Molecular characterization and clinical interpretation of BRCA1/BRCA2 variants in families from Murcia (south-eastern Spain) with hereditary breast and ovarian cancer: clinical-pathological features in BRCA carriers and non-carriers.

This is the first study performed in Murcia (south-eastern Spain) in which 592 families with hereditary breast and ovarian cancer were identified thanks to Genetic Counselling Units from this area over 6 years. Diagnostic performance was 18.1% and 194 different genetic variants were obtained. Variants with uncertain significance accounted for only 5.6% of the total number of reports, so our population has been well characterised. In BRCA1 gene, two novel variants were found (c.1859delT and c.3205C > T) and the most frequently detected mutations were c.68_69delAG, c.212 + 1G > A, c.5123C > A, c.211A > G and c.1918C > T, which together represented 56.67% of total pathogenic mutations. In BRCA2 gene, four recurrent variants were described (deletion of entire exon 2, c.9117G > A, c.3264dupT and c.3455T > G) representing 43.5% of the mutations in this gene. Mutation c.68_69delAG and deletion of entire exon 2 in BRCA1 and BRCA2 genes respectively were the most prevalent variants in our population. Regarding the genotype-phenotype relation, mutation c.212 + 1G > A appeared in an important percentage of breast and ovarian cancer cases, c.5123C > A in bilateral breast cancer and c.9117G > A in bilateral breast cancer and ovarian cancer. With respect to clinical-pathological characteristic, BRCA1/BRCA2 mutation carriers showed earlier onset age of breast tumour and higher risk of developing contra lateral breast cancer than non-informative cases. Moreover, association between either molecular subtype triple negative breast cancer or ovarian cancer and BRCA1 carriers was obtained.

Second trimester angiotensing-converting enzyme and uterine artery Doppler as predictors of preeclampsia in a high-risk population.

OBJECTIVE: To investigate whether serum angiotensing-converting enzyme (ACE) and uterine artery Doppler (UAD) are useful markers as predictors of preeclampsia (PE) in a high-risk population. METHODS: Patients at risk of PE (n = 68) were subclassified as having PE (n = 8) or no PE (n = 60). Blood samples were obtained between 19 and 22 weeks of gestation. Doppler ultrasound of the uterine arteries was done at the time of blood sampling. Maternal serum ACE was determined through spectrophotometry assay (A15 Biosystem, ATOM, Barcelona, Spain). RESULTS: Comparing the group who presented PE with the one who was not developed it, we found significant differences for ACE (54.2 ± 21.2, 38.1 ± 12.3 U/L; p = 0.003); the pulsatility index (PI) (1.2 ± 0, 3.1 ± 0.3; p = 0.032) and resistance index (RI) (0.7 ± 0.1, 0.5 ± 0.1; p = 0.004). The AUC for ACE was 0.724, so we selected the cutoff of 36.5 U/L (sensitivity: 62.5% and specificity: 86.7%). The AUC for PI was 0.652 choosing a cutoff of 1.4 (sensitivity: 57.1% and specificity: 93.1%). The AUC for RI was 0.712 and the cutoff of 0.7 (sensitivity of 71.4% and specificity: 89.6%). The combination that allowed us to increase the diagnostic performance was the ACE+RI with Doppler study, increasing the AUC to 0.872. CONCLUSIONS: ACE, PI and RI as parameters of Doppler study were useful predictors of PE in the second trimester of gestation. The best combination to increase the diagnostic performance was ACE with the RI.

Placental growth factor, soluble fms-like tyrosine kinase 1 and progesterone as diagnostic biomarkers for ectopic pregnancy and missed abortion.

