Assessment of the functional impact of germline BRCA1/2 variants located in non-coding regions in families with breast and/or ovarian cancer predisposition.

PURPOSE: The molecular mechanism of breast and/or ovarian cancer susceptibility remains unclear in the majority of patients. While germline mutations in the regulatory non-coding regions of BRCA1 and BRCA2 genes have been described, screening has generally been limited to coding regions. The aim of this study was to evaluate the contribution of BRCA1/2 non-coding variants. METHODS: Four BRCA1/2 non-coding regions were screened using high-resolution melting analysis/Sanger sequencing or next-generation sequencing on DNA extracted from index cases with breast and ovarian cancer predisposition (3926 for BRCA1 and 3910 for BRCA2). The impact of a set of variants on BRCA1/2 gene regulation was evaluated by site-directed mutagenesis, transfection, followed by Luciferase gene reporter assay. RESULTS: We identified a total of 117 variants and tested twelve BRCA1 and 8 BRCA2 variants mapping to promoter and intronic regions. We highlighted two neighboring BRCA1 promoter variants (c.-130del; c.-125C > T) and one BRCA2 promoter variants (c.-296C > T) inhibiting significantly the promoter activity. In the functional assays, a regulating region within the intron 12 was found with the same enhancing impact as within the intron 2. Furthermore, the variants c.81-3980A > G and c.4186-2022C > T suppress the positive effect of the introns 2 and 12, respectively, on the BRCA1 promoter activity. We also found some variants inducing the promoter activities. CONCLUSION: In this study, we highlighted some variants among many, modulating negatively the promoter activity of BRCA1 or 2 and thus having a potential impact on the risk of developing cancer. This selection makes it possible to conduct future validation studies on a limited number of variants.

Breast and ovarian cancer predisposition due to de novo BRCA1 and BRCA2 mutations.

BRCA1 and BRCA2 are the two major genes predisposing to breast and ovarian cancer. Whereas high de novo mutation rates have been demonstrated for several genes, only 11 cases of de novo BRCA1/2 mutations have been reported to date and the BRCA1/2 de novo mutation rate remains unknown. The present study was designed to fill this gap based on a series of 12 805 consecutive unrelated patients diagnosed with breast and/or ovarian cancer who met the inclusion criteria for BRCA1/2 gene analysis according to French guidelines. BRCA1/2 mutations were detected in 1527 (12%) patients, and three BRCA1 mutations and one BRCA2 mutation were de novo. The BRCA1/2 de novo mutation rate was estimated to be 0.3% (0.1%; 0.7%). Although rare, it may be useful to take the possibility of de novo BRCA1/2 mutation into account in genetic counseling of relatives and to improve the understanding of complex family histories of breast and ovarian cancers.

Tracking camera control in endoscopic dacryocystorhinostomy surgery.

OBJECTIVES: Poor camera control during endoscopic dacryocystorhinostomy (EnDCR) surgery can cause inadequate visualisation of the anatomy and suboptimal surgical outcomes. This study investigates the feasibility of using computer vision tracking in EnDCR surgery as a potential formative feedback tool for the quality of endoscope control. DESIGN: A prospective cohort analysis was undertaken comparing junior versus senior surgeons performing routine EnDCR surgery. Computer vision tracking was applied to endoscopic video footage of the surgery: Total number of movements, camera path length in pixels and surgical time were determined for each procedure. A Mann-Whitney U-test was used to test for a significant difference between juniors and seniors (P < 0.05). SETTING: Operating theatre. PARTICIPANTS: Ten junior surgeons (<20 completed procedures) and 10 senior surgeons (>100 completed procedures). MAIN OUTCOME MEASURES: Total number of movements of the endoscope per procedure. Path length of the endoscope per procedure. RESULTS: Twenty videos, 10 from junior surgeons and 10 from senior surgeons were analysed. Feasibility of our tracking system was demonstrated. Mean camera path lengths were significantly different at 119,329px (juniors) versus 43,697px (seniors), P ≪ 0.05. The mean number of movements was significantly different at 9134 (juniors) versus 3690 (seniors), P ≪ 0.05. These quantifiable differences demonstrate construct validity for computer vision endoscope tracking as a measure of surgical experience. CONCLUSIONS: Computer vision tracking is a potentially useful structured and objective feedback tool to assist trainees in improving endoscope control. It enables juniors to examine how their pattern of endoscope control differs from that of seniors, focusing in particular on sections where they are most divergent.

