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A weekly administration of alternating irinotecan or oxaliplatin associated to 5-Fluorouracil in advanced colorectal cancer was planned in order to evaluate a new schedule maintaining dose intensities of each drug as in double combinations and tolerability of the triplet association. The following weekly schedule was administered: irinotecan, days 1 and 15; oxaliplatin, days 8 and 22; 5-fluorouracil (5-FU) over 12-h (from 10:00 p.m. to 10:00 a.m.) timed flat infusion, days 1-2, 8-9, 15-16 and 22-23, every 4 weeks. Dose- finding and phase II study were planned. Thirteen patients were enrolled in the dose-finding study and 23 in the phase II study. The recommended doses of our study are: irinotecan 160 mg/m(2); oxaliplatin 80 mg/m(2); 5-FU 900 mg/m(2). The dose-limiting toxicity was diarrhea (35% of patients) but no cases of febrile neutropenia were observed. In 30 patients assessable for response two complete (6.7%) and 18 partial (60%) responses were observed, for an overall response rate of 66.7% (alpha 0.05, CI+/-17). The triplet association using this weekly alternating schedule is an active and well-tolerated outpatient regimen. Surgical removal of residual disease was considered in 5 patients and a radical resection was performed in 5 patients (147 %).

Purpose This phase II study, evaluated the activity and cardiotoxicity of first-line epirubicin plus low-dose trastuzumab (LD-T) in patients with HER2 positive MBC. Methods Patients received epirubicin 90 mg/sqm every 3 weeks plus weekly LD-T (2 mg/kg loading dose, then 1 mg/kg). After 6/8 cycles of epirubicin, single agent trastuzumab was continued. Cardiotoxicity was defined as signs or symptoms of congestive heart failure (CHF), or ≥15% decline in LVEF without symptoms, or

Background Recent data have shown that cardiotoxicity represents a potentially important side-effect in patients treated with sunitinib. We reviewed cardiac adverse events in patients with metastatic renal cell carcinoma (RCC) who underwent treatment with this agent. Patients and methods The medical records of 175 patients with metastatic RCC treated with sunitinib at eight Italian institutions were retrospectively reviewed. Alterations in left ventricular ejection fraction (LVEF) and blood pressure were evaluated. Patients with preexisting cardiac risk factors were specifically scrutinized for increased expression of cardiac changes. Results Grade 3 hypertension was seen in 17 patients (9.7%); in 12 of these 17, hypertension developed after receiving the third sunitinib cycle. Among these 17 patients, 12 (70.6%) also experienced left ventricular systolic (LVEF) dysfunction; in all, 33 of the 175 patients (18.9%) developed some degree of cardiac abnormality, of which 12 were classified as grade 3 LVEF dysfunction and/or congestive heart failure (CHF) (6.9%). Significant univariate associations for predictors of CHF were history of hypertension (P = 0.008), history of coronary heart disease (P = 0.0005) and prior treatment with an angiotensin-converting enzyme inhibitor (P = 0.04). Multivariate analysis suggested that a history of coronary artery disease [odds ratio (OR) 18, 95% confidence interval (CI) 4–160, P = 0.005] and hypertension (OR 3, 95% CI 1.5–80, P = 0.04) was the only significant independent predictors of CHF. Conclusions Patients undergoing sunitinib, especially those with a previous history of hypertension and coronary heart disease, are at increased risk for cardiovascular events and should be monitored for exacerbations of their hypertension and for evidence of LVEF dysfunction during treatment.

Erlotinib is a small-molecule tyrosine kinase inhibitor targeted EGFR, known to be overexpressed in a variety of cancers, including prostate cancer. Clinical trials showed insignificant clinical benefit in patients with castration resistant prostate cancer both when EGFR inhibitors were administered as monotherapy or in association with antiandrogens or chemotherapeutics. Why, differently to other tumors, have EGFR inhibitors been so ineffective in human prostate cancer? This is the question that we have set in this report.For this purpose, the effectiveness of erlotinib, a selective EGFR inhibitor, in a wide range of prostate cancer cells (wild type or engineered to overexpress peculiar proteins including androgen receptor and PTEN).We demonstrated that the effectiveness of erlotinib was inversely correlated to the EGFR/Her2 ratio rather than EGFR/p-EGFR or Her2/p-Her2 levels. Chronic treatment with bicalutamide induced overexpression of Her2 and reduction of EGFR/Her2ratio and this was associated with increased Akt and Erk activity. In these conditions of treatment a reduced efficacy of erlotinib was observed. At the same time, an increased efficacy versus erlotinib was documented in cancer cells chronically exposed to DHT. In these culture conditions low levels of Her2 and increased EGFR/Her2 ratio were evidenced.Taken together, our results seem to suggest that a low EGFR/Her2 ratio and PTEN absence are the main factors responsible of erlotinib inefficacy. Therefore the inhibition of EGFR could have important antitumor effects in hormone-naive rather than in hormonally treated patients.

