Real-time detection of BRAF V600E mutation from archival hairy cell leukemia FFPE tissue by nanopore sequencing

Abstract

The MinION is a miniaturized high-throughput next generation sequencing platform of novel conception. The use of nucleic acids derived from formalin-fixed paraffin-embedded samples is highly desirable, but their adoption for molecular assays is hurdled by the high degree of fragmentation and by the chemical-induced mutations stemming from the fixation protocols. In order to investigate the suitability of MinION sequencing on formalin-fixed paraffin-embedded samples, the presence and frequency of BRAF c.1799T > A mutation was investigated in two archival tissue specimens of Hairy cell leukemia and Hairy cell leukemia Variant. Despite the poor quality of the starting DNA, BRAF mutation was successfully detected in the Hairy cell leukemia sample with around 50% of the reads obtained within 2 h of the sequencing start. Notably, the mutational burden of the Hairy cell leukemia sample as derived from nanopore sequencing proved to be comparable to a sensitive method for the detection of point mutations, namely the Digital PCR, using a validated assay. Nanopore sequencing can be adopted for targeted sequencing of genetic lesions on critical DNA samples such as those extracted from archival routine formalin-fixed paraffin-embedded samples. This result let speculating about the possibility that the nanopore sequencing could be trustably adopted for the real-time targeted sequencing of genetic lesions. Our report opens the window for the adoption of nanopore sequencing in molecular pathology for research and diagnostics.

Cytokines secreted from bone marrow derived mesenchymal stem cells promote apoptosis and change cell cycle distribution of K562 cell line as clinical agent in cell transplantation

“…Cancer is one of the major causes of morbidity and mortality throughout the world. Unlike the various therapeutic strategies such as radiotherapy, chemotherapy, and surgery, these approaches are often limited by the recurrence of metastasis, drug resistance, off-target effects or complications caused by these methods [27]. With this explanation, stem cell-based therapy as an alternative treatment has attracted researchers and clinicians [28].…”

Protein-Catalyzed Capture Agents

“…Theranostic probes can be peptide based. 162,163 When equipped with one radiolabel, they are an in vivo positron emission tomography (PET) imaging probe. 164–166 Substitution of a second radiolabel doesn’t influence the targeting, but converts the same targeting molecule into a potent vehicle for site-directed radiotherapy.…”

iCrotoK-PseAAC: Identify lysine crotonylation sites by blending position relative statistical features according to the Chou’s 5-step rule

“…Chou’s peptide formulation [30] was used for an accurate elaboration of samples, which had generally utilized in computational biology and bioinformatics; for example, in predicting the signal peptide cleavage sites [31], nitrotyrosine sites [32], methylation sites [33], hydroxylysine and hydroxyproline sites [34], lysine ubiquitination sites [35], protein-protein binding sites [36], phosphorylation sites [37, 38], lysine succinylation sites [39], and, numerous lysine PTM sites [40].…”

“…To keep the crucial information safe regarding the protein samples’ sequence order, as it is necessary that the benchmark dataset of protein samples have the amino acid residues to be in a specific order. Different types of factors were considered while gathering the information of protein samples using different variables such as order of moments in which some of the moments for data evaluation and some for peculiarity of data and some for orientation from enormous amount of gathered protein samples and some of them were elaborated by the mathematicians using distributions and polynomial functions [13, 21, 37, 38, 56–65].…”