti-spike antibody levels. All the patients received two doses of the mRNA-1273 vaccine or the BNT162b2 vaccine. Healthcare workers served as a control group of healthy subjects., Results: Among the 243 patients included in the present analysis, 208 (85.60%) and 238 (97.94%) resulted seroconverted after the first and the second dose of vaccine, respectively. Only five patients (2.06%) had a negative titer after the second dose. No significant differences in the rate of seroconversion after two vaccine doses were observed in patients as compared with the control group of healthy subjects. Age and anticancer treatment class had an independent impact on the antibody titer after the second dose of vaccination. In a subgroup of 171 patients with available data about the third timepoint, patients receiving immunotherapy with immune checkpoint inhibitors seem to have a higher peak of antibodies soon after the second dose (3 weeks after), but a more pronounced decrease at a late timepoint (3 months after)., Conclusions: The systemic immunosuppression characterizing cancer patients did not seem to dramatically affect the humoral response to anti-SARS-CoV-2 mRNA vaccines in our population of patients with solid tumors. Further investigation is needed to dissect the interplay between immunotherapy and longitudinal dynamics of humoral response to mRNA vaccines, as well as to analyze the cellular response to mRNA vaccines in cancer patients., Competing Interests: Conflict of interest statement: The authors declare that there is no conflict of interest., (© The Author(s), 2022.)