One of the major obstacles in curing prostate cancer is the development of drug resistance. It is not only imperative to discover the molecular basis of resistance but also to find therapeutic agents that can disrupt the resistant pathways. Tumor necrosis factor TNF-related apoptosis-inducing ligand TRAIL-like ligands or agonist TRAIL-receptor monoclonal antibodies have entered phase I and II clinical trials with a very limited cytotoxic profile when used systemically in a variety of cancers. Therefore, TRAIL-receptor agonists are new proapoptotic pharmaceutical agents with great potential as new cancer therapeutic agents. Although many cancer cells undergo TRAIL-mediated apoptosis, some are resistant to TRAIL. Therefore, we have been investigating mechanisms to overcome TRAIL resistance in cancer cells so that TRAIL-associated compounds can be used effectively in clinical trials. Epigenetic inactivation of proapoptotic genes, or activation of survival signaling, can cause cross-resistance to several anti-tumor therapies and to immune cytotoxic lymphocytes. We hypothesize that 5-aza-2 deoxycytidine aza-dCR, decitabine may render TRAIL-resistant prostate cancer cells sensitive to caspase-8-mediated apoptosis and may, therefore, be therapeutically efficient. We evaluated the antiproliferative effects of decitabine on the following four prostate cancer cell lines: well-differentiated AR positive LnCaP p53(+), PTEN- and 22rv1 p53(+) and PTEN(+)]; poorly-differentiated AR negative PC3 p53-, PTEN- and DU145 p53 mutant, PTEN(+). Here, we provide evidence that treatment with sub-optimal concentrations of decitabine are additive to TRAIL effects in well-differentiated PCa cells whereas the same treatment shows synergistic effects in poorly-differentiated PCa cells through increased caspase-8 expression, down-modulation of Akt activation and through the expression of certain anti-apoptotic molecules including FLIP, PED/PEA-15, survivin and c-IAP-1. Our findings demonstrate that decitabine at relatively low concentrations restores caspase-8 expression and sensitises resistant PCa cells to TRAIL-induced apoptosis leading to important implications in novel therapeutic strategies targeting defective apoptosis pathways in advanced prostate tumors.

The carcinoembryonic antigen (CEA) test was studied in 54 patients with advanced stages of colon cancer which was treated with high doses of folinic acid + fluorouracil. The CEA test correlates evaluated included: prognostic value, performance status, metastatic pattern, histologic grading, predictive value for response to chemotherapy, and value differences in cases with partial response to therapy. CEA levels less than 5 ng/ml corresponded to a greater survival time than did levels greater than 5 ng/ml. A correlation of CEA with performance status and with metastatic pattern was demonstrated. A progressive increase in average CEA values corresponded to increases in neoplastic mass. Although CEA levels were not found to be an index for predicting the response to chemotherapy, there was a significant different between pre- and posttreatment levels for partial response. The results suggest that CEA offers an additional criterion for evaluating the response of colon cancer to chemotherapy and it also has a role in the staging of advanced disease.

The clinical efficacy of ErbB1 kinase inhibitors is limited by the development of acquired autoresistance. The activation of alternative signaling pathways can contribute to gefitinib resistance. In this study, we demonstrate that the continuous in vitro exposure of the phosphatase and tensin homologue (deleted from chromosome 10)-negative prostate cancer (PC)3 cell line to gefitinib resulted in a sustained growth inhibition of 50% for about 2 months, but afterwards the surviving cells resumed their usual proliferation rate. During chronic treatment, gefitinib-treated cells developed drug resistance undergoing a G0/G1 cell cycle arrest, with a corresponding reduction in the G2/M cells without evident cell apoptosis, and thus a tyrosine kinase inhibitor-resistant (TKI-R) PC3 cell subline was isolated. TKI-R cells show i) an increment in basal ERK activation, ii) an epidermal growth factor-mediated and gefitinib insensitive ERK phosporylation, iii) increased levels of Her2/Neu, iv) a significant decrement in epidermal growth factor receptor (EGFR) expression, v) a very low sensitivity against EGFR TKIs and blocking antibodies, vi) a moderate increase in the sensitivity to growth inhibition by the Her2 inhibitor, AG825 or by 2C4, the humanized monoclonal antibody which blocks Her2 heterodymerization, vii) an increased expression of the neutrophine receptors, TrkA and TrkB, and viii) a significantly increased sensitivity to growth inhibition by the TrkA inhibitor, CEP701. Treatment with a mitogen-activated-protein kinase inhibitor abolished gefitinib resistance completely. Therefore, the ability of tumor cells to maintain high ERK activity under EGFR inhibition could represent a potential mechanism of the resistance to gefitinib.

RATIONALE Neurotrophine tyrosine kinase receptors (NTR) are expressed in prostate carcinoma (PCa), and their distribution seems to be related to disease malignancy. MATERIAL AND METHODS In this article we analyzed the expression of NTRK1 (TrkA), NTRK2 (TrkB), NTRK3 (TrkC), and p75NTR in a 102 patient cohort with clinically localized tumors, which had been surgically treated with radical prostatectomy (RP). Among these, 61 patients received RP as sole treatment, and 41 patients received neoadjuvant hormone therapy (NHT) for 120 days with bicalutamide 150 mg/day. In addition, we analyzed the NTR expression in vitro in the androgen receptor positive, androgen-sensitive cell strains derived from CWR22R. RESULTS AND CONCLUSIONS We demonstrated that: (i) TrkA and TrkC levels were significantly upmodulated, whereas (ii) p75NTR seemed to be reduced, and (iii) TrkB expression seemed to be not affected by NHT. TrkA were constitutively activated when its levels were very high. In vitro studies showed that the dehydrotestosterone (DHT) was able to maintain low TrkA and TrkC protein levels. Conversely, DHT was able to maintain p75NTR at high levels. Bicalutamide treatment induced TrkA and TrkC and reduced p75NTR expression. Antiproliferative effects of CEP701 were dependent to TrkA levels. A therapeutical effect of CEP701 was seen in all culture conditions, and bicalutamide seemed to sensitize prostate cancer cells to the effects of a pan TrkA inhibitor CEP701, suggesting that a sequential therapy between these drugs could further increase the efficacy of Trk inhibition. Prostate 67: 1255–1264, 2007. © 2007 Wiley-Liss, Inc.

