Low dose TamOxifen and LifestylE changes for bReast cANcer prevention (TOLERANT study): Study protocol of a randomized phase II biomarker trial in women at increased risk for breast cancer.

BACKGROUND: Breast Cancer (BC) prevention strategies range from lifestyle changes such as increasing physical activity and reducing body weight to preventive drugs like tamoxifen, known to reduce BC incidence in high-risk women. Sex Hormone Binding Globulin (SHBG) is related to BC risk due to its ability to bind circulating estradiol at high affinity and to regulate estradiol action. A study protocol is presented based on the assessment of the effect of different interventions such as tamoxifen at 10 mg every other day (LDT), intermittent caloric restriction (ICR) two days per week, lifestyle intervention (LI, step counter use) and their combination on the modulation of SHBG and several other biomarkers associated to BC. METHODS: A randomized phase II biomarker study will be conducted in 4 Italian centers. Unaffected women aged between 18 and 70 years, carriers of a germline pathogenetic variant (BRCA1, BRCA2, PALB2, or other moderate penetrance genes), or with a >5% BC risk at 10 years (according to the Tyrer-Cuzick or the Breast Cancer Surveillance Consortium Risk models) or with a previous diagnosis of intraepithelial neoplasia will be eligible. A total of 200 participants will be randomized to one of the four arms: LDT; LDT + ICR; LI; LI + ICR. Interventions will span six months, with baseline and follow-up clinic visits and interim phone calls. DISCUSSION: The aim of the study is to verify whether LDT increases circulating SHBG more than LI with or without ICR after 6 months. Secondary objectives include assessing HOMA-index, inflammatory markers, adiponectin/leptin ratio, quality of life (QoL), safety, toxicity, mammographic density, and changes in microbiome composition across groups. The study's innovation lies in its inclusion of diverse BC risk categories and combination of pharmaceutical and behavioral interventions, potentially enhancing intervention efficacy while balancing tamoxifen's side effects on QoL, especially menopausal symptoms. TRIAL REGISTRATION: EuCT number:2023-503994-39-00; Clinical trials.gov NCT06033092.

Drug and biomarker tissue levels in a randomized presurgical trial on exemestane alternative schedules.

The drug's activity at the target tissue could help to define the minimal effective dose to promote cancer preventive therapy. Here we present exemestane and sex hormone concentrations within breast tissue from a pre-surgical study of alternative exemestane schedules. Postmenopausal women candidate for breast surgery for estrogen receptor-positive breast cancer were randomized to exemestane 25 mg once daily (QD), 25 mg three times/week (TIW), or 25 mg per/week (QW) for 4-6 weeks before surgery. Drug and sex hormones were analyzed from homogenized frozen tissue using a QTRAP 6500+ LC-MS/MS System. Tissue drug concentrations were detectable only in the QD arm with higher concentrations in non-malignant tissue. Estradiol was nearly suppressed in all groups in the non-malignant tissue (QD vs TIW p = .364 and QD vs QW p = .693). In contrast, a dose-response trend was observed in cancer tissue. Based on estradiol suppression in non-malignant tissue, lower exemestane schedules should be explored for breast cancer preventive therapy.

Serum Biomarkers to Dynamically Predict the Risk of Cardiovascular Events in Patients under Oncologic Therapy. A Multicenter Observational Study.