Purpose: In GI cancers, anaplastic lymphoma kinase ( ALK ) rearrangements are extremely less frequent than in non-small-cell lung cancer but may be important to offer personalized strategies of treatment in selected patients. Data about the activity and efficacy of ALK inhibitors (ALKi) in GI cancers are scarce., Materials and Methods: We assembled a clinical and molecular international data set of pretreated patients with metastatic or nonresectable cancers of GI primary tumor origin with documented ALK rearrangement treated with at least one line of ALKi. Measurable disease as per RECIST 1.1 was required for response analysis., Results: Primary tumor sites were distributed as follows: 5 (38%) pancreas, 3 (23%) right colon, and 1 (8%) for each one of gastric, duodenal, rectal, left colon, and biliary tract sites. Seven patients (54%) were treated with alectinib, 5 (38%) with crizotinib, and 1 (8%) with entrectinib. After disease progression, five patients (38%) received a subsequent ALKi treatment line, and at the time of data cutoff date, treatment was still ongoing in two patients. Five of 12 evaluable patients (41%) achieved a partial response to first-line ALKi, five patients (41%) had stable disease, and 2 (17%) had progressive disease. No complete responses were registered. At a median follow-up of 39.6 months (interquartile range: 19.8-59.5), the median progression-free survival was 5.0 months (95% CI, 3.68 to no response) and the median overall survival was 9.3 months (95% CI, 5.46 to no response)., Conclusion: Treatment with ALKi provides remarkable responses and clinical benefit in pretreated patients with ALK fusion-positive GI malignancies. Despite the rarity, ALK rearrangements represent an important therapeutic target in individual pretreated patients with GI solid tumors. Further work providing prospective clinical validation of this target is needed., Competing Interests: Marzia Del ReConsulting or Advisory Role: Sanofi, Celgene, Janssen-Cilag, Ipsen, Roche Molecular DiagnosticsSpeakers' Bureau: Sanofi, Pfizer, Novartis, Janssen-Cilag, Ipsen, AstraZeneca, Astellas Pharma, Roche, MSD Oncology Paolo MancaPatents, Royalties, Other Intellectual Property: I have a patent for a method for the identification of gene panels optimal for TMB estimation (Inst) Andrew HendifarConsulting or Advisory Role: Novartis, Ipsen, Perthera, Celgene, AbbVie, EisaiResearch Funding: Ipsen, NGM Biopharmaceuticals (Inst)Travel, Accommodations, Expenses: HalozymeOther Relationship: FibroGen Alexander DrilonHonoraria: Pfizer, Loxo/Bayer/Lilly, IASLCConsulting or Advisory Role: Ignyta, Loxo, AstraZeneca, Pfizer, Blueprint Medicines, Genentech/Roche, BeiGene, Hengrui Therapeutics, Exelixis, Bayer, Tyra Biosciences, Takeda/Millennium, BerGenBio, MORE Health, Lilly, AbbVie, 14ner Oncology/Elevation Oncology, Monopteros Therapeutics, Novartis, EMD Serono/Merck, Repare Therapeutics, Melendi, Archer, Nuvalent Inc, Janssen, Amgen, Merus, Axis Pharma, Medscape, Liberum, Med Learning, PeerView, EPG Health, JNCCN/Harborside, Ology, Clinical Care Options, touchIME, Entos, Treeline, Prelude Therapeutics, Applied Pharmaceutical Science IncSpeakers' Bureau: Helsinn Therapeutics, Beigene, Remedica Ltd, TP Therapeutics, Verastem, Ignyta/Genentech/Roche, AstraZeneca, Liberum, Loxo/Bayer/Lilly, Lungevity, NIH, PER, OncLive/MJH Life Sciences, Clinical Care Options/NCCN, Lung Cancer Canada, AIOT, Chugai Pharma, Chugai Pharma, Sirio Libanes Hospital, Answers in CME, Faculty RTP, RV MoreResearch Funding: Foundation MedicinePatents, Royalties, Other Intellectual Property: Wolters Kluwer (Royalties for Pocket Oncology)Other Relationship: Merck, GlaxoSmithKline, Teva, Taiho Pharmaceutical, Pfizer, PharmaMar, Puma Biotechnology, Merus, Boehringer Ingelheim Guilherme HaradaSpeakers' Bureau: MSD, AstraZeneca, Pfizer Anne Hansen ReeHonoraria: MSD Oncology, BMSiResearch Funding: BMSi (Inst) Samuel KlempnerStock and Other Ownership Interests: TP Therapeutics, Nuvalent IncHonoraria: NateraConsulting or Advisory Role: Lilly, Astellas Pharma, Bristol Myers Squibb, Pieris Pharmaceuticals, Merck, Daiichi Sankyo/UCB Japan, Sanofi/Aventis, MersanaResearch Funding: Leap Therapeutics (Inst), BeiGene (Inst), Silverback Therapeutics (Inst)Other Relationship: NCCN Gunhild Mari MælandsmoPatents, Royalties, Other Intellectual Property: Patent application submitted for a nine-protein/gene panel predicting response to anti VEGF therapies in combination with chemotherapy (Inst) Hege G. RussnesHonoraria: AstraZeneca (Inst), Pfizer (Inst), InCyte (Inst), Merck (Inst), Roche (Inst)Research Funding: Foundation Medicine (Inst) James M. ClearyHonoraria: Blueprint Medicines, Syros PharmaceuticalsConsulting or Advisory Role: Bristol Myers SquibbResearch Funding: Merck, Tesaro, AstraZeneca, Esperas Pharma, AbbVie (Inst), Merus (Inst), Roche/Genentech (Inst), BMS (Inst)Travel, Accommodations, Expenses: Roche, Agios, Bristol Myers Squibb Andrea Sartore-BianchiConsulting or Advisory Role: Amgen, Bayer, Sanofi, Servier, Novartis Maria Di BartolomeoHonoraria: Lilly, MSD Oncology, Servier, BMSiConsulting or Advisory Role: Lilly, MSD OncologyResearch Funding: LillyTravel, Accommodations, Expenses: Roche, Sanofi Filippo PietrantonioHonoraria: Servier, Bayer, AstraZeneca/MedImmune, Lilly, Sanofi, MSD Oncology, AmgenConsulting or Advisory Role: Amgen, Servier, MSD Oncology, MerckResearch Funding: Bristol Myers Squibb (Inst), Astrazeneca (Inst)No other potential conflicts of interest were reported.

Introduction: In patients with metastatic colorectal cancer (mCRC), baseline circulating tumour DNA (ctDNA) variant allele fraction (VAF) might serve as a surrogate of disease burden and should be evaluated in comparison with CEA and RECIST-defined sum of target lesions., Methods: In this pre-planned analysis of the VALENTINO trial, we included patients with RAS wild-type mCRC receiving upfront FOLFOX/panitumumab with available baseline liquid biopsy. CtDNA was analysed by means of a 14-gene NGS panel. For each patient, the gene with the highest VAF in ctDNA was selected., Results: The final cohort included 135 patients. The median VAF was 12.6% (IQR: 2.0-45.2%). Higher VAF was observed in patients with liver metastases and with synchronous metastases presentation. Patients with high VAF had poorer median OS compared to those with low VAF (21.8 vs 36.5 months; HR: 1.82, 95%CI: 1.20-2.76; p = 0.005). VAF outperformed baseline CEA and target lesion diameter in the prognostic stratification and remained significantly correlated with OS (p = 0.003) in a multivariate model. VAF was not significantly correlated with dimensional response and PFS., Conclusion: CtDNA measured by VAF is prognostic in patients with RAS wild-type mCRC. Response and PFS after an anti-EGFR-based first-line strategy are independent from initial tumour burden., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

