Gianmaria Miolo

Prevalence of BRCA1 genomic rearrangements in a large cohort of Italian breast and breast/ovarian cancer families without detectable BRCA1 and BRCA2 point mutations

The presence of genomic rearrangements of the BRCA1 gene in breast and/or ovarian cancer families has been intensively investigated in patients from various countries over the last years. A number of different rearrangements have been reported by several studies that clearly document the involvement of this mutation type in genetic predisposition to breast and ovarian cancer. Population‐specific studies are now needed to evaluate the prevalence of genomic rearrangements before deciding whether to include ad hoc screening procedures into standard diagnostic mutation detection approaches. Indeed, the vast majority of the studies have been performed on small, highly selected, sample sets because of the limitations imposed by the laborious technical approaches. Moreover, prevalence figures are likely to differ across different countries according to the ethnic origin of each specific population. Here we analyze a large cohort of 653 Italian probands, negative for BRCA1 and BRCA2 point mutations, gathered from four National Institutions. We report the identification of BRCA1 genomic rearrangements in 12 independent families. Noteworthy, half of the probands carry mutations that recur in more than one Italian family. Considering the whole spectrum of Italian BRCA1 gene rearrangements identified thus far in consecutive patients, we estimate that alterations of this type account for 19% (95% CI: 0.11 < 0.19 < 0.28) of the BRCA1 mutation positive families. We conclude that the search for major genomic rearrangements is essential for an accurate and comprehensive BRCA1 mutation detection strategy in Italy.

Clinical validity of a DPYD-based pharmacogenetic test to predict severe toxicity to fluoropyrimidines.

Pre‐therapeutic DPYD pharmacogenetic test to prevent fluoropyrimidines (FL)‐related toxicities is not yet common practice in medical oncology. We aimed at investigating the clinical validity of DPYD genetic analysis in a large series of oncological patients. Six hundred three cancer patients, treated with FL, have been retrospectively tested for eight DPYD polymorphisms (DPYD‐rs3918290, DPYD‐rs55886062, DPYD‐rs67376798, DPYD‐rs2297595, DPYD‐rs1801160, DPYD‐rs1801158, DPYD‐rs1801159, DPYD‐rs17376848) for association with Grade ≥3 toxicity, developed within the first three cycles of therapy. DPYD‐rs3918290 and DPYD‐rs67376798 were associated to Grade ≥3 toxicity after bootstrap validation and Bonferroni correction (p = 0.003, p = 0.048). DPYD‐rs55886062 was not significant likely due to its low allelic frequency, nonetheless one out of two heterozygous patients (compound heterozygous with DPYD‐rs3918290) died from toxicity after one cycle. Test specificity for the analysis of DPYD‐rs3918290, DPYD‐rs55886062 and DPYD‐rs67376798 was assessed to 99%. Among the seven patients carrying one variant DPYD‐rs3918290, DPYD‐rs55886062 or DPYD‐rs67376798 allele, not developing Grade ≥3 toxicity, 57% needed a FL dose or schedule modification for moderate chronic toxicity. No other DPYD polymorphism was associated with Grade ≥3 toxicity. Our data demonstrate the clinical validity and specificity of the DPYD‐rs3918290, DPYD‐rs55886062, DPYD‐rs67376798 genotyping test to prevent FL‐related Grade ≥3 toxicity and to preserve treatment compliance, and support its introduction in the clinical practice.

Metabolomics Biomarkers of Frailty in Elderly Breast Cancer Patients

Metabolome analysis has emerged as a powerful technique for detecting and define specific physio‐pathological phenotypes. In this investigation the diagnostic potential of metabolomics has been applied to better characterize the multiple biochemical alterations that concur in the definition of the frailty phenotype observed in elderly breast cancer patients. The study included 89 women with breast cancer (range 70–97 years) classified as Fit (n = 49), Unfit (n = 23), or Frail (n = 17) according to comprehensive geriatric assessment. The serum metabolomic profile was performed by tandem mass spectrometry and included different classes of metabolites such as amino acids, acylcarnitines, sphingo‐, and glycerol‐phospolipids. ANOVA was applied to identify the metabolites differing significantly among Fit, Unfit, and Frail patients. In patients carrying the frail phenotype, the amino acid perturbations involve serine, tryptophan, hydroxyproline, histidine, its derivate 3‐methyl‐hystidine, cystine, and β‐aminoisobutyric acid. With regard to lipid metabolism, the frailty phenotype was characterized by a decrease of a wide number of glycerol‐ and sphingo‐phospholipid metabolites. These metabolomics biomarkers may give a further insight into the biochemical processes involved in the development of frailty in breast cancer patients. Moreover, they might be useful to refine the comprehensive geriatric assessment model. J. Cell. Physiol. 229: 898–902, 2014.