Rare germline large rearrangements in the BRCA1/2 genes and eight candidate genes in 472 patients with breast cancer predisposition.

Hereditary breast cancers account for up to 5-10 % of breast cancers and a majority are related to the BRCA1 and BRCA2 genes. However, many families with breast cancer predisposition do not carry any known mutations for BRCA1 and BRCA2 genes. We explored the incidence of rare large rearrangements in the coding, noncoding and flanking regions of BRCA1/2 and in eight other candidate genes--CHEK2, BARD1, ATM, RAD50, RAD51, BRIP1, RAP80 and PALB2. A dedicated zoom-in CGH-array was applied to screen for rearrangements in 472 unrelated French individuals from breast-ovarian cancer families that were being followed in eight French oncogenetic laboratories. No new rearrangement was found neither in the genomic regions of BRCA1/2 nor in candidate genes, except for the CHEK2 and BARD1 genes. Three heterozygous deletions were detected in the 5' and 3' flanking regions of BRCA1. One large deletion introducing a frameshift was identified in the CHEK2 gene in two families and one heterozygous deletion was detected within an intron of BARD1. The study demonstrates the usefulness of CGH-array in routine genetic analysis and, aside from the CHEK2 rearrangements, indicates there is a very low incidence of large rearrangements in BRCA1/2 and in the other eight candidate genes in families already explored for BRCA1/2 mutations. Finally, next-generation sequencing should bring new information about point mutations in intronic and flanking regions and also medium size rearrangements.

Italian consensus statement on management of HIV-infected individuals with advanced disease naïve to antiretroviral therapy.

BACKGROUND: Individuals with advanced HIV infection naïve to antiretroviral therapy represent a special population of patients frequently encountered in clinical practice. They are at high risk of disease progression and death, and their viroimmunologic response following the initiation of highly active antiretroviral therapy may be more incomplete or slower than that of other patients. Infection management in such patients can also be complicated by underlying conditions, comorbidities, and the need for concomitant medications. AIM: To provide practical guidelines to those clinicians providing care to HIV-infected patients in terms of diagnostic assessment, monitoring, and treatment. CONCLUSIONS: The principals of antiretroviral treatment in asymptomatic naïve patients with advanced HIV infection are the same as those applicable to the general population with asymptomatic HIV infection. Naïve patients with advanced HIV infection and a history of AIDS-defining illnesses urgently need antiretroviral treatment, with the choice of antiretroviral regimen and timetable based on such factors as concomitant treatment and prophylaxis, drug interactions, and potential concomitant drug toxicity. Finally, an adequate counseling program - both before and after HIV-testing - that includes aspects other than treatment adherence monitoring is a crucial step in disease management.

[Functional outcome and quality of life after pancreaticoduodenectomy]. / Séquelles fonctionnelles et qualité de vie après duodénopancréatectomie céphalique.