This phase II study was conducted to evaluate tumor response rate and safety profile of dose-dense epirubicin plus cyclophosphamide followed by docetaxel plus capecitabine given preoperatively to patients with stage II or IIIA breast cancer.Patients underwent four cycles of dose-dense cyclophosphamide 600 mg/m(2) and epirubicin 90 mg/m(2) every 2 weeks followed by two cycles of docetaxel 36 mg/m(2) on days 1, 8, and 15 plus capecitabine 1250 mg/m(2) on days 5-18 every 4 weeks, with prophylactic pegfilgrastim. The primary objective of the study was to determine the incidence of pathologic complete response defined as the absence of invasive or in situ cancer in the breast and the axillary nodes at definitive surgery.Forty-four patients were enrolled in the study and 41 (93%) were assessable for response to chemotherapy. An objective clinical response was observed in 38 (93%) patients. Seven patients (17.1%) exhibited a pathologic complete response. Breast-conserving surgery was carried out in 36 (88%) patients. Grade 3/4 neutropenia occurred in 4.3% of 252 administered chemotherapy cycles. No febrile neutropenia, cardiac toxicity, thrombocytopenia or other serious adverse event was registered.The sequential combination of dose-dense epirubicin plus cyclophosphamide followed by docetaxel plus capecitabine is an effective and well-tolerated neo-adjuvant chemotherapy for stage II, IIIA breast cancer.

Background Breast cancer (BC) is a leading cause of death among women. Among the major risk factors, an important role is played by familial history of BC. Germ-line mutations in BRCA1/2 genes account for most of the hereditary breast and/or ovarian cancers. Gene expression profiling studies have disclosed specific molecular signatures for BRCA1/2-related breast tumors as compared to sporadic cases, which might help diagnosis and clinical follow-up. Even though, a clear hallmark of BRCA1/2-positive BC is still lacking. Many diseases are correlated with quantitative changes of proteins in body fluids. Plasma potentially carries important information whose knowledge could help to improve early disease detection, prognosis, and response to therapeutic treatments. The aim of this study was to develop a comprehensive approach finalized to improve the recovery of specific biomarkers from plasma samples of subjects affected by hereditary BC. Methods To perform this analysis, we used samples from patients belonging to highly homogeneous population previously reported. Depletion of high abundant plasma proteins, 2D gel analysis, liquid chromatography-tandem mass spectrometry (LC-MS/MS) and bioinformatics analysis were used into an integrated approach to investigate tumor-specific changes in the plasma proteome of BC patients and healthy family members sharing the same BRCA1 gene founder mutation (5083del19), previously reported by our group, with the aim to identify specific signatures. Results The comparative analysis of the experimental results led to the identification of gelsolin as the most promising biomarker. Conclusions Further analyses, performed using a panel of breast cancer cell lines, allowed us to further elucidate the signaling network that might modulate the expression of gelsolin in breast cancer.

e13031 Background: EDO-S101 combines the DNA damaging effect of bendamustine with the pan-histone deacetylase inhibitor vorinostat. The molecule was tested in models of human MGMT negative and posi...

Inherited mutations of the BRCA1/2 genes confer a significantly increased risk for breast and/or ovarian cancer development. Several models were elaborated to help genetic counsellors in selecting individuals with high probability of being mutation carriers. The IC software, a country-customized version of the Brcapro model, was recently shown to be particularly accurate in the prediction of carrier probability status in the Italian population. Here, we used our independent series of 70 breast/ovarian cancer families to analyze the performances of the IC software and compare it to widely used models, such as Brcapro and the Myriad mutation prevalence tables. Analysis of the areas under the receiver operator characteristics (ROC) curves indicated that overall the models performed well. However, the IC software and Myriad tables were more efficient in predicting mutated cases, showing a higher sensitivity (94 and 88%, respectively) and negative predictive value (NPV, 94 and 92%, respectively) compared to Brcapro (sensitivity 71 and NPV 83%). IC software also appeared particularly accurate in the identification of families belonging the low mutation risk group (

Purpose: To assess the value of changes in the expression of topoisomerase IIα (TopoII) and the proto-oncogene erbB-2 (HER-2) as predictors of relapse-free survival in women with operable breast cancer treated with anthracycline-based neoadjuvant chemotherapy. Patients and Methods: Seventy-seven patients with primary breast cancer who had undergone neoadjuvant anthracycline-based chemotherapy were included in the present study. TopoII and HER-2 were measured by immunohistochemistry in prechemotherapy and postchemotherapy (at the time of surgery) tumor specimens, and the value of their changes as predictors of relapse-free survival were evaluated by Kaplan-Meier and Cox proportional hazard regression analyses. Results: Neoadjuvant chemotherapy resulted in a significant reduction in the percentage of cells expressing TopoII (P < 0.0001). No significant change was observed for HER-2. TopoII and HER-2 expression before chemotherapy predicted tumor response to treatment. Changes in TopoII expression after chemotherapy were strongly associated with a poor relapse-free survival (P < 0.0001) in a Cox multivariate analysis adjusted for other clinicopathologic prognostic factors. Conclusion: Changes in TopoII expression after anthracycline-based neoadjuvant chemotherapy is an independent predictor of a poor relapse-free survival in patients with breast cancer. Tumor cells displaying an increased TopoII expression after treatment may be responsible for relapses, and may, therefore, define a group of patients with anthracycline-resistant breast cancer.