OBJECTIVE: The aim of this study is to investigate if progesterone, placental growth factor (PlGF) and soluble fms-like tyrosine kinase-1 (sFlt1) serum levels are useful markers to differentiate between ectopic pregnancy (EP), missed abortion (MA) and viable intrauterine implantation pregnancy (IUP). METHODS: We designed a retrospective case-control study which included 100 pregnant women (50 EP and 50 MA) at 6-8 weeks of gestation with ßhCG serum levels between 800 and 3500 UI/L and a viable IUP group. Progesterone, PlGF and sFlt-1 levels were measured with an electrochemiluminescence assay (Roche Diagnostics, Manheim, Alemania). A non parametric test was used to compare the markers in the different groups and we used receiver operating characteristic (ROC) curve analysis to calculate the area under the curve (AUC). RESULTS: When we compared the EP group with the MA group, we didn't find significant differences for PlGF (15.1[13.2-17.4]/16.7[12.8-18.7] pg/mL) (p=0.275). We only obtained significant differences for progesterone (9.1[3.1-16.8]/2.6[1.3-6.1] ng/mL) (p<0.001) and sFlt-1 (84[65-96]/126[94-256] pg/mL) (p<0.001). The AUC for progesterone was 0.756 and the cutoff point with better sensitivity and lower false positive rate was 6 ng/mL (sensitivity=60%, specificity=72.7%). The AUC for sFlt-1 was 0.842 and the cutoff point was 93 pg/mL (sensitivity=84.5%, specificity=86.3%). The combination of both markers allowed us to increase the AUC to 0.910. CONCLUSIONS: In conclusion, the present study suggests that sFlt-1 could be a useful marker to differentiate between an EP or a MA when ßhCG levels are similar in both groups. The combination of sFlt-1 with progesterone helps to increase the diagnostic performance.

Performance evaluation of biochemical and ultrasound markers for the screening of chromosomal numerical abnormalities in the first trimester of pregnancy.

BACKGROUND: To estimate the effectiveness of the first-trimester combined screening test in our population, departing from the results of diagnostic sensitivity and false positive rate (FPR), and checking some important parameters in prenatal screening. METHODS: The test was evaluated on 14250 pregnant women. The following variables were studied: the number of invasive techniques and the reasons for using such techniques, newborns with chromosomal abnormalities, total number of births, variation of biochemical markers throughout the gestational weeks, and MoM (multiple of the median) for biochemical and ultrasound markers. RESULTS: An important coverage and a decreased number of invasive techniques were obtained. For our population of pregnant women, the best gestational week to determine free beta-hCG and PAPP-A would be week 11 in which the best discrimination was found between affected and non affected fetuses for the three trisomies researched. We propose the cut-off 1/350, because it is the best one to increase sensitivity without exceeding the 5% FPR. CONCLUSIONS: Combined screening should be offered to pregnant woman, preferentially at week 11. Although different cut-offs for this prenatal test have been recommended by scientific societies, biochemical laboratories should set their own cut-off for getting the best sensitivity and FPR results. There should be a good level of collaboration between the laboratory and each participating ultrasound unit in order to ensure an optimal use of the first-trimester combined screening test.

Analysis of cholinesterases in human prostate and sperm: implications in cancer and fertility.

Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are postulated to play non-cholinergic roles in cellular physiology. The probable implication of cholinesterases (ChEs) in several human pathologies prompted us to study the cholinergic components in the male reproductive system. Surgical pieces of prostatic cancer (PC) and benign prostatic hyperplasia (BPH) were analyzed for AChE and BChE activity. Loosely (S1) and tightly (S2) bound AChE and BChE forms were characterized by sedimentation analysis. The mean AChE activity in BHP samples was 2.38+/-0.56 mU/mg (nmol of the substrate hydrolysed per minute and per milligram protein) and 2.57+/-0.61 mU/mg in S1 and S2, respectively. The AChE activity did not vary with cancer, showing 2.46+/-0.45 mU/mg in S1 and 2.70+/-0.53 mU/mg in S2 from PC samples. Amphiphilic dimers and monomers and hydrophilic dimers of AChE were identified in BHP and PC tissues. Their contribution was affected by cancer with a great increase in hydrophilic dimers in the cancerous samples. Significant levels of both AChE and BChE activities were found in seminal fluid and homogenates from spermatozoids. Enzymatic activity dropped in samples with abnormal seminal parameters as sperm count and mobility.