Background: Immune-checkpoint inhibitors (ICIs) have significantly improved outcome of advanced non-small cell lung cancer (aNSCLC) patients. However, their efficacy remains uncertain in uncommon histologies (UH)., Materials and Methods: Data from ICI treated aNSCLC patients (April,2013-January,2021) in one Institution were retrospectively collected. Univariate and multivariate survival analyses were estimated by Kaplan-Meier and Cox proportional hazards regression model, respectively. Objective response rate (ORR) and disease control rate (DCR) were assessed., Results: Of 375 patients, 79 (21.1%) had UH: 19 (24.1%) sarcomatoid carcinoma, 15 (19.0%) mucinous adenocarcinoma, 10 (12.6%) enteric adenocarcinoma, 8 (10.1%) adenocarcinoma not otherwise specified, 7 (8.9%) large-cell neuroendocrine carcinoma, 6 (7.6%) mixed histology non-adenosquamous, 5 (6.3%) adenosquamous carcinoma, 9 (11.4%) other UH. In UH group, programmed death-ligand 1 (PD-L1) <1%, 1-49%, ≥50% and unknown expression were reported in 27.8%, 22.8%, 31.7% and 17.7% patients respectively and ICI was the second/further-line in the majority of patients. After a median follow-up of 35.64 months (m), median progression-free survival (mPFS) was 2.5 m in UH [95% CI 2.2-2.9 m] versus (vs.) 2.7 m in CH [95% CI 2.3-3.2 m, P-value = .584]; median overall survival (mOS) was 8.8 m [95% CI 4.9-12.6 m] vs. 9.7 m [95% CI 8.0-11.3 m, P-value = .653]. At multivariate analyses only ECOG PS was a confirmed prognostic factor in UH. ORR and DCR were 25.3% and 40.5% in UH vs. 21.6% and 49.5% in CH [P-value = .493 and .155 respectively]., Conclusions: No significant differences were detected between UH and CH groups. Prospective trials are needed to understand ICIs role in UH population., (Copyright © 2021. Published by Elsevier Inc.)

Background: Cabozantinib strongly inhibits osteoclast differentiation and bone resorption in vitro. We aimed to evaluate its effect on bone turnover markers (BTMs) in metastatic renal cell carcinoma., Methods: This is a monocentric prospective study on patients with mRCC treated with cabozantinib between October 2016 and July 2018. We collected blood samples at baseline and after 3 and 6 months of treatment. We compared sets of data obtained from plasma samples in the whole population with unpaired 2-tailed Student t tests and data for a subset of patients for which all timepoints were available with paired 2-tailed Student t tests. We used the Kaplan-Meier method for survival analyses and the log-rank test to compare the curves., Results: Our analysis included 39 patients. At month 3, the mean C-terminal cross-linked telopeptides of type I collagen (CTx) and the mean N-terminal propeptide of type 1 collagen (PINP) levels were significantly decreased in the whole population ( p = 0.013 and p < 0.0001, respectively), as well as at paired analysis ( p = 0.015 and p = 0.045, respectively). No differences were observed between baseline and 6 months ( p = 0.053 and p = 0.087, respectively). After 3 months, the mean parathyroid hormone (PTH) levels significantly increased in the whole population ( p = 0.004), as well as at paired analysis; the mean PTH levels increased significantly at 3 and 6 months, respectively ( p = 0.019 and p = 0.041, respectively). Changes in BTM levels were not associated with outcome., Conclusions: Cabozantinib significantly reduced bone resorption as demonstrated by the decrease of CTx and showed a transient secondary increase of PTH.