A dose-finding study was designed to determine the maximum tolerated dose (MTD) of a bimonthly 12-h (10:00 p.m to 10:00 a.m), timed flat infusion (TFI) of 5-fluorouracil (5-FU) plus irinotecan (CPT-11), without leucovorin (LV), for metastatic colorectal carcinoma (CRC). A total of 33 patients were treated. Seven dose levels included a fixed CPT- 11 dose of 180 mg/m 2 on days 1 and 15 (d 1,15 ) and escalating doses of 5-FU 600-1200 mg/m 2 on days 1-4 and 15-18 (d 1-4,15-18 ). Dose-limiting toxicities (DLTs) were: grade 3-4 non-hematologic, grade 4 hematologic and any toxicity causing a more than a 2-week delay in treatment. The MTD was reached at the seventh dose level. DLTs were observed in 5/8 patients (63%): G3 diarrhea, 2 patients, associated with G3 mucositis in one instance; G4 neutropenia, 2 patients, associated with severe asthenia in 1 patient; G3 hand-foot syndrome, 1 patient. The recommended doses (RDs) were established at the sixth dose level: 5-FU, 1100 mg/m 2 /d 1-4,15-18 ; CPT-11 180 mg/m 2 /d 1,15 [5-FU and CPT-11 dose intensity (DI), 2200 and 90 mg/m 2 per week (w), respectively]. At the recommended dose, the DLTs in 38 cycles were: mucositis, 2 cycles (5%); afebrile G4 neutropenia and hand-foot syndrome, 1 cycle (3%). In 24 assessable patients, the overall response rate was 37.5%. The present CPT-11/5-FU schedule is highly tolerable in an outpatient setting using the highest recommended 5-FU dose effective in advanced CRC.

Familial aggregations of breast/ovarian cancer cases frequently depend on BRCA1/2 pathogenic mutations. Here we counselled 120 Italian breast/ovarian cancer families and selected 73 probands for BRCA1/2 mutation screening. Through this analysis we defined the prevalence of BRCA1/2 pathogenic mutations occurring in Italian breast/ovarian cancer families, enlarged the spectrum of Italian BRCA1/2 mutations by 15% and report on the identification of 13 novel variants, including two deleterious truncating mutations and two potentially pathogenic missense mutations, on the BRCA1 and BRCA2 genes. Finally in hereditary breast cancer families with three or more female breast cancer cases we observed a low mutation prevalence and a significant association with BRCA2 mutations.

Nine patients with advanced cancer suffering from idiopathic severe sweating were treated with gabapentin (600-1,800 mg/day). All patients responded to the treatment; five patients experienced transient drowsiness. Gabapentin seems to be a safe and effective treatment for idiopathic sweating in advanced cancer patients. Further studies are warranted to confirm or refute these results.

Background Breast cancer is an extremely complex disease, characterized by a progressive multistep process caused by interactions of both genetic and non-genetic factors. A combination of BRCA1 and BRCA2 gene mutations appears responsible for about 20%–30% of the cases with breast cancer familial history. The prevalence of BRCA1/2 pathogenic mutations largely varies within different populations; in particular, the rate of mutations in Italian breast and/or ovarian cancer families is rather controversial and ranges from 8% to 37%. Patients and methods Of the 152 breast/ovarian cancer families counseled in our centre, 99 were selected for BRCA1/2 mutation screening according to our minimal criteria. The entire coding sequences and each intron/exon boundary of BRCA1/2 genes were screened by direct sequencing (PTT limited to BRCA1 exon 11). For each proband, the a priori probability of carrying a pathogenic BRCA1/2 germline mutation was calculated by means of different mutation prediction models (BRCApro, IC and Myriad Table) in order to evaluate their performances. Results Our analysis resulted in the identification of 25 and 52 variants in the BRCA1 and BRCA2 genes, respectively. Seventeen of them represent novel variants, including four deleterious truncating mutations in the BRCA2 gene (472insA, E33X, C1630X and IVS6+1G>C). Twenty-seven of the 99 probands harbored BRCA1 (n = 15) and BRCA2 (n = 12) pathogenic germline mutations, indicating an overall detection rate of 27.3% and increasing by more than 15% the spectrum of mutations in the Italian population. Furthermore, we found the lowest detection rate (19.4%) in pure hereditary breast cancer family subset. All of the prediction models showed praises and faults, with the IC software being extremely sensitive but poorly specific, compared to BRCApro. In particular all models accumulated most false-negative prediction in the HBC subset. Interestingly preliminary results of a study addressing the presence of genomic rearrangements in HBC probands with BRCApro or IC prediction scores ≥95%, provided evidence for additional mutations undetectable with our conventional screening for point mutations. Conclusions : Altogether our results suggest that HBC families, the largest pool in our series, represent an heterogeneous group where the apparently faulty performances of the prediction models might be at least partially explained by the presence of additional kinds of BRCA1/2 alteration (such as genomic rearrangements) or by mutations on different breast cancer related genes.

