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Wednesday, February 26, 2020

Single Molecule Real Time (SMRT) sequencing sensitively detects polyclonal and compound BCR-ABL in patients who relapse on kinase inhibitor therapy.

Secondary kinase domain (KD) mutations are the most well-recognized mechanism of resistance to tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia (CML) and other cancers. In some cases, multiple drug resistant KD mutations can coexist in an individual patient (“polyclonality”). Alternatively, more than one mutation can occur in tandem on a single allele (“compound mutations”) following response and relapse to sequentially administered TKI therapy. Distinguishing between these two scenarios can inform the clinical choice of subsequent TKI treatment. There is currently no clinically adaptable methodology that offers the ability to distinguish polyclonal from compound mutations. Due to the size of…

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Wednesday, October 23, 2019

Cas9-mediated allelic exchange repairs compound heterozygous recessive mutations in mice.

We report a genome-editing strategy to correct compound heterozygous mutations, a common genotype in patients with recessive genetic disorders. Adeno-associated viral vector delivery of Cas9 and guide RNA induces allelic exchange and rescues the disease phenotype in mouse models of hereditary tyrosinemia type I and mucopolysaccharidosis type I. This approach recombines non-mutated genetic information present in two heterozygous alleles into one functional allele without using donor DNA templates.

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Sunday, September 22, 2019

Clonal distribution of BCR-ABL1 mutations and splice isoforms by single-molecule long-read RNA sequencing.

The evolution of mutations in the BCR-ABL1 fusion gene transcript renders CML patients resistant to tyrosine kinase inhibitor (TKI) based therapy. Thus screening for BCR-ABL1 mutations is recommended particularly in patients experiencing poor response to treatment. Herein we describe a novel approach for the detection and surveillance of BCR-ABL1 mutations in CML patients.To detect mutations in the BCR-ABL1 transcript we developed an assay based on the Pacific Biosciences (PacBio) sequencing technology, which allows for single-molecule long-read sequencing of BCR-ABL1 fusion transcript molecules. Samples from six patients with poor response to therapy were analyzed both at diagnosis and follow-up. cDNA was…

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Sunday, September 22, 2019

Current developments in molecular monitoring in chronic myeloid leukemia.

Molecular monitoring plays an essential role in the clinical management of chronic myeloid leukemia (CML) patients, and now guides clinical decision making. Quantitative reverse-transcriptase-polymerase-chain-reaction (qRT-PCR) assessment of BCR-ABL1 transcript levels has become the standard of care protocol in CML. However, further developments are required to assess leukemic burden more efficiently, monitor minimal residual disease (MRD), detect mutations that drive resistance to tyrosine kinase inhibitor (TKI) therapy and identify predictors of response to TKI therapy. Cartridge-based BCR-ABL1 quantitation, digital PCR and next generation sequencing are examples of technologies which are currently being explored, evaluated and translated into the clinic. Here we…

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Sunday, September 22, 2019

Rare compound heterozygous mutations in gene MSH6 cause constitutive mismatch repair deficiency syndrome.

Few studies reported patients who harbored three kinds of primary tumors simultaneously. Here, we present a 9-year-old boy with colon carcinoma, brain medulloblastoma, and lymphoma. Genetic mutation detection was explored with next-generation sequencing, and compound heterozygous mutations in gene MSH6 c.3103C>T p.Arg1035Ter and c.3261dupC p.Phe1088LeufsTer were discovered.

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