Purpose: The routine use of liquid biopsy is not recommended for the choice of initial treatment for patients with metastatic colorectal cancer (mCRC)., Experimental Design: We included patients with left-sided, RAS/BRAF wild-type, HER2-negative, and microsatellite stable mCRC, treated with upfront panitumumab/FOLFOX-4 in the Valentino study. We performed amplicon-based genomic profiling of 14 genes in baseline plasma samples and compared these data with tumor tissue ultra-deep sequencing results. Specific gene mutations in circulating tumor DNA (ctDNA) and their clonality were associated with progression-free survival (PFS), overall survival (OS), and radiological dynamics., Results: Ten and 15 of 120 patients had a mutation of RAS and PIK3CA in ctDNA, with a positive concordance with tissue deep sequencing of only 31.3% and 47.1%, respectively. Presence of RAS or PIK3CA mutations in baseline ctDNA was associated with worse median PFS [8 vs. 12.8 months; HR, 2.49; 95% confidence interval (CI), 1.28-4.81; P = 0.007 and 8.5 vs. 12.9 months; HR, 2.86; 95% CI, 1.63-5.04; P < 0.001] and median OS (17.1 vs. 36.5 months; HR, 2.26; 95% CI, 1.03-4.96; P = 0.042 and 21.1 vs. 38.9 months; HR, 2.18; 95% CI, 1.16-4.07; P = 0.015). RAS mutations in ctDNA were associated with worse RECIST response, early tumor shrinkage, and depth of response, while PIK3CA mutations were not. Patients with higher levels of RAS/PIK3CA variant allele fraction (VAF) in ctDNA had the worst outcomes (VAF ≥ 5% vs. all wild-type: median PFS, 7.7 vs. 13.1 months; HR, 4.02; 95% CI, 2.03-7.95; P < 0.001 and median OS, 18.8 vs. 38.9 months; HR, 4.07; 95% CI, 2.04-8.12; P < 0.001)., Conclusions: Baseline ctDNA profiling may add value to tumor tissue testing to refine the molecular hyperselection of patients with mCRC for upfront anti-EGFR-based strategies., (©2021 American Association for Cancer Research.)

Objectives: Despite the initial clinical benefit, resistance to antiangiogenic therapies develops through the activation of alternative pathways. We measured plasma levels of circulating angiogenic factors to explore their predictive role in metastatic renal cell carcinoma (mRCC) patients treated with pazopanib., Materials and Methods: mRCC patients receiving first-line pazopanib were prospectively enrolled. The levels of circulating interleuchine (IL)-6, IL-8, stromal derived factor-1, vascular endothelial growth factor-A, hepatocyte growth factor (HGF), osteopontin, and E-selectin were quantified at baseline and every 4 weeks until disease progression (PD). Patients were dichotomized into "low" and "high" subgroups by a cutoff point defined by the respective median circulating angiogenic factor (CAF) value at baseline. Then, association with the objective response was determined. Changes in CAF levels between baseline and PD were also compared., Results: Among 25 patients included in the final data set, 6 patients were still on treatment. As best response, 12 patients presented a partial response (48%), 9 showed stable disease, and 4 showed PD. The median follow-up was 31.9 months. The median progression-free survival was 14.8 months. Low baseline levels of IL-6, IL-8, HGF, and osteopontin were found to be significantly associated with objective response. In addition, patients with low baseline levels of HGF showed longer progression-free survival and overall survival, whereas patients with low baseline levels of IL-8 showed longer overall survival. Among patients experiencing PD, the median plasma levels of stromal derived factor-1 and vascular endothelial growth factor-A were significantly higher compared with the baseline (P=0.01; P=0.011). Conversely, the median levels of E-selectin were significantly lower compared with the baseline (P=0.017)., Conclusion: Changes in levels of selected CAFs were associated with response/resistance to pazopanib in mRCC patients.

Enzalutamide and abiraterone have been shown to improve progression-free survival (PFS) and overall survival (OS) in metastatic castration-resistant prostate cancer (mCRPC) patients. Moreover, some patients may not benefit from the inhibition of androgen receptor (AR) activity or, alternatively, may develop secondary resistance. Detection in patients' circulating tumor cells (CTCs) of ARV7, a splicing variant of AR lacking the ligand-binding domain, showed a link with treatment failure. Independent confirmation of the predictive role of CTC status combined with ARV7 detection is, therefore, a priority for extending personalized biomarker-driven treatments to all patients. In this prospective observational study, CTC status and the expression of AR and ARV7 were measured in 37 mCRPC patients, before starting treatment with enzalutamide or abiraterone, by employing commercially available kits. CTC status was positive in 21/37 patients: 46% and 24% of CTC-positive patients were defined as AR- and ARV7-positive, respectively. Kaplan-Meier estimates showed that positivity for each variable was significantly associated with poorer radiological PFS, PSA-PFS, and OS. All considered treatment outcomes worsened when going from CTC-negative to CTC-positive/ARV7-negative to CTC-positive/ARV7-positive patients, both in the global case series and in patients stratified into three groups based on basal PSA levels. Presently, technical approaches appear to be mature for introducing CTC/ARV7 tests in clinical practice.