This work was conducted to evaluate symptoms assessment and use of patient-tailored protocols in clinical practice among Italian medical oncologists.A questionnaire based on four topics (assessment of symptoms, assessment of a specific symptom, assessment of pain, use of patient-tailored protocols of treatment) was administered to 250 Italian medical oncologists.Of these oncologists, 43.7% used multiple symptoms tools and 37.9% used symptom specific tools; 58.9% used some instrument to assess pain. More than a third of the respondents (35.5%) used patient-tailored protocols. No statistical differences were found regarding region of residency, availability of consultants in pain therapy and/or palliative care, colleagues with main interest on palliative care, and beds dedicated to palliative care. Statistically significant differences were found regarding the position (staff/resident) in three out four topics.Among Italian medical oncologists, the instruments used for assessment of symptoms are poorly employed. Even when these instruments are used, patient-tailored protocols are rarely administered.

The Supportive Care Task Force (SCTF) was established within the Medical Oncology Department at the University of L’Aquila in May 2002. The missions of the SCTF were to allow systematic evaluation and treatment of symptoms, to warrant continuity of care in all phases of disease and to provide medical oncology residents with training in the treatment of symptoms. A medical oncologist, two senior residents in medical oncology and a registered nurse comprised the SCTF. A psychiatrist, two neurologists, a dietician, and two physiotherapists served as consultants or on a part-time basis. Four beds in two-bedded rooms inside the Medical Oncology Department were reserved to SCTF. A close integration with the physicians of the Medical Oncology Department was realised. The only criterion to admission was the presence of uncontrolled symptoms. Patients were evaluated and monitored with the visual analogue scale for pain and with the Edmonton Symptom Assessment Scale (ESAS). The Palliative Prognostic Score (PaP Score) was employed to assess the prognosis. Non-clinical needs were evaluated with the Need Evaluation Questionnaire (NEQ). Protocols for the treatment of common symptoms were available in written form for consultation by physicians, residents and nurses. From 1 May 2002 to 31 May 2004, we observed 208 patients: 111 women and 97 men. The median age was 64.7 (range 28–90) years. Fifty-four patients (25.9%) were admitted more than once, for a total of 285 admissions. One hundred ninety-nine admissions (69.5%) were for supportive care while 86 admissions (30.5%) were for supportive care and active treatment. The most frequent symptoms were asthenia and anorexia. We registered excellent results regarding the treatment of pain, nausea and dyspnea while psychological symptoms, anorexia and asthenia proved more difficult to treat. Two hundred twenty patients were discharged: 142 (49.8%) home; 76 (26.7%) to the Home Care Service and two (0.7%) to others units of the hospital. Sixty-five (22.8%) died in our unit.

Familial breast cancer, whether associated or not with particular other breast cancer features (male, early onset, bilateral breast cancer), determines a wide and variable risk of developing breast cancer in the 'unpatients' (unaffected individuals) of these families, particularly in those harboring a genetic predisposition. The antiestrogen tamoxifen has been proposed in different trials to prevent breast cancer in women at risk. The NSABP-P1 study demonstrated that tamoxifen drastically reduced (by approximately 50%) the incidence of breast cancer in women at risk selected according to the Gail score. The preventive effect was particularly consistent in postmenopausal women and in those showing familial breast cancer (three or more affected patients). BRCA1/BRCA2 (BRCA1/2) gene analysis in women accrued in the NSABP-P1 trial who developed breast cancer showed that tamoxifen chemoprevention reduced breast cancer incidence in BRCA2 carriers. Different chemoprevention trials are ongoing to compare different selective estrogen receptor modulators and aromatase inhibitors with tamoxifen. The Italian Consortium of Hereditary Breast Ovarian Cancer recently developed the Aromasin Prevention Study, a multicenter, double-blind, randomized, placebo-controlled phase III study evaluating the effect of the aromatase inhibitor exemestane for chemoprevention in postmenopausal women carriers of BRCA1/2 genetic predisposition. Women who are postmenopausal unaffected carriers of BRCA1/2 mutations will be selected by participating institutions and randomly assigned to receive either oral exemestane or oral placebo every day for 3 years in order to reduce the incidence of breast cancer. Genetic counseling and the detection of predisposing BRCA1/2 mutations are mandatory before accrual into the study. Signed informed consents for the performing of BRCA1 and BRCA2 genetic analysis and for enrollment into the study are required. Eligible women will be followed thereafter in order to evaluate the efficacy of exemestane in reducing the incidental rate of breast cancer in unaffected postmenopausal carriers of BRCA1/2 mutations.

