Proteomics and mathematical modeling of longitudinal CSF differentiates fast versus slow ALS progression

Objective: Amyotrophic lateral sclerosis (ALS) is a heterogeneous disease with a complex etiology that lacks biomarkers predicting disease progression. The objective of this study was to use longitudinal cerebrospinal fluid (CSF) samples to identify biomarkers that distinguish fast progression (FP) from slow progression (SP) and assess their temporal response.Methods: We utilized mass spectrometry (MS)-based proteomics to identify candidate biomarkers using longitudinal CSF from a discovery cohort of SP and FP ALS patients. Immunoassays were used to quantify and validate levels of the top biomarkers. A state-transition mathematical model was created using the longitudinal MS data that also predicted FP versus SP.Results: We identified a total of 1148 proteins in the CSF of all ALS patients. Pathway analysis determined enrichment of pathways related to complement and coagulation cascades in FPs and synaptogenesis and glucose metabolism in SPs. Longitudinal analysis revealed a panel of 59 candidate markers that could segregate FP and SP ALS. Based on multivariate analysis, we identified three biomarkers (F12, RBP4, and SERPINA4) as top candidates that segregate ALS based on rate of disease progression. These proteins were validated in the discovery and a separate validation cohort. Our state-transition model determined that the overall variance of the proteome over time was predictive of the disease progression rate.Interpretation: We identified pathways and protein biomarkers that distinguish rate of ALS disease progression. A mathematical model of the CSF proteome determined that the change in entropy of the proteome over time was predictive of FP versus SP

Metastatic melanoma and vemurafenib: novel approaches

Metastatic melanoma (MM) presents a treatment challenge to oncologists worldwide. Dacarbazine is the first line chemotherapy treatment for MM, though the overall response rates are very poor. Recently, the v-raf murine sarcoma viral oncogene homolog B1 (BRAF) V600 mutation was found to play a main role in MM. This mutation is present in 40-60% of melanoma patients. Vemurafenib is a BRAF kinase inhibitor that showed impressive results in phase I-III trials and was thus recently approved for the treatment of MM. This paper will briefly focus on vemurafenib in the treatment of MM and highlight concerns

Direct detection of a BRAF mutation in total RNA from melanoma cells using cantilever arrays.

Malignant melanoma, the deadliest form of skin cancer, is characterized by a predominant mutation in the BRAF gene. Drugs that target tumours carrying this mutation have recently entered the clinic. Accordingly, patients are routinely screened for mutations in this gene to determine whether they can benefit from this type of treatment. The current gold standard for mutation screening uses real-time polymerase chain reaction and sequencing methods. Here we show that an assay based on microcantilever arrays can detect the mutation nanomechanically without amplification in total RNA samples isolated from melanoma cells. The assay is based on a BRAF-specific oligonucleotide probe. We detected mutant BRAF at a concentration of 500 pM in a 50-fold excess of the wild-type sequence. The method was able to distinguish melanoma cells carrying the mutation from wild-type cells using as little as 20 ng µl(-1) of RNA material, without prior PCR amplification and use of labels

Collaborative study to establish a World Health Organization International genotype panel for parvovirus B19 DNA nucleic acid amplification technology (NAT)-based assays

BACKGROUND AND OBJECTIVES: The aim of the collaborative study was to evaluate a panel of plasma samples containing different genotypes of parvovirus B19 (B19V) for use in nucleic acid amplification technology (NAT)-based assays. MATERIALS AND METHODS: The panel of samples [Center for Biologics Evaluation and Research Parvovirus B19 Genotype Panel 1; National Institute for Biological Standards and Control (NIBSC) code number 09/110] comprises four different members, i.e. Member 1, Member 2, Member 3, and Member 4 (M1-M4); these represent genotypes 1, 2, 3a B19V, and a negative plasma control, respectively. Thirty-five laboratories from 13 different countries participated in the study. Participants assayed the panel members concurrently with the 2nd World Health Organization (WHO) International Standard for B19V DNA (NIBSC code 99/802) on four separate occasions. RESULTS: A total of 44 sets of data were returned, 34 from quantitative assays and 10 from qualitative assays. The majority of assays used were in-house and based on real-time PCR. The results showed that all three genotypes were detected consistently by the majority of participants, although a small number of assays detected genotypes 2 and 3 less efficiently, or not at all. Real-time stability studies have indicated that the panel of B19V samples is stable under normal conditions of storage, i.e. 64-70\ub0C. CONCLUSIONS: The four-member panel is intended for use in evaluating the ability of NAT assays to detect different B19V genotypes (M1-M3). Based on the results of the collaborative study, the panel was established as the 1st WHO International Reference Panel for parvovirus B19 genotypes