Background: Oral vinorelbine administered at the maximum tolerated dose has already showed activity and a good safety profile in advanced non-small-cell lung cancer (NSCLC). The MA.NI.LA study was a phase II, multicenter, randomized, controlled trial that aimed to assess the effects of a 'switched maintenance' regimen with oral metronomic vinorelbine (OMV) in patients with NSCLC who had not progressed after first-line platinum-based chemotherapy., Patients and Methods: Patients were randomly assigned in a 1:1 ratio to either OMV (50 mg three-times weekly) as maintenance treatment or best supportive care (BSC). The primary endpoint was progression-free survival (PFS). Secondary endpoints included overall survival (OS), objective disease control rate (DCR, CR + PR + SD), safety and quality of life., Results: In total, 61 and 59 patients were assigned to OMV and BSC, respectively. At a median follow-up of 23.9 (IQR 10.2-38.2) months, patients treated with OMV reported a significantly lower progression rate compared to patient in the BSC arm (89% [54/61] vs 96% [56/58]; HR 0.73; 90% CI 0.53-0.999, p = 0.049). Median PFS for patients treated with vinorelbine was 4.3 months (95% CI 2.8-5.6) vs 2.8 months (95% CI 1.9-4.5) for patients receiving BSC. This benefit was specifically evident in patients aged ≥70 years, in current smokers, and in those who reported disease stabilization as best response to induction chemotherapy. OS and response rate and quality of life were similar in the two arms. Drop-out rate for major toxicity with OMV was unexpectedly high (25%, 14/61) mainly due to grade 3-4 neutropenia (11%, 7/61). Conclusions In patients with unselected NSCLC achieving disease control after platinum-based chemotherapy switch maintenance therapy with OMV prolonged PFS compared to BSC; however, the optimal dose of OMV requires further investigation., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Purpose: Immune-checkpoint inhibitors (ICI) have improved the survival of patients with non-small cell lung cancer (NSCLC). However, only a subset of patients benefit from ICIs, and the role of PD-L1 as predictive biomarker is still debated. A plasma immune-related miRNA-signature classifier (MSC) was established in lung cancer screening settings to identify the lethal form of the disease in early stages. In this exploratory study, we tested the efficacy of the MSC as prognostic marker in patients with advanced NSCLC treated with ICIs., Experimental Design: The MSC risk level was prospectively assessed in a consecutive series of 140 patients with NSCLC before starting treatment with ICIs. Overall response rate (ORR), progression-free survival (PFS), and overall survival (OS) in strata of PD-L1 and MSC alone or combined were considered as endpoints. Multiple plasma samples to monitor MSC risk level during treatment were also profiled., Results: Adequate tissue and plasma samples were available from 111 (79%) and 104 (75%) patients with NSCLC, respectively. MSC risk level was associated with ORR ( P = 0.0009), PFS [multivariate HR = 0.31; 95% confidence interval (CI), 0.17-0.56; P = 0.0001], and OS (multivariate HR = 0.33; 95% CI, 0.18-0.59; P = 0.0002). The combination of MSC and PD-L1 stratified patients into three risk groups having 39%-18%-0% 1-year PFS ( P < 0.0001) and 88%-44%-0% 1-year OS ( P < 0.0001), according to the presence of 2-1-0 favorable markers. During treatment, MSC risk level decreased or remained low until tumor progression in patients with responsive or stable disease., Conclusions: The plasma MSC test could supplement PD-L1 tumor expression to identify a subgroup of patients with advanced lung cancer with worse ORR, PFS, and OS in immunotherapy regimens., (©2019 American Association for Cancer Research.)