Accurate estimates of breast and ovarian cancer penetrance in BRCA1/2 mutation carriers are crucial in genetic counseling. Estimation is difficult because of the low frequency of mutated alleles and the often-uncertain mechanisms of family ascertainment. We estimated the penetrances of breast and ovarian cancers in carriers of BRCA1/2 mutations by maximizing the retrospective likelihood of the genetic model, given the observed test results, in 568 Italian families screened for germline mutations. The software BRCAPRO was used as a probability calculation tool in a Markov Chain Monte Carlo approach. Breast cancer penetrances were 27% (95% CI 20-34%) at age 50 years and 39% (27-52%) at age 70 in BRCA1 carriers, and 26% (0.18-0.34%) at age 50 and 44% (29-58%) at age 70 in BRCA2 carriers, and ovarian cancer penetrances were 14% (7-22%) at age 50 and 43% (21-66%) at age 70 in BRCA1 carriers and 3% (0-7%) at age 50 and 15% (4-26%) at age 70 in BRCA2 carriers. The new model gave a better fit than the current default in BRCAPRO, the likelihood being 70 log units greater; in addition, the observed numbers of mutations in families stratified by gene and by cancer profile were not significantly different from those expected. Our new penetrance functions are appropriate for predicting breast cancer risk, and for determining the probability of carrying BRCA1/2 mutations, in people who are presently referred to genetic counseling in Italy. Our approach could lead to country-customized versions of the BRCAPRO software by providing appropriate population-specific estimates.

Mouse bone marrow cells cultured with human breast cancer MCF-7 cell-conditioned media showed osteoclastogenesis with an increment of bone resorption, although conditioned media from an adriamycin-selected MCF-7 clone (MCF-7ADR) had no effect. Consistently, MCF-7 cells induced 5-fold more in vivo experimental osteolytic bone metastases, with no soft tissue lesions, compared to MCF-7ADR cells. Paracrine factors stimulating (interleukin (IL)-6, IL-1beta, tumor necrosis factor-alpha (TNF-alpha)) or inhibiting (IL-12, IL-18, granulocyte macrophage-colony stimulating factor (GM-CSF)) osteoclastogenesis were significantly increased in MCF-7ADR relative to MCF-7 cells, suggesting that the inhibitory cytokines could selectively overwhelm the effects of the stimulatory ones. Treatment of osteoblast primary cultures with MCF-7-conditioned medium induced a selective upregulation of IL-6 expression, suggesting an indirect stimulation of osteoclastogenesis via the osteoblasts. MCF-7 and MCF-7ADR showed no difference in proliferation rate. However, a higher ability to migrate and invade gelatin and matrigel was observed in MCF-7ADR. Enhanced invasiveness might result from increased metalloproteinase (MMP) activity and cytoskeleton rearrangement. MCF-7ADR cells expressed higher levels of c-Src, focal adhesion kinase (FAK), and protein tyrosine kinase 2 (PYK2) involved in cell adhesion and motility. MCF-7 and MCF-7ADR expressed high and faint levels of functional estrogen receptor alpha (ERalpha), respectively. MCF-7ADR also showed significantly higher levels of the protein kinase C (PKC) alpha and beta2 and a selective activation of PKC compared to MCF-7, where the most abundant isoforms were beta1 and delta. Heat shock protein 27 (Hsp27) was more abundant in MCF-7 cells, but failed to translocate to the nucleus in response to heat shock. In conclusion, we have demonstrated that despite the fact that MCF-7ADR cells showed a more invasive phenotype relative to MCF-7, they have low potential to induce osteolytic bone lesions and stimulate osteoclastogenesis and osteoclast activity. Therefore, we believe that reduced aggressiveness of breast carcinomas could correlate with a greater osteolytic activity featuring their bone metastases.

The occurrence of mutations in the p53 tumor suppressor gene is a specific and recurring genetic event in solid tumors. P53 plays a pivotal role in multiple cellular processes such as cell growth control, DNA repair and programmed cell death. Genotoxic damage, also induced by chemotherapy or radiotherapy, induces p53 overexpression in order to control the rate of proliferating damaged cells, thus triggering the mismatch repair or apoptotic pathways. P53 inactivation determines a condition of genetic instability, justifying the subsequent susceptibility to acquire mutations of different other genes. P53 mutations are associated with worse prognosis and with chemo/radioresistance, due to the inability to trigger p53-dependent programmed cell death. Molecular diagnostic strategies show 32% p53 mutations in breast cancer. The analysis of the p53 gene performed by FAMA (Fluorescence Assisted Mismatch Analysis) in high-risk breast cancer patients withor = 10 involved axillary nodes may help identify a subset of very high risk BC patients (vHR-BC) with poorer prognosis and a subset with better prognosis, potentially responsive to medical treatments. The accurate evaluation of the p53 status can predict prognosis and sensitivity to chemotherapy, thus representing the first step toward better definition of therapeutic strategies according to the molecular characterization of the individual patient.

Alterations of the NBS1 gene are responsible for Nijmegen breakage syndrome (NBS), which is characterized by chromosomal instability, radiosensitivity, and cancer predisposition. NBS1 protein (Nibrin) is part of a molecular complex (NBS1– MRE11A–RAD50) that is functionally involved in DNA double-strand–break repair. Defects in recombination or in repair mechanisms at the level of DNA breakage can lead to chromosomal aberrations, genetic instability, as well as cancer predisposition syndromes (i.e., NBS, ataxia-telangiectasia, Bloom syndrome). In this study, we examined 20 cancer cell lines to evaluate the potential involvement of NBS1 in tumoral pathogenesis. Three different mutations, generating truncated or aberrant NBS1 transcripts, were identified at the level of NBS1 mRNA. In addition, two shorter NBS1 protein variants were detected in two cell lines. These data suggest a possible involvement of NBS1 in tumor development. © 2003 Wiley-Liss, Inc.

