Quantitative copy number analysis by Multiplex Ligation-dependent Probe Amplification (MLPA) of BRCA1-associated breast cancer regions identifies BRCAness

Our group has previously employed array Comparative Genomic Hybridization (aCGH) to assess the genomic patterns of BRCA1-mutated breast cancers. We have shown that the so-called BRCA1-like(aCGH) profile is also present in about half of all triple-negative sporadic breast cancers and is predictive for benefit from intensified alkylating chemotherapy. As aCGH is a rather complex method, we translated the BRCA1(aCGH) profile to a Multiplex Ligation-dependent Probe Amplification (MLPA) assay, to identify both BRCA1-mutated breast cancers and sporadic cases with a BRCA1-like(aCGH) profile. The most important genomic regions of the original aCGH based classifier (3q22-27, 5q12-14, 6p23-22, 12p13, 12q21-23, 13q31-34) were mapped to a set of 34 MLPA probes. The training set consisted of 39 BRCA1-like(aCGH) breast cancers and 45 non-BRCA1-like(aCGH) breast cancers, which had previously been analyzed by aCGH. The BRCA1-like(aCGH) group consisted of germline BRCA1-mutated cases and sporadic tumours with low BRCA1 gene expression and/or BRCA1 promoter methylation. We trained a shrunken centroids classifier on the training set and validation was performed on an independent test set of 40 BRCA1-like(aCGH) breast cancers and 32 non-BRCA1-like(aCGH) breast cancer tumours. In addition, we validated the set prospectively on 69 new triple-negative tumours. BRCAness in the training set of 84 tumours could accurately be predicted by prediction analysis of microarrays (PAM) (accuracy 94%). Application of this classifier on the independent validation set correctly predicted BRCA-like status of 62 out of 72 breast tumours (86%). Sensitivity and specificity were 85% and 87%, respectively. When the MLPA-test was subsequently applied to 46 breast tumour samples from a randomized clinical trial, the same survival benefit for BRCA1-like tumours associated with intensified alkylating chemotherapy was shown as was previously reported using the aCGH assay. Since the MLPA assay can identify BRCA1-deficient breast cancer patients, this method could be applied both for clinical genetic testing and as a predictor of treatment benefit. BRCA1-like tumours are highly sensitive to chemotherapy with DNA damaging agents, and most likely to poly ADP ribose polymerase (PARP)-inhibitors. The MLPA assay is rapid and robust, can easily be multiplexed, and works well with DNA derived from paraffin-embedded tissue

Amorphous aluminosilicate scaling characterization in a reverse osmosis membrane

This paper describes the results of experiments performed in a high-recovery system to elucidate the silica scaling phenomenon and characterize the scaling. In this research, cation exchange pretreatment is used to reduce Ca2+, Ba2+, and Mg2+ levels to prevent scaling during subsequent nanofiltration (NF) and reverse osmosis (RO) filtration, in which RO is fed with NF concentrate. In a pilot plant, a series of experiments were carried out at a total (NF + RO) recovery of 91, 94, 96 and 98% with locally available tap water as feed water. Autopsy studies were performed with the RO membranes after each experiment. The fouling layer was studied using SEM-EDX, ATR-FTIR and fouling extraction to determine the structure and the composition of the fouling deposits. A thin dense fouling layer was observed, which covered approximately half of the membrane surface, after operating for 20 days at 91 and 94% recovery. At 96 and 98% recovery, the fouling layer was thicker and completely covered the membrane surface. The scaling layer was mainly composed of Si, Al, Fe and O. The amount of Si increased with increasing recovery. To work at these high recoveries for an extended period, further measures need to be taken to prevent silica scaling