A different immunologic profile characterizes patients with HER-2-overexpressing and HER-2-negative locally advanced breast cancer: implications for immune-based therapies

IntroductionThe clinical efficacy of trastuzumab and taxanes is at least partly related to their ability to mediate or promote antitumor immune responses. On these grounds, a careful analysis of basal immune profile may be capital to dissect the heterogeneity of clinical responses to these drugs in patients with locally advanced breast cancer undergoing neoadjuvant chemotherapy.MethodsBlood samples were collected from 61 locally advanced breast cancers (36 HER2- and 25 HER2+) at diagnosis and from 23 healthy women. Immunophenotypic profiling of circulating and intratumor immune cells, including regulatory T (Treg) cells, was assessed by flow cytometry and immunohistochemistry, respectively. Serum levels of 10 different cytokines were assessed by multiplex immunoassays. CD8+ T cell responses to multiple tumor-associated antigens (TAA) were evaluated by IFN-γ-enzyme-linked immunosorbent spot (ELISPOT). The Students t test for two tailed distributions and the Wilcoxon two-sample test were used for the statistical analysis of the data.ResultsThe proportion of circulating immune effectors was similar in HER2+ patients and healthy donors, whereas higher percentages of natural killer and Treg cells and a lower CD4+/CD8+ T cell ratio (with a prevalence of naïve and central memory CD8+ T cells) were observed in HER2- cases. Higher numbers of circulating CD8+ T cells specific for several HLA-A*0201-restricted TAA-derived peptides were observed in HER2+ cases, together with a higher prevalence of intratumor CD8+ T cells. Serum cytokine profile of HER2+ patients was similar to that of controls, whereas HER2- cases showed significantly lower cytokine amounts compared to healthy women (IL-2, IL-8, IL-6) and HER2+ cases (IL-2, IL-1β, IL-8, IL-6, IL-10).ConclusionsCompared to HER2- cases, patients with HER2-overexpressing locally advanced breast cancer show a more limited tumor-related immune suppression. This may account for the clinical benefit achieved in this subset of patients with the use of drugs acting through, but also promoting, immune-mediated effects.

Improved Natural Killer cell activity and retained anti-tumor CD8(+) T cell responses contribute to the induction of a pathological complete response in HER2-positive breast cancer patients undergoing neoadjuvant chemotherapy.

BackgroundLocally advanced HER2-overexpressing breast cancer (BC) patients achieve a high rate of pathological complete responses (pCR) after neoadjuvant chemotherapy (NC). The apparently unaltered immune proficiency of these patients together with the immune-modulating activities of NC drugs suggest a potential contribution of host immunity in mediating clinical responses. We thus performed an extensive immunomonitoring in locally advanced BC patients undergoing NC to identify immunological correlates of pCR induction.MethodsThe immune profile of 40 HER2-positive and 38 HER2-negative BC patients was characterized at diagnosis and throughout NC (Paclitaxel and Trastuzumab, or Docetaxel and Epirubicin, respectively). The percentages of circulating immune cell subsets including T and B lymphocytes, Natural Killer (NK) cells, regulatory T cells, T helper 17 lymphocytes, were quantified by multiparametric flow cytometry. NK cells functional activity was evaluated through the analysis of NF-kB nuclear translocation by Multispectral flow cytometry, and with the in vitro monitoring of Trastuzumab-mediated antibody-dependent cell cytotoxicity (ADCC). CD8+ T cell responses against six different tumor-associated antigens (TAA) were characterized by IFN-γ ELISPOT and IFN-γ/IL-2 DualSpot assays.ResultsAfter NC, HER2-positive patients showed a significant increase in the number of NK cells and regulatory T cells irrespective of the pathological response, whereas patients undergoing a pCR disclosed higher percentages of T helper 17 cells. Notably, a significant increase in the number of activated NK cells was observed only in HER2-positive patients achieving a pCR. Characterization of anti-tumor T cell responses highlighted sustained levels of CD8+ T cells specific for survivin and mammaglobin-A throughout NC in patients undergoing a pCR in both arms. Moreover, HER2-positive patients achieving a pCR were characterized by a multi-epitopic and polyfunctional anti-tumor T cell response, markedly reduced in case of partial response.ConclusionsThese results indicate that maintenance of functional T cell responses against selected antigens and improvement of NK cell proficiency during NC are probably critical requirements for pCR induction, especially in HER2-positive BC patients.Trail registration: Trial registration number: NCT02307227, registered on ClinicalTrials.gov (http://www.clinicaltrials.gov, November 26, 2014).