Background: Serum biomarkers have been investigated as predictive risk factors for cancer-related cardiovascular (CV) risk, but their analysis is limited to their baseline level rather than their overtime change. Besides historically validated causal factors, inflammatory and oxidative stress (OS) related markers seem to be correlated to CV events but this association needs to be further explored. We conducted an observational study to determine the predictive role of the longitudinal changes of commonly used and OS-related biomarkers during the cancer treatment period. Methods: Patients undergoing anticancer therapies, either aged 75+ years old or younger with an increased CV risk according to European Society of Cardiology guidelines, were enrolled. We assessed the predictive value of biomarkers for the onset of CV events at baseline and during therapy using Cox model, Subpopulation Treatment-Effect Pattern Plot (STEPP) method and repeated measures analysis of longitudinal data. Results: From April 2018 to August 2021, 182 subjects were enrolled, of whom 168 were evaluable. Twenty-eight CV events were recorded after a median follow up of 9.2 months (Interquartile range, IQR: 5.1-14.7). Fibrinogen and troponin levels were independent risk factors for CV events. Specifically, patients with higher than the median levels of fibrinogen and troponin at baseline had higher risk compared with patients with values below the medians, hazard ratio (HR) = 3.95, 95% CI, 1.25-12.45 and HR = 2.48, 0.67-9.25, respectively. STEPP analysis applied to Cox model showed that cumulative event-free survival at 18 and 24 months worsened almost linearly as median values of fibrinogen increased. Repeated measure analysis showed an increase over time of D-Dimer (p-interaction event*time = 0.08), systolic (p = 0.07) and diastolic (p = 0.05) blood pressure and a decrease of left ventricular ejection fraction (p = 0.15) for subjects who experienced a CV event. Conclusions: Higher levels of fibrinogen and troponin at baseline and an increase over time of D-Dimer and blood pressure are associated to a higher risk of CV events in patients undergoing anticancer therapies. The role of OS in fibrinogen increase and the longitudinal monitoring of D-dimer and blood pressure levels should be further assessed.

Prebiotics, probiotics, synbiotics and postbiotics to adolescents in metabolic syndrome.

The prevalence of childhood and adolescent obesity has globally reached alarming dimensions and many adolescents affected by obesity already present one or more obesity-related comorbidities. In recent years, emerging evidence supporting the role of gut microbiota in the pathophysiology of metabolic diseases has been reported and the use of prebiotics, probiotics, synbiotics and postbiotics as a strategy to manipulate gut microbiota has become popular. The aim of this review is to explore the relationship between gut microbiota and metabolic syndrome in adolescents and to discuss the potential use of prebiotics, probiotics, synbiotics and postbiotics for the prevention and treatment of this clinical picture in adolescence. According to the most recent literature, prebiotics, probiotics and synbiotics have no clear effect on MetS, but a possible modulation of anthropometric parameters has been observed after synbiotic supplementation. Only one study has examined the role of postbiotics in alleviating metabolic complications in children with obesity but not in adolescents. More extensive research is needed to support the conclusions drawn so far and to develop effective microbiome-based interventions that may help improving the quality of life of children and adolescents exposed to the increasing prevalence of MetS.

Alternative dosing regimen of exemestane in a randomized presurgical trial: the role of obesity in biomarker modulation.

In a 3-arm presurgical trial, four-six weeks exemestane 25 mg three times/week (TIW) was non-inferior to 25 mg/day (QD) in suppressing circulating estradiol in postmenopausal women with ER-positive breast cancer. Since obesity may decrease exemestane efficacy, we analyzed changes in sex steroids, adipokines, Ki-67, and drug levels in relation to obesity. Postmenopausal women with early-stage ER-positive breast cancer were randomized to either exemestane 25 mg QD (n = 57), 25 mg TIW (n = 57), or 25 mg/week (QW, n = 62) for 4-6 weeks before breast surgery. Serum and tissue pre- and post-treatment biomarkers were stratified by body mass index (BMI)< or ≥30 kg/m2. Post-treatment median exemestane and 17-OH exemestane levels were 5-6 times higher in the QD arm compared to the TIW arm. For obese women, TIW maintained comparable reductions to QD in systemic estradiol levels, although the reduction in estrone was less with the TIW regimen. There was less suppression of SHBG with the TIW versus the QD dose schedule in obese women which should result in less systemic bioavailable estrogens. Metabolically, the effect of the TIW regimen was similar to the QD regimen for obese women in terms of leptin suppression and increase in the adiponectin-leptin ratio. Reduction in tissue Ki-67 was less for obese women on the TIW regimen than QD, although changes were similar for non-obese women. Our findings suggest that TIW exemestane should be explored further for primary cancer prevention in both normal weight and obese cohorts.