Background: An increase in naturally-occurring porphyrins has been described in the blood of subjects bearing different kinds of tumors, including colorectal, and this is probably related to a systemic alteration of heme metabolism induced by tumor cells. The aim of our study was to develop an artificial neural network (ANN) classifier for early detection of colorectal adenocarcinoma based on plasma porphyrin accumulation and risk factors., Methods: We measured the endogenous fluorescence of blood plasma in 100 colorectal adenocarcinoma patients and 112 controls using a conventional spectrofluorometer. Height, weight, personal and family medical history, use of alcohol, red meat, vegetables and tobacco were all recorded. An ANN model was built up from demographic data and from the integral of the fluorescence emission peak in the range 610-650 nm. We used the Receiver Operating Characteristic (ROC) curve to assess performance in distinguishing colorectal adenocarcinoma patients and controls. A liquid chromatography-high resolution mass spectrometry (LC-HRMS) analytical method was employed to identify the agents responsible for native fluorescence., Results: The fluorescence analysis indicated that the integral of the fluorescence emission peak in the range 610-650 nm was significantly higher in colorectal adenocarcinoma patients than controls (p < 0.0001) and was weakly correlated with the TNM staging (Spearman's rho = 0.224, p = 0.011). LC-HRMS measurements showed that the agents responsible for the fluorescence emission were mainly protoporphyrin-IX (PpIX) and coproporphyrin-I (CpI). The overall accuracy of our ANN model was 88% (87% sensitivity and 90% specificity) with an area under the ROC curve of 0.83., Conclusions: These results confirm that tumor cells accumulate a diagnostic level of endogenous porphyrin compounds and suggest that plasma porphyrin concentrations, indirectly measured through fluorescence analysis, may be useful, together with risk factors, as a clinical decision support tool for the early detection of colorectal adenocarcinoma. Our future efforts will be aimed at examining how plasma porphyrin accumulation correlates with survival and response to therapy.

Background: The differential diagnosis between diffuse malignant peritoneal mesothelioma (DMPM) and other peritoneal surface malignancies (PSM) is still challenging. Serum mesothelin and osteopontin are increasingly used as markers of pleural mesothelioma, but their role in DMPM is unclear. We assessed the diagnostic and prognostic values of mesothelin, osteopontin, CEA, CA19.9, CA125, and CA15.3 in DMPM patients., Methods: Markers were dosed before cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (HIPEC) by enzyme-linked immunosorbent assay (ELISA) in 30 DMPM patients and 14 controls with other PSMs. Receiver-operating characteristics (ROC) curve were plotted. The performance of each marker was assessed by the area under the ROC curve (AUC-ROC)., Results: Mean mesothelin levels were 7.84 ng/dl (SD = 5.14) in DMPM group and 3.00 ng/dl (SD = 1.25) in controls (P = 0.001). Mean CEA levels were 5.3 ng/dl (SD = 4.7), and 61.96 ng/dl (SD = 112.5) in the two groups (P = 0.008). No statistical difference was seen for osteopontin (P = 0.738), CA19.9 (P = 0.081), CA125 (P = 0.600), and CA15.3 (P = 0.365). AUC-ROC was 0.836 for CA19.9, 0.812 for mesothelin, 0.793 for CEA, and lower for CA125 (0.652), osteopontin (0.531), and CA15.3 (0.481). Using diagnostic cut-offs selected by ROC methodology, sensitivity, specificity, positive and negative predictive values were 70.0%, 100.0%, 100.0%, and 60.9% for mesothelin >5.21 ng/dl, and 90.0%, 85.7%, 93.1%, and 80.0% for CA19.9 < 8.8 U/dl. At multivariate analysis, osteopontin correlated with survival (hazard rate 6.46; 95%CI 1.81-23.05; P = 0.004)., Conclusion: When assessing PSMs of unknown origin, elevated mesothelin with low CA19.9 may increase the suspicion index for DMPM. Ospeopontin warrants further investigations as a prognostic marker for DMPM., (Copyright © 2018 Elsevier Ltd, BASO ~ The Association for Cancer Surgery, and the European Society of Surgical Oncology. All rights reserved.)

Genomic characterization of circulating tumor cells (CTCs) enables the monitoring of tumor progression and of adaption occurring during treatment. CTC molecular characterization represents indeed a precious tool to implement in the clinical practice for better dealing with acquired resistance to systemic treatment and tumor evolution. Unfortunately CTCs are very rare and enrichments from blood samples and subsequent identification of these cells are technically very challenging. We describe here the main steps leading to the development of a technical protocol for visualization, enumeration and recovery of single CTCs exploiting the recently developed DEPArray™platform. Our description of the technical workflow starts with evaluation of pre-analytical aspects related to blood sample collection warning about the possible effects on immunoreactivity profiles which may bias the interpretation. Subsequently, other CTC-enrichment approaches are critically discussed and compared in relation to their performances with the DEPArray™. Identification of CTCs represents another critical point due to their heterogeneity and due to the still-to-be clarified role of different subpopulations, typically epithelial, mesenchymal or mixed. Finally, issues related to single cell analysis are illustrated. The chapter ends with an overview of results obtained on real clinical samples which support the reliability of the protocol and its transferability to the daily clinical routine.

