Late hematological toxicity after chemotherapy: Exome analysis with long-term follow-up in Mexican population.

Abstract

e24145 Background: Genomic alterations chemotherapy-induced are associated with cytopenias, myelodysplasia, or acute leukemia. Methods: Patients with cancer who achieved a complete response after chemotherapy, had a normal blood cytology and thereafter developed a cytopenia (group 2), + myelodysplasia or acute leukemia (group 3, G-3) were included. Control group (group 1): cases with the same malignancy, age, treatment, and follow-up, except for the development of a late cytopenia. Samples: The genomic DNA (gDNA) was extracted from peripheral blood by standard methods. Thereafter, the Whole Exome Sequencing (WES) libraries were prepared using Illumina DNA Prep with Exome 1.0 Enrichment (Illumina, San Diego CA, United States). All libraries were quantified with the Qubit dsDNA BR Assay Kit (Invitrogen, Carlsbad, CA, USA), libraries size was analysed in S2 Standard DNA Cartridge for Sep 400 (BiOptic, New Taipei City, Taiwan), and sequencing was performed in a NovaSeq 6000 (Illumina, San Diego CA, United States) in a 150 bp pair-end configuration. Computational Analysis: As an initial step, the raw sequence was pre-processed using Trimmomatic 0.40. Pre-processed sequences were aligned to the human reference sequence (hg38) using the Illumina-Dragen Enrichment pipeline (lllumina, San Diego CA, United States). The BAM files resulting from the enrichment were removed from PCR duplicates using Picard Tools (http://broadinstitute.github.io/picard.). Each BAM file was used to obtain the somatic variants using the GATK pipeline, and variants were annotated using ANNOVAR the following databases: Clinvar, gnomAD, refGene, cytoBand, exac03, avsnp147, dbnsfp30a. The somatic variants were then transformed to MAF using Funkotator from the GATK. Additionally, converted annotated variant files were analyzed with the Maftools package from the R programming language to visualize the landscape of critical mutations. Results: After comparing the 3 groups: The 10 top genes with mutations and allelic variants were: MUC12,HLA-DRB1, MUC 16, TTN, HLA-B, HLA-A, HLA-C, FLG, ZNF717, MUC6 & FLG . A greater dispersion in the MUC6, MUC 12, HLA-A, FLG, and TTN genes was found in G-3. The gen MUC6 had the highest number of mutations, including shallow amplification, deep deletion, and amplification. Most of the mutations in G-3 included missense mutations, in frame deletions, and shallow amplification. After analyzing with Oncogenic Signaling pathways in TCGA cohorts, the main pathway involved was RTK-RAS. No statistical difference was found between the level of mutations and the ratio of SNP’s. Conclusions: Mutations and allelic variants vary between patients with cytopenias and secondary myelodysplasia/leukemia. A deeper analysis of these changes is required.