Background: Myeloproliferative Neoplasms (MPNs) are clonal hematopoietic disorders, classified into three different phenotypes Polycythemia Vera (PV), Essential Thrombocytosis (ET) and Primary Myelofibrosis (PMF). This disorder is associated with the presence Jak2V617F mutation in 90% of PV and 50% of ET and PMF patients as well as other mutations such as Jak2 exon 12 and MPL. However, there are still significant numbers of MPNs cases that are negative to most common genetic anomalies and many mutations are still unknown Aim: To identify and elucidate genetic defects causing MPNs that could serve as disease biomarker. Methods: 7 MPN patients and healthy individuals from 3 consanguineous families were consented into the study. Peripheral blood samples were collected from 6 patients and 4 healthy individuals. Further, genomic DNA was extracted and used for multiplex PCR amplification of 190 amplicons targeting 739 cancer associated mutations in 604 loci from 46 key cancer genes, using the Ion AmpliSeq™ Kit. Next Generation Sequencing (NGS) was performed through the Ion Torrent Personal Genome Machine using the 318 chip and was analyzed with the Torrent Suite Software (Full list of mutations targeted can be found at http://www.bcm.edu/geneticlabs/index.cfm?pmid=21681). Mutation details were obtained from the Catalogue Of Somatic Mutations In Cancer (COSMIC) database. A human genome (hg) 19 DNA sequence (http://www.ncbi.nlm.nih.gov/refseq/rsg/) was used as references for this panel of genes. The nomenclature of mutations is based on the convention recommended by the Human Genome Variation Society (http://www.hgvs.org/mutnomen/). The confirmation of NGS data was performed using RQ-PCR or Sanger sequencing and Jak2 exon 12 mutations were studied. Results: Out of four PV patients, three were positive for the JAK2 V617F mutation. One patient had mutations in PDGFRA 838-840delGLA, APC Q1285fs, NPM1 Splice Site Loss and PTEN S10N. The second patient had mutations in KIT M541L, KDR Q472H, TP53 P72R, while the third patient had PI3KCA I391M, KIT M541L, KDR Q472H and P53 P72R mutations. The fourth patient was negative to the JAK2 V617F mutation, but showed PDGFR A838-840delGLA and SMARCB1 T72K mutations. Out of three ET patients, one patient was positive for JAK2 V617F, SMARCB1 T72K, IDH1 K115Q and PDGFRA 838-840delGLA mutations. Two patients were negative for the JAK2 V617F and MPL mutations, one patient had mutations in PDGFRA 838-840delGLA, APC Q1285fs and SMAD4 198-200delPAL, while the other patient had mutations in CDKN2A S73R, APC Q1285fs and PDGFRA 838-840delGLA. Furthermore, the PDGFRA 838-840delGLA and APC Q1285fs mutations were also found in healthy individuals. Conclusion: This study was able to identify a list of deleterious somatic mutations such as missense and frame shift mutations, in-frame deletions, splice loss sites and Single Nucleotide Variation (SNV) in MPNs patients and healthy individuals. Our preliminary results suggest that the MPNs patients in Qatar have complex mutations. Evidences show that there exists a possibility of the disease arising out of the accumulation of genetic alteration and not as the consequence of a single genetic event. This could possibly be due to the high rate of consanguineous marriages in Qatar i.e. the "Founder Effect".


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