Therapeutic management of breast cancer in the elderly

Introduction: Over the last few decades, the population of developed countries has aged. Breast cancer is the most common cancer among the increased numbers of older women. The choice of adjuvant treatment is particularly difficult in older women because the oncologist has to balance reduction of the risk of recurrence with patient-related comorbidities that may increase the risk of treatment-related toxicity and influence patient survival. Areas covered: This article describes the concept of a comprehensive geriatric assessment and reviews the current literature on biological and pathological characteristics of breast cancer in the elderly, including genomic assays recently available in the clinic. Endocrine, targeted and chemotherapy treatments both in adjuvant and metastatic setting are also covered. Expert opinion: A new generation of studies aimed to re-evaluate treatments in the various subtypes of breast cancer is needed. Whether this will be possible through prospective studies (especially in the adjuvant setting) is unknown. An alternative direction for further research in the elderly could be a reappraisal of old studies with carefully planned subtype analyses. Whatever the direction, the management of elderly breast-cancer patients is inherently multidisciplinary: the contribution of medical and allied health professionals is essential to the provision of optimal care.

Genotype-guided dosing study of FOLFIRI plus bevacizumab in patients with metastatic colorectal cancer

Purpose: UGT1A1*28 confers a higher risk of toxicity in patients treated with irinotecan. Patients with *1/*1 and *1/*28 genotypes might tolerate higher than standard doses of irinotecan. We aimed to identify the MTD of irinotecan in patients with metastatic colorectal cancer (mCRC) with *1/*1 and *1/*28 genotypes treated with FOLFIRI plus bevacizumab, and to determine whether bevacizumab alters irinotecan pharmacokinetics. Experimental Design: Previously untreated patients with mCRC (25 *1/*1; 23 *1/*28) were given FOLFIRI plus bevacizumab every 2 weeks. The irinotecan dose was escalated using a 3 + 3 design in each genotype group as follows: 260, 310, and 370 mg/m2. The MTD was the highest dose at which <4/10 patients had a dose-limiting toxicity (DLT). Pharmacokinetics of irinotecan and SN-38 were measured on days 1 to 3 (without bevacizumab) and 15 to 17 (with bevacizumab). Results: For *1/*1 patients, 2 DLTs were observed among 10 patients at 310 mg/m2, while 370 mg/m2 was not tolerated (2 DLTs in 4 patients). For *1/*28 patients, 2 DLTs were observed among 10 patients at 260 mg/m2, while 310 mg/m2 was not tolerated (4 DLTs in 10 patients). Neutropenia and diarrhea were the most common DLTs. Changes in the AUCs of irinotecan and SN-38 associated with bevacizumab treatment were marginal. Conclusions: The MTD of irinotecan in FOLFIRI plus bevacizumab is 310 mg/m2 for UGT1A1 *1/*1 patients and 260 mg/m2 for *1/*28 patients. Bevacizumab does not alter the pharmacokinetics of irinotecan. The antitumor efficacy of these genotype-guided doses should be tested in future studies of patients with mCRC treated with FOLFIRI plus bevacizumab. Clin Cancer Res; 23(4); 918–24. ©2016 AACR.

Malignant cardiac tumors: diagnosis and treatment

Primary malignant cardiac tumors are represented by sarcomas and non-Hodgkin lymphomas. They are rare, affect mostly patients in the fourth decade of life and have a severe prognosis. Both the diagnosis and the treatment require a multidisciplinary approach, and the cardiologist plays a central role both in the diagnosis and in the follow-up. The prognosis may be improved by a careful planning of surgery and by the use of multimodality treatment, including complementary chemotherapy and radiation therapy. A strict follow-up must be planned even after apparently complete cure.