Inflammatory and Metabolic Biomarker Assessment in a Randomized Presurgical Trial of Curcumin and Anthocyanin Supplements in Patients with Colorectal Adenomas.

Colorectal cancer prevention is crucial for public health, given its high mortality rates, particularly in young adults. The early detection and treatment of precancerous lesions is key to preventing carcinogenesis progression. Natural compounds like curcumin and anthocyanins show promise in impeding adenomatous polyp progression in preclinical models. We conducted a randomized, double-blind, placebo-controlled, phase II presurgical trial in 35 patients with adenomatous polyps to explore the biological effects of curcumin and anthocyanins on circulating biomarkers of inflammation and metabolism. No significant difference in biomarker changes by treatment arm was observed. However, the network analysis before treatment revealed inverse correlations between adiponectin and BMI and glycemia, as well as direct links between inflammatory biomarkers and leptin and BMI. In addition, a considerable inverse relationship between adiponectin and grade of dysplasia was detected after treatment (corr = -0.45). Finally, a significant increase in IL-6 at the end of treatment in subjects with high-grade dysplasia was also observed (p = 0.02). The combined treatment of anthocyanins and curcumin did not result in the direct modulation of circulating biomarkers of inflammation and metabolism, but revealed a complex modulation of inflammatory and metabolic biomarkers of colon carcinogenesis.

Novel Treatments for Obesity: Implications for Cancer Prevention and Treatment.

It is now established that obesity is related to a higher incidence of cancer during a lifespan. The effective treatment of obesity opens up new perspectives in the treatment of a relevant modifiable cancer risk factor. The present narrative review summarizes the correlations between weight loss in obesity and cancer. The current knowledge between obesity treatment and cancer was explored, highlighting the greatest potential for its use in the treatment of cancer in the clinical setting. Evidence for the effects of obesity therapy on proliferation, apoptosis, and response to chemotherapy is summarized. While more studies, including large, long-term clinical trials, are needed to adequately evaluate the relationship and durability between anti-obesity treatment and cancer, collaboration between oncologists and obesity treatment experts is increasingly important.

Taste and Smell Disorders in Cancer Treatment: Results from an Integrative Rapid Systematic Review.

Taste and smell disorders (TSDs) are common side effects in patients undergoing cancer treatments. Knowing which treatments specifically cause them is crucial to improve patients' quality of life. This review looked at the oncological treatments that cause taste and smell alterations and their time of onset. We performed an integrative rapid review. The PubMed, PROSPERO, and Web of Science databases were searched in November 2022. The article screening and study selection were conducted independently by two reviewers. Data were analyzed narratively. Fourteen studies met the inclusion criteria and were included. A high heterogeneity was detected. Taste disorders ranged between 17 and 86%, while dysosmia ranged between 8 and 45%. Docetaxel, paclitaxel, nab-paclitaxel, capecitabine, cyclophosphamide, epirubicin, anthracyclines, and oral 5-FU analogues were found to be the drugs most frequently associated with TSDs. This review identifies the cancer treatments that mainly lead to taste and smell changes and provides evidence for wider studies, including those focusing on prevention. Further studies are warranted to make conclusive indication possible.

Case report: Variable response to immunotherapy in ovarian cancer: Our experience within the current state of the art.

Despite recent advances in ovarian cancer (OC) treatment, including the introduction of bevacizumab and PARP-inhibitors, OC remains a lethal disease. Other therapeutic options are being explored, such as immunotherapy (IT), which has been proved effective in many solid tumors. Findings about tumor-infiltrating cytotoxic and regulatory T cells, together with the expression of PD-1 on immune cells and of PD-L1 on tumor cells, gave the rationale for an attempt to the use of IT also in OC. We treated two patients with avelumab, an anti-PD-L1 monoclonal antibody, after the first line of chemotherapy: Patient A underwent 19 cycles of maintenance therapy with avelumab with a disease-free interval of 12 months, whereas patient B showed a slight progression of disease after only eight cycles. A higher PD-L1 expression in tumor cells of patient A was detected. She also underwent a genomic assessment that described the presence of a high Tumor Mutational Burden (TMB) and a status of Loss of Heterozygosity (LoH). This different response to the same treatment puts in evidence that some genomic and immune features might be investigated.