Cancer-associated differences in acetylcholinesterase activity in bronchial aspirates from patients with lung cancer.

In non-neuronal contexts, ACh (acetylcholine) is thought to be involved in the regulation of vital cell functions, such as proliferation, differentiation, apoptosis and cell-cell interaction. In airways, most cells express the non-neuronal cholinergic system, each containing a specific set of components required for synthesis, signal transduction and ACh hydrolysis. The aim of the present study was determine the expression of cholinergic system components in bronchial aspirates from control subjects and patients with lung cancer. We conducted an analysis of cholinergic components in the stored soluble and cellular fraction of bronchial aspirates from non-cancerous patients and patients diagnosed with lung cancer. The results show that the fluid secreted by human lung cells contains enough AChE (acetylcholinesterase) activity to control ACh levels. Thus these findings demonstrate that: (i) AChE activity is significantly lower in aspirates from squamous cell carcinomas; (ii) the molecular distribution of AChE in both bronchial cells and fluids consisted of amphiphilic monomers and dimers; and (iii) choline acetyltransferase, nicotinic receptors and cholinesterases are expressed in cultured human lung cells, as demonstrated by RT-PCR (reverse transcriptase-PCR). It appears that the non-neuronal cholinergic system is involved in lung physiology and lung cancer. The physiological consequences of the presence of non-neuronal ACh will depend on the particular cholinergic signalling network in each cell type. Clarifying the pathophysiological actions of ACh remains an essential task and warrants further investigation.

Cholinesterase activity of human lung tumours varies according to their histological classification.

The probable involvement of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) in cancer and the relevance of cholinergic responses for lung cancer growth prompted us to study whether cholinesterase activity of human lung is altered by malignancy. Surgical pieces of non-small lung carcinomas (NSLC) and their adjacent non-cancerous tissues (ANCT) were analysed for AChE and BChE activities. AChE activity in adenocarcinoma (AC) was 7.80 +/- 5.59 nmol of substrate hydrolysed per min and per mg of protein (mU/mg), the same as in their ANCT (8.83 +/- 4.72 mU/mg; P = 0.823); in large cell carcinoma (LCC), 7.52 +/- 3.32 mU/mg, approximately 50% less than in their ANCT (15.39 +/- 5.66 mU/mg; P = 0.043); and in squamous cell carcinoma (SCC), 1.39 +/- 0.58 mU/mg, 80% less than in ANCT (6.08 +/- 2.88 mU/mg; P = 0.003). BChE activity was 5.85 +/- 3.20 mU/mg in AC and 9.56 +/- 3.38 mU/mg in ANCT (P = 0.022); 2.94 +/- 2.01 mU/mg in LCC and 6.50 +/- 6.63 mU/mg in ANCT (P = 0.068); and 4.49 +/- 2.30 mU/mg in SCC and ANCT 6.56 +/- 4.09 mU/mg (P = 0.026). Abundant AChE dimers and fewer monomers were identified in lung and, although their distribution was unaffected by cancer, the binding with concanavalin A revealed changes in AChE glycosylation between SCC and their ANCT. The fall in BChE activity affected all molecules, with a strong decrease of the amphiphilic tetramers. Western blotting revealed protein bands with the expected mass of the principal AChE subunits, and the deeper intensity of the protein signal in SCC than in healthy lung, in lanes loaded with the same units of AChE activity, supported an augment in the amount of AChE protein/unit of AChE activity in SCC. The increased availability of acetylcholine in neoplastic lung, resulting from the fall of cholinesterase activity, may enhance cholinergic signalling and contribute to tumour progression.

[Biochemical bone markers and bone mass study in recently postmenopausal women without osteoporosis]. / Marcadores bioquímicos de remodelado en el estudio de la masa ósea en la mujer con menopausia reciente sin osteoporosis.