Lapatinib-based therapy in heavily pretreated HER2-positive metastatic breast cancer: a single institution experience

AIMS AND BACKGROUND Lapatinib in combination with capecitabine is feasible in patients with HER2-positive metastatic breast cancer pretreated with anthracyclines, taxanes and trastuzumab, but inferior results were reported in the global lapatinib expanded access program in comparison with the phase III registration trial. METHODS and study design. Women with HER2-positive metastatic breast carcinoma after antracycline, taxane and trastuzumab-based regimens were treated at progression with lapatinib plus capecitabine. The outcome of these patients was evaluated. From April 2007 to August 2010, 68 patients were treated overall. RESULTS Median progression-free survival was 6 months (range, 1-29), and median overall survival was 26 months (range, 1-39). Eight (12%; 95% CI, 4-25) patients experienced a complete response. Partial response was observed in 22 patients (31%; 95% CI, 20-42), for an overall response rate of 43% (95% CI, 31-55). The treatment with lapatinib plus capecitabine was well tolerated, with grade 3-4 toxicity reported in few patients, and no treatment-related deaths were noted. Of note, no cardiac toxicity was reported in this highly pretreated group of patients or in the subgroup of 10 elderly patients. CONCLUSIONS Our data confirm that lapatinib plus capecitabine is an active regimen even in heavily pretreated patients with visceral and brain metastases and is feasible and active also in selected elderly patients.

Improved outcome with multimodal treatment and imatinib rechallenge in advanced GIST

Dear Editor: Gastrointestinal stromal tumor (GIST) is the most common mesenchymal neoplasm of the digestive tract. Surgery and imatinib are the recommended treatments for advanced highrisk GIST. However, even in most patients showing an initial response, imatinib resistance is developed within a few years. After progression, current therapeutic options include the switch to other tyrosine kinase inhibitors (TKI) or surgical debulking. We here report long-term survival in a metastatic GIST patient treated with reiterated multimodal therapeutic approaches and as a fourth-line treatment, imatinib rechallenge. Molecular analysis of the tumors provided a framework for interpretation of the pattern of response observed. A 52-year-old man was referred to surgical intervention for an abdominal mass in January 2003. Surgery revealed a major (15 cm) intestinal tumor, together with other smaller peritoneal nodules, which were also resected. At pathological evaluation, the lesions displayed a spindle cell morphology, were CD117 (KIT) positive, CD34 positive, desmin negative, focally positive for smooth muscle actin and S100, and negative for cytokeratins, consistent with a diagnosis of metastatic, high-risk GIST. Six months later, the patient relapsed and underwent a II surgery with complete resection (R0) of a mass of 12 cm at the right iliac fossa, pathologically confirmed as GIST. Adjuvant imatinib (400 mg/day) was administered for 24 months, without evidence of recurrence. Eight months after imatinib discontinuation, a CT scan revealed a mass of 6 cm at the right iliac fossa and multiple peritoneal nodules without clinical symptoms. Imatinib was therefore resumed at the same dose. Radiological assessment showed an initial response, but 14 months later, disease progression was observed. Patient then underwent curative III surgery (R0) for a major mass (8 cm) proximal to the rectum and multiple peritoneal and omental nodules of smaller size. After surgery, imatinib was continued at 400 mg/day until, 8 months later, disease recurrence was documented by CT scan with a larger pararectal mass and multiple small peritoneal nodules. Patient was again subjected to tumor debulking (IV surgery) with microscopic residual disease (R=1). Patient resumed imatinib at a higher dose (800 mg/day). Due to disease progression, after 3 months, he was switched to sunitinib (50 mg/day), reducing the dosage (37.5 mg/day) after the first cycle because of poor tolerance. Treatment was continued for 15 months until pelvic progression was detected. For his third line treatment, the patient was enrolled in a clinical trial evaluating nilotinib efficacy in patients with unresectable/metastatic GIST and refractory to imatinib and/ or sunitinib. Nilotinib was given at 800 mg/day for 2 months, until CT scan showed abdominal progression. Surgical Gianmaria Miolo and Elena Torrisi contributed equally to this work.