AIM OF STUDY: Pancreaticoduodenectomy is a major surgical procedure whose physiological effects may weigh heavily on quality of life. The goal of this retrospective unicentric pilot study was to assess the the functional outcome after pancreaticoduodenectomy and its effect on the patient's quality of life. PATIENTS AND METHOD: Thirty patients free from tumor recurrence more than one year after pancreaticoduodenectomy responded to the GIQLI questionnaire (Gastro Intestinal Quality of Life Index) and to a specific questionnaire evaluating long-term functional outcome. RESULTS: The acceptability rate was 100%. The internal coherence of the GIQLI questionnaire was good (a Cronbach rate=0.85). The average total score of the GIQLI was 94 (IC-95%=[86-101]) compared to an ideal rate of 144. The quality of life was significantly impaired by steatorrhea, need for treatment of diarrhea, or need for enzymatic substitutive treatment. CONCLUSION: Compared to the reference for the normal population, patients post-pancreaticoduodenectomy have an average 25% decrease of quality of life scores (although more than 25% of patients experience a normal quality of life). The impairment of quality of life after pancreaticoduodenectomy appears to be related to the functional digestive consequences of the procedure. The GIQLI score could be used to assess the technical surgical variants.

First Italian consensus statement on diagnosis, prevention and treatment of cardiovascular complications in HIV-infected patients in the HAART era (2006).

The present document contains recommendations for assessment, prevention and treatment of cardiovascular risk for HIV-infected patients. All recommendations were graded according to the strength and quality of the evidence and were voted on by 73 members of the Italian Cardiovascular Risk Guidelines Working Group which includes both experts in HIV/AIDS care and in cardiovascular and metabolic medicine. Since antiretroviral drug exposure represents only one risk factor, continued emphasis on an integrated management is given. This should include prevention and treatment of known cardiovascular risk factors (such as dyslipidaemia, diabetes, insulin resistance, healthy diet, physical activity, avoidance of smoking), but also rational switch of antiretroviral drugs. A rational switch strategy should consider both metabolic and anthropometric disturbances and effectiveness of antiretroviral regimens.

Effects of coexisting hypertension and type II diabetes mellitus on arterial stiffness.

Hypertension (HT) is frequently associated with diabetes mellitus (DM) and its prevalence doubles in diabetics compared to the general population. This high prevalence is associated with increased stiffness of large arteries, which often precedes macrovascular events. The aim of our study was to evaluate the influence of HT and type II DM on aortic stiffness in patients with one disease or the other compared to those with both HT and type II DM. We studied 220 patients, 50 with type II DM (Group A), 50 with HT (Group B), 85 with both diseases (Group C), and 35 healthy subjects (HS). Regional arterial stiffness was assessed by automatic measurement of the carotid-femoral pulse wave velocity (PWV). For each patient, we evaluated: age, sex, body mass index, smoking habit, heart rate, SBP/DBP, pulse pressure (PP), mean BP, fasting glucose, lipid profile, uric acid, and fibrinogen. Group C had significantly more women and non smokers and the highest PP (61+/-14 mmHg). Of biochemical parameters, only fibrinogen was higher in Group A and in Group C (P<0.01 and P<0.001, respectively). Group C had a significantly higher PWV than the other four groups (P<0.0001). Stepwise forward regression analysis showed that fasting glucose was the first independent determinant of PWV (P<0.0001). In conclusion, this study shows that patients with DM and HT have higher arterial stiffness compared to HS and those with one disease or the other. Fasting glucose is the major independent determinant of PWV, which may be used as a relevant tool to assess the influence of cardiovascular risk factors on arterial stiffness in high-risk patients.

[Microcalcification and breast pathology: differential diagnosis and oncogenic risk assessment]. / Le microcalcificazioni nella patologia mammaria: diagnosi differenziale e valutazione del rischio oncogeno.

Breast cancer is one of the first cause of death in women. The investigation and the study of microcalcifications can discover about half of hidden breast carcinomas and this is an excellent help for the diagnosis. The aim of this study is to give some guidelines for a correct interpretation of these microcalcifications.

Chronic taurine supplementation ameliorates oxidative stress and Na+ K+ ATPase impairment in the retina of diabetic rats.