BACKGROUND AND OBJECTIVE: To date, there have not been performed enough studies in Spain to analyze the usefulness of biochemical bone markers in postmenopausal and non-osteoporotic women. POPULATION AND METHOD: We studied several groups of women, ages between 17 and 30 years, 30-45 years, premenopausal and recently postmenopausal in order to know the reference values of some biochemical bone markers along 2 years in premenopausal and postmenopausal women (postmenopausal divided in 2 groups: those under treatment with ordinary hormonal replacement therapy [HRT], and those without it). RESULTS: Values of biochemical bone markers were higher in postmenopausal and in 17-30 years group than in premenopausal and 30-45 years group (p < 0.05). Absolute values and the change in bone markers (%) were higher in postmenopausal and non-osteoporotic women under HRT than in those without HRT (% change: OC, 35,72 vs 4.45 [p < 0.01]; ALPo, 13,13 vs 27.06 [p < 0.01]; NTx, 42.74 vs 10.93 [p < 0.01]; fDPD, 21.07 vs 28.49 [p < 0.05]; betaCrossLaps, 50,17 vs 23.04 [p < 0.01]). CONCLUSIONS: We have defined reference values in our region and have proved that biochemical bone markers (absolute values and their change from the basal value) represent a useful tool in monitoring hormonal replacement therapy in recently postmenopausal and non-osteoporotic women.

Acetylcholinesterase biogenesis is impaired in lung cancer tissues.

Studies cited by Cowan et al. [J. Appl. Toxicol. 23, 177 (2003)] indicate existence of inflammatory and cholinergic pathways in both nerve agents and sulfur mustard (HD) injury. Increase in AChE synthesis and neurite extension was noted after exposure to HD [K.W. Lanks et al., Exp. Cell Res. 355 (1975)]. Moreover, anti-inflammatory drugs reduce the dermal, respiratory and ocular damage caused by exposure to HD. On the other hand, recent studies have noted the involvement of neuro-inflammatory processes during exposure to the nerve agents sarin or soman [Cowan et al., 2003]. The use of various anti-inflammatory drugs in addition to the classical antidotal drugs (e.g. atropine and oximes) caused decrease in certain toxic symptoms and inflammation-induced brain damage. Our new bifunctional drugs (Scheme 1) are based on CNS-permeable molecular combination of pseudo-reversible AChE inhibitor (pyridostigmine, PYR) coupled via a hydrophobic spacer (octyl or decyl hydrocarbon chain) to a non-steroidal anti-inflammatory drug (NSAID) such as Ibuprofen or Diclofenac (Scheme 1). This study evaluates the efficacy of certain bifunctional compounds against HD and soman poisoning in mice in vivo.

[Addition of an angiotensin II receptor blocker to maximal dose of ACE inhibitors in heart failure]. / Adición de un ARAII al tratamiento con dosis máximas de IECA en la insuficiencia cardíaca.

In heart failure, the benefits of adding angiotensin-receptor blockade to ACE inhibitors have been studied only with submaximal doses of ACE inhibitors. We included 20 patients (LVEF 24 7%, NYHA II-III), with no clinical or therapeutic variations in the previous three months, who were receiving maximal doses of ACE inhibitors. We added losartan 50 mg once a day. At six months, SBP decreased (115 8 vs. 106 9 mmHg; p = 0.001), LVEF increased (24.4 7 vs. 34.1 7%; p < 0.001), ventricular end-diastolic volumes decreased (220 58 vs 190 46 ml; p = 0.007), and SPAP decreased (43 8 vs. 35 7 mmHg; p < 0.001). Seven patients improved one degree on the NYHA scale (p = 0.004), but VO2max did not change (20.8 5.2 vs. 21.8 5.0 ml/kg/min, p = 0.120). Plasma levels of norepinephrine, at rest and maximal exercise, brain natriuretic peptide, and renin were similar. After maximum ACE inhibitor doses, the addition of losartan is safe and associated with an improvement in ventricular function and NYHA functional class, but with no change in neurohormonal status.