Objective: Mutations of the rhodopsin gene are responsible for autosomal dominant or recessive retinitis pigmentosa (RP). The present study reports the first prenatal diagnosis performed on chorionic villi biopsy of a pregnant woman affected by a severe form of autosomal dominant transmitted RP, due to the Arg135Trp substitution. Methods: The rhodopsin gene was analysed by automated direct sequencing and, for the first time, by fluorescence-assisted mismatch analysis (FAMA). The latter is an inexpensive, rapid and particularly sensitive method, based on the chemical cleavage of the mismatch in heteroduplex DNA molecules marked with strand-specific fluorophores. Results: FAMA is a feasible procedure for prenatal molecular diagnosis of rhodopsin mutations. The redundancy of signals obtained by FAMA and its sensitivity make it suitable for identifying exactly the position of the mutation and the nucleotide substitution. Conclusions: An association is proposed between FAMA and automated direct sequencing procedures, in order to achieve optimal results in terms of reliability for prenatal diagnosis of rhodopsin mutations.

e14575 Background: Bevacizumab (BEV) addiction to chemotherapy can predict increased efficacy in KRAS wild-type and mutant metastatic colorectal cancer (MCRC) patients. Codon 12 KRAS mutations diff...

512 Background: KRAS genotype does not significantly affect prognosis of MCRC patients treated with bevacizumab-containing chemotherapy. Codon 12 mutations increase aggressiveness by differently modifying mutated KRAS and downstream signalling, thus conferring worse clinical behavior. Prognostic relevance of the prevalent c.35 G > A KRAS mutation in MCRC patients fit and unfit for first-line bevacizumab added to triplet chemotherapy (FIr-B/FOx) was evaluated. Methods: KRAS codon 12/13 mutations were screened. Efficacy was evaluated and compared by log-rank. Results: Fifty-nine fit patients were treated with firstline FIr-B/FOx: wild-type, 31 (53%); mutant, 28 (47%). KRAS mutations: c.35 G > A, 15 (25.4%); c.35 G > T, 7 (11.8%); c.38 G > A, 3 (5%); other, 3 (5%). Wild-type compared to mutant status did not significantly affect PFS (14 and 11 months) and OS (38 and 20 months). c.35 G > A mutant showed: no significantly different PFS compared to wild-type; significantly worse OS (14 months) compared to wild-type and to other mutant (39 months). After progression, wild-type compared with mutant did not show significantly different PFS (10 months equivalently) nor OS (17 and 12 months); c.35 G > A compared to wild-type and/or other mutant showed significantly worse PFS and OS. Among 39 unfit patients, treated with conventional regimens, wild-type compared to mutant significantly affected PFS (8 and 6 months), and not OS (13 and 8 months); c.35 G > A mutation significantly affected worse PFS and OS compared to wild-type and/or other mutant. Conclusions: KRAS genotype does not significantly affect clinical outcome of MCRC patients treated with first line FIr-B/FOx, while mutant status may significantly affect worse PFS in patients unfit for intensive treatments. c.35 G > A mutant status, compared to wild type and other mutant, may significantly affect worse clinical outcome in MCRC patients treated with first line intensive FIr-B/FOx and after progression, as well as in patients unfit for FIr-B/FOx, due to increased biological aggressiveness. It did not affect worse PFS only in patients treated with FIr-B/FOx, probably due to effectiveness of triplet chemotherapy plus VEGF inhibitor.

Twenty patients with locally advanced cervical cancer (FIGO stage Ib-IIa “bulky”/VIIb) were treated with three courses of weekly PVB (day 1: cisplatin, 50 mg/m2; vincristin, 1 mg/m2; bleomycin, 30 IU over 24-hr) in a neoadjuvant setting. Toxicity was generally mild (no grade 3-4 toxicity was observed), and the treatment was well tolerated without reduction of programmed dose intensity. Fourteen patients (70%) experienced a clinical response and underwent surgery within 20 days after the third course of chemotherapy. Six patients (30%) with stable disease were treated with salvage radiotherapy. Two of the 14 responders experienced a pathologic complete response (14.2%); microscopic disease was detected in one patient with clinical complete response. Pelvic node metastases were found in 4/14 patients (28.5%) and microscopic parametrium involvement in 3/14 (21.4%). All 14 patients had free margins of resection. A short-term weekly platinum-based chemotherapy is highly effective, has little toxicity, and allows a prompt salvage therapy for nonresponding patients.

Cisplatin-based drug combinations are now currently investigated in an attempt to surpass the results obtainable by 5-FU alone or by 5-FU-containing regimens. We evaluated 29 patients with advanced and measurable gastric adenocarcinoma treated with etoposide, epirubicin and cisplatin. One patient had complete response (3 ± 7%); 9 patients had partial response (31 ± 17%); 5 patients (17±14%) were considered stable (S) and 14 (48 ± 18%) were classified as progressive. The average survival time for the entire group was 11 months with 25% of the patients alive at 24 months. We feel that cisplatin-containing regimens are promising and deserve further investigations. To clearly explore the potential innovative role of these regimens randomized trials versus FAM or 5-FU alone are mandatory.