This study evaluates the effect of 4 months supplementation with 2% and 5% taurine (w/w) on the retina of diabetic rats. In non-diabetic rats, taurine does not modify glycemia, body weight, retinal conjugated dienes (CD), lipid hydroperoxide (LP), and Na(+)K(+)ATPase activity. In diabetic rat, at 2, 4, 8, 16 weeks following the onset of diabetes, retinal CD and LP are significantly and progressively increased, while pump activity is gradually and significantly reduced. In taurine supplemented diabetic rats, glycemia is not affected but lipid peroxidation is significantly decreased. Finally, taurine preserves ATPase activity being 5% more effective than 2% taurine. We conclude that taurine supplementation ameliorates biochemical retinal abnormalities caused by diabetes, thereby suggesting that taurine may have a role in the prevention of retinal changes in diabetes.

[Helicobacter pylori and gastric cancer. The incidence of infection in personal experience]. / Helicobacter pylori e carcinoma gastrico. Incidenza della colonizzazione batterica nella nostra esperienza.

BACKGROUND: Helicobacter pylori is an identified carcinogen for gastric cancer, but the underlying mechanisms remain to be defined. The aim of this study is to analyze the incidence of Hp infection in our series of patients with gastric carcinoma. METHODS: Between 1988 and 1998, 60 patients with diagnosis of gastric adenocarcinoma underwent partial or total gastrectomy. Forty-one were males and 19 females with an average age of 62 years (range 36-79). Twenty-seven cancers (45%) were localized in the lower third of the stomach, 17 (28%) in the middle third and eight (13%) in the upper third or cardias. In six patients (10%) the tumor was multicentric, while a recurrence on gastric stump after subtotal gastrectomy was present in two cases (3%). According to Lauren's criteria 39 cancers (65%) were of intestinal type, 16 (27%) of diffuse type and five (8%) of mixed type. The histologic preparations have been re-examined in order to verify the presence or not of Hp on gastric mucosa around neoplasm. RESULTS: Hp was found in 35 (58%) of the analyzed specimens and therefore a significant percentage of patients was Hp-positive at the time of diagnosis and surgery. Between 35 Hp-positive samples, 24 were adenocarcinomas of intestinal type, nine of diffuse type and two of mixed type, with a prevalence of Hp in intestinal type cancer. CONCLUSIONS: This study confirmed the high incidence of Hp infection in patients with gastric carcinoma, particularly in those with intestinal type cancer.

[Prognosis and treatment of cervix uteri carcinoma in pregnancy]. / Prognosi e trattamento del carcinoma della cervice uterina in gravidanza.

Carcinoma of the cervix is tha most common ginecologic cancer found during pregnancy. Management and treatment of this condition depend on cancer stage and estimated gestational age. Pregnancy represents an ideal time for cervical cancer screening, and all pregnant women presenting for prenatal care should be carefully examinated. The prognosis for pregnant women is similar to that for non-pregnant one. The aim of this study is to review the incidence, treatment and prognosis of this condition during pregnancy.

Inhibition of protein farnesylation enhances the chemotherapeutic efficacy of the novel geranylgeranyltransferase inhibitor BAL9611 in human colon cancer cells.

Proteins belonging to the ras superfamily are involved in cell proliferation of normal and neoplastic tissues. To be biologically active, they require post-translational isoprenylation by farnesyl-transferase and geranylgeranyl-transferase. Enzyme inhibition by drugs may thus represent a promising approach to the treatment of cancer. Therefore, the combined effect of BAL9611, a novel inhibitor of geranylgeranylation, and manumycin, a farnesyl-transferase inhibitor, was evaluated on the SW620 human colon cancer cell line which harbours a mutated K-ras gene. BAL9611 and manumycin dose-dependently inhibited SW620 cell growth with 50% inhibitory concentration (IC(50)) of 0.47 +/- 0.03 and 5.24 +/- 1.41 microM (mean +/- SE), respectively. The isobologram analysis performed at the IC(50)level revealed that the combined treatment was highly synergistic with respect to cell growth inhibition. BAL9611 and manumycin were able to inhibit the geranylgeranylation of p21rhoA and farnesylation of p21ras; both drugs inhibited p42ERK2/MAPK phosphorylation, but their combination was more effective than either drug alone. Moreover, the enhanced inhibition of cell growth in vitro by the BAL9611-manumycin combination was also observed in vivo in CD nu/nu female mice xenografted with SW620 tumours. Finally, both drugs were able to induce cell death by apoptosis in vitro and in vivo, as demonstrated by perinuclear chromatin condensation, cytoplasm budding and nuclear fragmentation, and interoligonucleosomal DNA digestion. In conclusion, the inhibition of protein farnesylation enhances the chemotherapeutic effect of BAL9611 in vitro and in vivo in a synergistic fashion, as a result of the impairment of post-translational isoprenylation of proteins and phosphorylation of p42ERK2/MAPK, whose activation is associated with post-translational geranylgeranylation and farnesylation of p21rhoA and p21ras.

