O-013 Mesenchymal stromal cells support significant engraftment of low-risk myelodysplastic syndromes (MDS) in a murine xenograft model

chondrial DNA (mtDNA) mutations. These mitochondrial-associated phenotypes suggest that mitochondrial dysfunction plays a possible role in the pathophysiology of MDS. Introduction: Next generation sequencing (NGS) is ideal to study mtDNA mutations because it allows for the detection of low-level heteroplasmy between flow-sorted samples. These changes would be challenging to detect by other methods. We hypothesized that in MDS, mtDNA changes would be found in genes or tRNAs involved in oxidative phosphorylation (OXPHOS) as had previously been shown in a wide range of cancers, including leukemia. Purpose: Using NGS, we sequenced mtDNA from hematopoietic progenitor cells of six MDS patients, flow-sorted by cell surface markers CD3+ (T-cell), CD33+ (myeloid) and CD34+ (hematopoietic progenitor). We analyzed the changes in mtDNA observed in the sorted hematopoietic progenitor and myeloid cells and compared them to T-cells. Materials and Methods: Bone marrow cells were flow-sorted prior to DNA extraction. mtDNA was enriched by long-range PCR (LRPCR). The LRPCR amplicons were equimolarly pooled and libraries constructed prior to Illumina NGS sequencing. Paired-end sequences were aligned to NC_012920 and PCR duplicates removed using BWA/Samtools and CLCBio. Variants were called using quality based metrics. Heteroplasmy levels were calculated using allele counts at a given m. position. Results: Three of the 6 samples had no sequence discordants between cell-sorted populations. One sample acquired two variants in CD34+ cells, a novel MT-TY m.5850T>C, and MT-ATP8 m.8412T>C p.Met16Thr previously reported as a polymorphism. Another sample had a novel variant with increasing heteroplasmy between CD33+ and CD34+ cells (23% to 92.8% alternate allele, respectfully) in MTCO2, m.7605T>C p.Val7Ala. The last sample had three novel variants in CD34+ cells, MT-ND1 m.3810A>G p.Thr168Thr; MT-CO2 m.7983T>A p.Leu133Gln and MT-CO3 m.9817A>G p.His204Arg. This patient also had a homoplasmic reversion back towild type compared with CD33+ cells in MT-C03 m.9477G p.Val61. Of the seven variants detected in 3/6 patients, five were novel. These novel variants are associated with OXPHOS complexes I, IV, and V. Conclusions: As most information regarding mtDNA variants is from mitochondrial disorder research, the identification of multiple novel variants from these MDS samples may be significant. Further NGS analysis and functional studies of novel mtDNA variants associated with MDSmay improve our understanding of the disease biology and possibly bear relevance to prognosis and treatment.