Predictors of poor seroconversion and adverse events to SARS-CoV-2 mRNA BNT162b2 vaccine in cancer patients on active treatment.

PURPOSE: Initial findings in patients with cancer suggest a lower seroconversion to SARS-CoV-2 vaccination possibly related to myelo-immunosuppressive therapies. We conducted a prospective study to assess factors predicting poor seroconversion and adverse events following immunisation (AEFI) to the BNT162b2 vaccine in patients on active treatment. PATIENTS AND METHODS: Cancer patients, candidates to two doses of BNT162b2 SARS-CoV-2 vaccination, were enrolled. Patients on active surveillance served as controls. The primary endpoint was poor seroconversion (anti S1/S2 IgG < 25 AU/mL) after 21 days from the second dose. RESULTS: Between March and July 2021, 320 subjects were recruited, and 291 were assessable. The lack of seroconversion at 21 days from the second dose was 1.6% (95% CI, 0.4-8.7) on active surveillance, 13.9% (8.2-21.6) on chemotherapy, 11.4% (5.1-21.3) on hormone therapy, 21.7% (7.5-43.7) on targeted therapy and 4.8% (0.12-23.8) on immune-checkpoint-inhibitors (ICI). Compared to controls, the risk of no IgG response was greater for chemotherapy (p = 0.033), targeted therapy (0.005) and hormonotherapy (p = 0.051). Lymphocyte count < 1 × 109/L (p = 0.04) and older age (p = 0.03) also significantly predicted poor seroconversion. Overall, 43 patients (14.8%) complained of AEFI, mostly of mild grade. Risk of AEFI was greater in females (p = 0.001) and younger patients (p = 0.009). CONCLUSION: Chemotherapy, targeted therapy, hormone therapy, lymphocyte count < 1 × 109/L, and increasing age predict poor seroconversion after two doses of BNT162b2 in up to 20% of patients, indicating the need for a third dose and long-term serological testing in non-responders. AEFI occur much more frequently in women and younger subjects who may benefit from preventive medications. CLINICALTRIALS. GOV IDENTIFIER: NCT04932863.

Patient- versus physician-reported outcomes in a low-dose tamoxifen trial in noninvasive breast cancer.

BACKGROUND: We recently conducted a de-escalation trial of low-dose tamoxifen 5 mg/day ("babytam", BT) or placebo given for 3 years in 500 women with noninvasive breast cancer. Women on babytam had a 52% reduction of recurrence (invasive breast cancer or DCIS) after 5 years. Since menopausal symptoms are major reasons for treatment withdrawal during tamoxifen preventive therapy, we compared and analyzed the patient-reported outcomes (PROs) with the physician-reported adverse events and studied their association with recurrence. METHODS: Menopausal symptoms recorded by physicians using the Common Terminology Criteria (CTCAEs) were compared with a patient self-reported validated questionnaire reviewed by a research nurse at baseline and every 6 months up to 36 months. Hot flashes (HF), the main outcome measure, were detected through a self-report 7-day diary for frequency and intensity. Treatment adherence and efficacy were assessed by the Kaplan-Meier curves and the Cox model. RESULTS: The number of HF events at 12, 24, and 36 months for PROs versus CTCAEs was 246 versus 12, 238 versus 8, and 210 versus 4, respectively. The majority of events were grade 1. There was no difference in PROs between babytam and placebo except for HF daily frequency, which increased by 1.5 events (95% CI, 1.1-1.8) on placebo to 2.1 on babytam (95% CI, 1.7-2.5, p = 0.05). The presence of HF at baseline was a favorable prognostic factor for recurrence and a predictive factor for response to babytam. Adherence was similar between babytam and placebo. CONCLUSIONS: The use of PROs is effective for identifying frequent mild grade menopausal symptoms which are underestimated by physicians but important prognostic and predictive factors. Research nurse can use these results as a tool to reassure patients about symptoms, improve adherence to treatment, and limit dropouts.