Unrestrained activation of the proteolytic systems in anastomotic tissue during repair has been implicated in the pathogenesis of anastomotic leakage. We hypothesized that this mechanism may promote an up-regulation of the urokinase-type plasminogen activator system and a spillover of soluble urokinase-type plasminogen activator receptor (suPAR) into blood. In this retrospective analysis patients with anastomotic leakage were compared with a group of matched uncomplicated patients. Anastomotic leakage complicated patients had significantly higher suPAR (p = 0.04) levels until day 3 after surgery. The area under the receiver-operating characteristic (ROC) for suPAR was higher than that CRP (0.874 vs. 0.836). Their analysis suggests the possible use of suPAR as serum marker to characterize the persistent inflammatory response that lead to tissue damage and surgical complication., (© 2016 by the Wound Healing Society.)

Purpose: Obesity is a known risk factor for breast cancer and has been linked to increased risk of recurrence and death in breast cancer patients. Little is known about the predictive value of obesity. As endocrine therapy is widely used for breast cancer treatment worldwide, we aimed at correlating baseline body mass index (BMI) with clinical benefit derived from fulvestrant in postmenopausal women with advanced breast cancer., Methods: We analyzed consecutive patients treated with fulvestrant in our center between January 2009 and March 2015. Patients were categorized as normal (BMI 18.5-24.9 kg/m2), overweight (BMI 25-29 kg/m2) and obese (BMI &gt;30 kg/m2). The antitumor activity of fulvestrant was evaluated in terms of the clinical benefit rate (CBR)., Results: Seventy-five consecutive patients matched the eligibility criteria for analysis. Fulvestrant was administered as first-line therapy in 4 (5%) cases, as second line in 27 (36%) and as third line and beyond in 44 (59%) cases. According to BMI, 44 (59%) patients were classified as normal weight, 19 (25%) as overweight, and 12 (16%) as obese. No difference in estrogen receptor expression was found in relation to BMI. CBR was 53% overall, but rose to 70.5% in normal-weight patients and dropped to 31.6% and 25% in overweight and obese patients, respectively (p&lt;0.001)., Conclusions: Increased BMI has a negative influence on treatment outcome. Even with the limitation of the relatively small sample size, it appears that patients of normal weight are 2.5-fold more likely to benefit from fulvestrant as overweight and obese patients.

A still relevant number of patients with RAS-BRAF wild-type colorectal cancer (CRC) do not respond to treatment with antiepidermal growth factor receptor (EGFR) monoclonal antibodies cetuximab and panitumumab, suggesting that additional biomarkers to guide patient selection are urgently needed. Circulating tumor cells (CTCs) may represent such a biomarker. In this prospective study, 38 patients with advanced RAS-BRAF-wild-type CRC received third-line therapy with cetuximab-irinotecan or panitumumab. Peripheral blood samples for CTC status determination were collected at baseline, during treatment at early (2-4 weeks) and at later (8-10 weeks) times. CTC enrichment was done with the AdnaTest ColonCancerSelect kit, whereas CTC detection was done with the AdnaTest ColonCancerDetect kit. CTC status positivity was defined according to the kit manufacturer's thresholds. Fifty percent of patients were defined as CTC positive at baseline and the overall RECIST response rate was 26%. CTC baseline status was not associated with treatment response, whereas early CTC status and CTC status changes during treatment were significantly associated with tumor response. Kaplan-Meier analysis showed a significantly shorter progression-free survival (median, 2.0 versus 4.0 months, p = 0.004) and overall survival (4.7 versus11.4, p = 0.039) in patients with early CTC + status compared with CTC - ones. In multivariable analysis including classical prognostic factors, the CTC status changes profile during treatment was an independent predictor of both progression-free survival (p < 0.001) and overall-survival (p = 0.001). CTC status assessed early during treatment with anti-EGFR monoclonal antibodies may predict treatment failure in advance compared to imaging-based tools., (© 2015 UICC.)