Mucosal and systemic immune activation is present in human immunodeficiency virus-exposed seronegative women.

Immune parameters were analyzed in peripheral blood mononuclear cells (PBMC) and cervical mucosa biopsy specimens of human immunodeficiency virus (HIV)-seronegative women sexually exposed to HIV (exposed seronegative [ESN]), HIV-infected women, and healthy women without HIV exposure. HIV was not detected in PBMC or cervical mucosa biopsy specimens of ESN women. However, interleukin (IL)-6, IL-10, IL-12, interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha and -beta mRNA were elevated in PBMC and cervical mucosa biopsy specimens of ESN and HIV-infected women; CCR5 and CXCR4 mRNA were augmented in cervical mucosa biopsy specimens, but not in PBMC, of ESN and HIV-infected women; HIV-specific IFN-gamma-secreting cells were detected in vaginal washes of ESN and HIV-infected women; and phenotypic alterations were present in PBMC of ESN women. These results suggest that active HIV infection is not required for T cell activation; immune alterations occur in women in whom HIV infection cannot be detected virologically or clinically.

HIV-specific mucosal and cellular immunity in HIV-seronegative partners of HIV-seropositive individuals.

HIV-specific mucosal and cellular immunity was analyzed in heterosexual couples discordant for HIV status in serum and in HIV-unexposed controls. HIV-specific IgA but not IgG was present in urine and vaginal wash samples from HIV-exposed seronegative individuals (ESN), whereas both IgA and IgG were observed in their HIV-seropositive partners; antibodies were not detected in low-risk controls. Envelope protein (Env) peptide-stimulated interleukin-2 (IL-2) production by peripheral blood mononuclear cells (PBMCs) was detected in 9 out of 16 ESNs, 5 out of 16 HIV-infected patients and 1 out of 50 controls. Env peptide-stimulated PBMCs of ESNs produced more IL-2 and less IL-10 compared with those of HIV-infected individuals; no differences were observed in chemokine production or in CCR5 expression. These data demonstrate that a compartmentalized immune response to pathogens is possible in humans and raise the possibility of protective roles for cell-mediated immunity and mucosal IgA in HIV-seronegative individuals exposed to HIV.

Leucocytoclastic vasculitis associated with hepatitis C virus antibodies.

The actiopathogenesis of leucocytoclastic vasculitis is still unknown, but recently hepatitis C virus (HCV) has been suggested as trigger of autoimmunity. We report a case of a 26-yr-old patient with purpura due to leucocytoclastic vasculitis associated with hepatitis C virus infection. Laboratory findings showed AST, ALT, gamma GT within normal limits, positive antibodies to HCV (IIF and Riba II) and polymerase chain reaction for HCV RNA. Anti-nuclear antibodies, IgG and IgM anti-cardiolipin antibodies, anti-platelet antibodies and anti-neutrophil cytoplasmic antibodies with perinuclear pattern were also present. A skin biopsy specimen of a purpuric lesion showed leucocytoclastic vasculitis with small vessel thrombosis and perivascular deposition of IgM and fibrinogen on immunofluorescence study. This case shows a role of HCV in leucocytoclastic vasculitis; it is possible that this HCV can induce autoimmunity independently of cryoglobulins and liver involvement.