Home Se-Cure: A Home Care Service for Cancer Patients during the COVID-19 Pandemic.

Cancer patients are exposed to a greater risk of COVID-19 infection, resulting in treatment delays and unnecessary hospitalizations. International authorities have suggested reducing visits to hospitals and guarantee continuity of care. We developed a home care project called Home Se-Cure (HSC) to guarantee the continuity of oral, intramuscular, and subcutaneous cancer therapy during COVID-19. The Home Se-Cure project included cancer patients living near Galliera Hospital. Patients received home visits by registered nurses (RNs), whoperformed blood tests and delivered cancer therapies. Patients were instructed to take drugs after blood test results and therapy confirmation by oncologists. Sixty-six patients decided to participate and 38 declined the service. A customer satisfaction questionnaire was administered to a subgroup of patients participating in the project. The most prevalent disease in the HSC group was prostate cancer. The mean age of the patients in HSC was 78.4 years and 68.9 in the decliner group. The majority of the HSC participants appreciated the project because they could stay at home (71%) and reduce the risk of COVID-19 contagion (67.7%). Compared to decliners, the time the study group saved was 2033 hours. HSC guaranteed the continuity of care during the COVID-19 pandemic by reducing the number of patients in the hospital and avoiding crowds in the waiting room.

A Presurgical Study of Curcumin Combined with Anthocyanin Supplements in Patients with Colorectal Adenomatous Polyps.

Adenomatous polyps are precancerous lesions associated with a higher risk of colorectal cancer (CRC). Curcumin and anthocyanins have shown promising CRC-preventive activity in preclinical and epidemiological studies. The objective of this window-of-opportunity, proof-of principle trial was to evaluate the effect of curcumin combined with anthocyanin supplements on tissue biomarkers of colorectal adenomatous polyps. Eligible patients received either anthocyanin and curcumin supplementation or related matching placebo for 4-6 weeks before polyp removal. Adenomatous polyps and adjacent tissue biopsies were collected at baseline and after supplementation for immunohistochemical assessment of ß-catenin, NF-kappa B (NF-κB), Ki-67, P53, and dysplasia. No differences were observed in baseline biomarker expression between normal and dysplastic tissues. The combination of anthocyanins and curcumin resulted in a significant borderline reduction of NF-κB immunohistochemistry (IHC) expression in adenoma tissue (geometric mean ratio (GMR): 0.72; 95% confidence interval (CI): 0.51-1.00; p-value: 0.05) and a trend to a reduction of Ki-67 (GMR: 0.73; 95% CI: 0.50-1.08; p-value: 0.11). No significant modulation of biomarkers in normal adjacent mucosa was observed. We concluded that the combined supplementation of anthocyanins and curcumin seems to lead to a potentially favorable modulation of tissue biomarkers of inflammation and proliferation in colon adenomas.

Association of CYP2D6 genotype and tamoxifen metabolites with breast cancer recurrence in a low-dose trial.