Aims: Triplet chemotherapy with fluoropyrimidines, oxaliplatin and irinotecan is a standard therapy for metastatic colorectal cancer (CRC). Single nucleotide polymorphisms (SNPs) in DPYD and UGT1A1 influence fluoropyrimdines and irinotecan adverse events (AEs). Low frequency DPYD variants (c.1905 + 1G > A, c.1679 T > G, c.2846A > T) are validated but more frequent ones (c.496A > G, c.1129-5923C > G and c.1896 T > C) are not. rs895819 T > C polymorphism in hsa-mir-27a is associated with reduced DPD activity. In this study, we evaluated the clinical usefulness of a pharmacogenetic panel for patients receiving triplet combinations., Methods: Germline DNA was available from 64 CRC patients enrolled between 2008 and 2013 in two phase II trials of capecitabine, oxaliplatin and irinotecan plus bevacizumab or cetuximab. SNPs were determined by Real-Time PCR. We evaluated the functional variants in DPYD (rare: c.1905 + 1G > A, c.1679 T > G, c.2846A > T; most common: c.496A > G, c.1129-5923C > G, c.1896 T > C), hsa-mir-27a (rs895819) and UGT1A1 (*28) genes to assess their association with grade 3-4 AEs., Results: None of the patients carried rare DPYD variants. We found DPYD c.496A > G, c.1129-5923C > G, c.1896 T > C in heterozygosity in 19%, 5% and 8%, respectively, homozygous rs895819 in hsa-mir-27a in 9% and homozygous UGT1A1*28 in 8%. Grade 3-4 AEs were observed in 36% patients and were associated with DPYD c.496A > G (odds ratio (OR) 4.93, 95% CI 1.29, 18.87; P = 0.021) and homozygous rs895819 in hsa-mir-27a (OR 11.11, 95% CI 1.21, 102.09; P = 0.020). Carriers of DPYD c.1896 T > C and homozygous UGT1A1*28 showed an OR of 8.42 (95% CI 0.88, 80.56; P = 0.052). Multivariate analysis confirmed an independent value for DPYD c.496A > G and c.1896 T > C., Conclusions: Concomitant assessment of DPYD variants and the UGT1A1*28 allele is a promising strategy needing further validation for dose personalization., (© 2015 The British Pharmacological Society.)

Fluoropyrimidines, the mainstay agents for the treatment of colorectal cancer, alone or as a part of combination therapies, cause severe adverse reactions in about 10%-30% of patients. Dihydropyrimidine dehydrogenase (DPD), a key enzyme in the catabolism of 5-fluorouracil, has been intensively investigated in relation to fluoropyrimidine toxicity, and several DPD gene (DPYD) polymorphisms are associated with decreased enzyme activity and increased risk of fluoropyrimidine-related toxicity. In patients carrying non-functional DPYD variants (c.1905+1G>A, c.1679T>G, c.2846A>T), fluoropyrimidines should be avoided or reduced according to the patients' homozygous or heterozygous status, respectively. For other common DPYD variants (c.496A>G, c.1129-5923C>G, c.1896T>C), conflicting data are reported and their use in clinical practice still needs to be validated. The high frequency of DPYD polymorphism and the lack of large prospective trials may explain differences in studies' results. The epigenetic regulation of DPD expression has been recently investigated to explain the variable activity of the enzyme. DPYD promoter methylation and its regulation by microRNAs may affect the toxicity risk of fluoropyrimidines. The studies we reviewed indicate that pharmacogenetic testing is promising to direct personalised dosing of fluoropyrimidines, although further investigations are needed to establish the role of DPD in severe toxicity in patients treated for colorectal cancer.

The need to identify biomarkers for bevacizumab-based treatment in advanced colorectal cancer is imperative. The aim of this study was to investigate the prognostic role of circulating VEGF, PDGF, SDF-1, osteopontin and CEA in patients randomly assigned to three bevacizumab-based regimens. Plasma samples from 50 patients treated at a single Institution were analysed using the multiplex assay BioPlex™ 2200 (Bio-Rad Laboratories, Inc, Berkeley, CA, USA) at baseline, before first three cycles and subsequently every three cycles until disease progression. Prognostic analyses of baseline values were performed using multivariable Cox models, including disease extension >10 cm or ≤10 cm (measured as the sum of the diameters for all target lesions) as adjustment factor. The association between progression-free and overall survival and biomarkers modulation during treatment was studied using multivariable Cox models, which included summary statistics synthesizing during-treatment modulation together with disease extension. The biomarkers significantly associated with disease extension were baseline CEA (p = 0.012) and SDF-1 (p = 0.030). High values of VEGF and SDF-1 tended to be associated with worse prognosis, especially in terms of overall survival. The negative prognostic trend was more marked for baseline CEA as compared to other biomarkers; increasing values during treatment was significantly related to worse prognosis independently of disease extension (p = 0.007 and 0.016 for progression-free and overall survival, respectively). VEGF is related to bevacizumab pharmacodynamics and is associated to other angiogenic cytokines; some of the proposed biomarkers such as SDF-1 and CEA should be further validated for prognosis assessment and monitoring of bevacizumab-based treatment of advanced colorectal cancer.