Background: Prognosis of MCRC patients treated with bevacizumab and chemotherapy is not significantly different according to KRAS genotype. Specific mutations confer different aggressiveness. In vitro, codon 12 mutations increase aggressiveness by differential KRAS pathways regulation. Different codon 12 mutations modify biological activity of mutated KRAS proteins. Biological aggressiveness of codon 12 mutant tumors confers worse clinical behavior. RAS/RAF/MEK/ERK pathway increases VEGF expression and represses negative angiogenesis regulators; its aberrations stimulate angiogenesis and influence response to anti-VEGF. Prognostic role of prevalent c.35 G>A KRAS mutation in first line FIr-B/FOx, post-progression, and in patients unfit for FIr-B/FOx was evaluated. Methods: KRAS codon 12/13 were screened by SNaPshot and/or sequencing. Efficacy was evaluated and compared according to KRAS genotype, using log-rank. Results: At 21.5 months follow-up, 59 fit MCRC patients were treated with first line FIr-B/FOx: wild-type, 31 (53%); mutant, 28 (47%). KRAS mutations: c.35 G>A (G12D), 15 (25.4%); c.35 G>T, 7 (11.8%); c.38 G>A, 3 (5%); other, 3 (5%). KRAS genotype wild-type compared to mutant did not significantly affect PFS (14 and 11 months) and OS, even if OS seems favorable in wild-type (38 and 20 months). PFS of c.35 G>A mutant compared to wild-type was not significantly different. OS was significantly worse in c.35 G>A patients (14 months) compared to: wild-type (p = 0.002); other mutant (39 months) (p = 0.05). Wild-type compared with mutant did not show significantly different PFS (10 and 10 months) nor OS (17 and 12 months), after progression. PFS and OS after progression were significantly worse in c.35 G>A compared to wild-type and/or other mutant patients. Among patients unfit for FIr-B/FOx, and treated with conventional medical treatments, KRAS genotype wild-type compared to mutant significantly affected PFS (8 and 6 months), while not OS (13 and 8 months). c.35 G>A mutation significantly affected worse PFS and OS compared to wild-type and/or other mutations. Conclusions: KRAS genotype, wild-type compared to mutant, did not significantly affect prognosis in MCRC patients treated with first line FIr-B/FOx, while it significantly affected worse efficacy (PFS) in patients unfit for intensive medical treatments. KRAS c.35 G>A mutant status compared to wild type and other mutant, may significantly affect worse prognosis of patients, treated with first line intensive regimen (FIr-B/FOx), after progression, or unfit for FIr-B/FOx, probably due to increased biological aggressiveness. It did not affect worse PFS only in patients treated with first line FIr-B/FOx, probably due to effectiveness of intensive medical treatment consisting of triplet chemotherapy and VEGF inhibitor. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C35. Citation Format: Gemma Bruera, Katia Cannita, Edoardo Alesse, Corrado Ficorella, Enrico Ricevuto. Different prognostic relevance of KRAS genotype in metastatic colorectal cancer (MCRC) patients fitting for triplet chemotherapy plus VEGF-inhibitor, unfit for intensive medical treatment and post-progression. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C35.

e14596 Background: Prognosis of MCRC patients (pts) treated with bevacizumab (BEV) and chemotherapy (CT) is not significantly different according to KRAS genotype. Specificmutations confer different aggressiveness. Prognostic relevance of the prevalent c.35 G > A KRAS mutation was retrospectively evaluated in pts treated with first line FIr-B/FOx, and post-progression. Methods: KRAS codon 12/13 and BRAF mutations were screened by SNaPshot and/or sequencing. FIr-B/FOx: weekly 2 days/12h-timed-flat-infusion/5-fluorouracil 900 mg/m2, weekly alternating irinotecan 160 mg/m2/BEV 5 mg/kg, or oxaliplatin 80 mg/m2. Second-line treatment was selected according to fitness, KRAS genotype, previous efficacy and safety. Efficacy was evaluated and compared according to treatment or KRAS genotype, using log-rank. Results: At 21.5 months, 59 pts were evaluated. KRAS mutant pts: c.35 G > A, 15 (25.4%); c.35 G > T, 7 (11.8%); c.38 G > A, 3 (5%); other, 3 (5%). KRAS wild-type, 31 pts (52.7%). ORR, PFS, OS were, respectively: c.35 G > A, 71%, 9 months, 14 months; other mutants, 61%, 12 months, 39 months. OS was significantly worse in c.35 G > A pts compared to KRAS wild-type (p = 0.002), KRAS/BRAFwild-type (p = 0.03), other pts (p = 0.002), other mutants (p = 0.05), other codon 12 (p = 0.03) mutant pts; PFS was not significantly different. Post-progression, at 14 months follow-up, PFS and OS were significantly worse in c.35 G > A compared to wild-type and/or other mutant pts. ORR of second line treatments was 38%, metastasectomies 12.5%, PFS 10 months, OS 14 months. PFS and OS were significantly favourable in pts treated with triplet CT plus targeted agent compared to triplet, respectively: PFS 13 months versus 8 months; OS not reached versus 11 months. Conclusions: KRAS c.35 G > A mutation may significantly affect worse OS of MCRC pts. It does not significantly affect worse PFS of intensive first line FIr-B/FOx, while after progression significantly affect worse efficacy of second line treatments. Re-challenge of intensive regimens may affect significantly favourable PFS and OS.