Low-dose tamoxifen halves recurrence in non-invasive breast cancer without significant adverse events. Some adjuvant trials with tamoxifen 20 mg/day had shown an association between low endoxifen levels (9-16 nM) and recurrence, but no association with CYP2D6 was shown in the NSABP P1 and P2 prevention trials. We studied the association of CYP2D6 genotype and tamoxifen metabolites with tumor biomarkers and recurrence in a randomized phase III trial of low-dose tamoxifen. Median (IQR) endoxifen levels at year 1 were 8.4 (5.3-11.4) in patients who recurred vs 7.5 (5.1-10.2) in those who did not recur (p = 0.60). Tamoxifen and metabolites significantly decreased C-reactive protein (CRP, p < 0.05), and a CRP increase after 3 years was associated with higher risk of recurrence (HR = 4.37, 95% CI, 1.14-16.73, P = 0.03). In conclusion, endoxifen is below 9 nM in most subjects treated with 5 mg/day despite strong efficacy and there is no association with recurrence, suggesting that the reason for tamoxifen failure is not poor drug metabolism. Trial registration: ClinicalTrials.gov, Identifier: NCT01357772 .

Effect Modifiers of Low-Dose Tamoxifen in a Randomized Trial in Breast Noninvasive Disease.

PURPOSE: Low-dose tamoxifen halved recurrence after surgery in a phase III trial in breast noninvasive disease without increasing adverse events. We explored the effect of low-dose tamoxifen in clinically relevant subgroups, including menopausal status, estradiol levels, smoking, body mass index, and proliferation of baseline lesion. PATIENTS AND METHODS: Incidence of invasive breast cancer or ductal carcinoma in situ was the primary endpoint. HRs and interaction terms were estimated using Cox models. RESULTS: A favorable HR and 95% confidence interval (CI) could be demonstrated for postmenopausal status (HR = 0.30; 95% CI, 0.11-0.82 vs. HR = 0.73; 95% CI, 0.30-1.76 in premenopausal women; P interaction = 0.13), women with estradiol less than 15.8 pg/mL, presence of menopausal symptoms at baseline, and never smoking (P interaction = 0.07), although the interaction P value was >0.05 for all characteristics. Efficacy was similar in all body mass index categories. Tumors with Ki-67 above the median level of 10% had a greater benefit (HR = 0.27; 95% CI, 0.09-0.81) than those with Ki-67 ≤10% (HR = 1.58; 95% CI, 0.45-5.60; P interaction = 0.04). CONCLUSIONS: The efficacy of low-dose tamoxifen seems to be greater in postmenopausal women and in women with lower estradiol levels. Benefits appear to be larger also in women with menopausal symptoms, never smokers, and tumors with Ki-67 >10%. Our results by menopausal status provide important insight into low-dose tamoxifen personalized treatment, although caution is necessary given their exploratory nature. Observation of an improved response in tumors with Ki-67 >10% is consistent but the use of the marker in this setting is investigational.See related commentary by Fabian, p. 3510.

Predictors of poor seroconversion and adverse events to SARS-CoV-2 mRNA BNT162b2 vaccine in cancer patients on active treatment. Role of the Research Nurse.

BACKGROUND: Vaccines have shown 95% protection from COVID-19 disease in healthy populations. Initial findings in cancer patients suggest a lower seroconversion and greater toxicity possibly related to myelo-immunosuppressive therapies. AIM: We conducted a prospective study to assess factors predicting poor seroconversion and adverse events following immunization (AEFI) to the BNT162b2 vaccine in cancer patients on active treatment. METHODS: Blood samples were collected by the research nurse at first dose (visit 1), second dose (visit 2), after 42 days (visit 3) and after 6 months (visit 4). At visit 1, 3 and 4 participants received: Hospital Anxiety and Depression Scale (HADS) and Distress Thermometer. Patients who ended treatment >6 months on active surveillance served as controls. RESULTS: Between March and July 2021, 320 subjects were recruited and 291 were assessable. The lack of seroconversion at 21 days from the second dose was 1.6% (95% CI, 0.4-8.7) on active surveillance, 13.9% (8.2-21.6) on chemotherapy, 11.4% (5.1-21.3) on hormone therapy, 21.7% (7.5-43.7) on targeted therapy and 4.8% (0.12-23.8) on immunotherapy. Compared to controls, the risk of no IgG response was greater for chemotherapy (P=0.033), targeted therapy (0.005) and hormonotherapy (P=0.051). Lymphocyte count less than 1x109/L, older age and advanced stage also significantly predicted poor seroconversion. Overall, 43 patients (14.8%) complained of AEFI, mostly of mild grade. Risk of AEFI was greater in females (P=0.001) and younger patients (P=0.009). CONCLUSIONS: A third booster dose and long-term serological testing is required in subjects who have not responded to the vaccine. NURSING IMPLICATIONS: nurses must take responsibility for promoting and protecting the health of cancer patients.

Estimating the magnitude of clinical benefit of systemic therapy in patients with DCIS or pre-invasive disease of the breast.

The challenge of effective management of ductal carcinoma in situ (DCIS) and other pre-malignant disorders of the breast is to select patients who will not progress to invasive carcinoma from those at the highest risk who require radiotherapy and/or endocrine therapy to minimize the risk of a subsequent invasive recurrence. Although IBIS-II and NSABP-B35 DCIS phase III trials proved that tamoxifen 20 mg/day and anastrozole reduce the risk of ipsilateral and contralateral events, the toxicities of both drugs have hampered the drug uptake by high-risk women. We recently reported results of a 3-year placebo-controlled trial of low-dose (5 mg/d) tamoxifen in 500 women with intraepithelial neoplasia (70% DCIS). At a median follow-up of 5 years, women randomly assigned to low-dose tamoxifen had half the number of subsequent diagnoses of DCIS or invasive cancer compared with those randomly assigned to placebo but no increase in thromboembolic events or endometrial cancers. The 5-year number needed to treat was 22 (95% CI, 20-27). Our attention is now focused on prognostic and predictive markers to identify patients who can derive the greatest benefits from low dose tamoxifen, such as for instance the expression of 23 genes involved in cell cycle progression (CCP). In conclusion, we endorse an active treatment of DCIS as the standard of care.

The ASAMET trial: a randomized, phase II, double-blind, placebo-controlled, multicenter, 2 × 2 factorial biomarker study of tertiary prevention with low-dose aspirin and metformin in stage I-III colorectal cancer patients.

BACKGROUND: Epidemiological studies and cardiovascular prevention trials have shown that low-dose aspirin can reduce colorectal cancer (CRC) incidence and mortality, including inhibition of distant metastases. Metformin has also been associated with decreased colon adenoma recurrence in clinical trials and lower CRC incidence and mortality in epidemiological studies in diabetics. While both drugs have been tested as single agents, their combination has not been tested in cancer prevention trials. METHODS/DESIGN: This is a randomized, placebo-controlled, double-blind, 2 × 2 biomarker trial of aspirin and metformin to test the activity of either agent alone and the potential synergism of their combination on a set of surrogate biomarkers of colorectal carcinogenesis. After surgery, 160 patients with stage I-III CRC are randomly assigned in a four-arm trial to either aspirin (100 mg day), metformin (850 mg bis in die), their combination, or placebo for one year. The primary endpoint biomarker is the change of IHC expression of nuclear factor kappa-B (NFκB) in the unaffected mucosa of proximal and distal colon obtained by multiple biopsies in two paired colonoscopies one year apart. Additional biomarkers will include: 1) the measurement of circulating IL-6, CRP and VEGF; 2) the IHC expression of tissue pS6K, p53, beta-catenin, PI3K; 3) the associations of genetic markers with treatment response as assessed by next generation sequencing of primary tumors; 4) the genomic profile of candidate genes, pathways, and overall genomic patterns in tissue biopsies by genome wide gene expression arrays; and 5) the evaluation of adenoma occurrence at 1 year. DISCUSSION: A favorable biomarker modulation by aspirin and metformin may provide important clues for a subsequent phase III adjuvant trial aimed at preventing second primary cancer, delaying recurrence and improving prognosis in patients with CRC. TRIAL REGISTRATION: EudraCT Number: 2015-004824-77; ClinicalTrial.gov Identifier: NCT03047837 . Registered on February 1, 2017.