The Role Of CD44 Isoform Expression In Niche Resident Chronic Myeloid Leukemia Stem Cell Evolution

Chronic myeloid leukemia (CML) is a myeloproliferative neoplasm characterized by the presence of the BCR-ABL fusion oncogene whose protein product has greatly increased ABL1 kinase activity. Although specific BCR-ABL tyrosine kinase inhibitors (TKIs) and the second generation TKIs like Nilotinib and the dual specific SCR and ABL inhibitor Dasatinib/Sprycel have dramatically improved CML therapy and significantly slowed disease progression by eradicating the bulk of CML cells in the circulation, they frequently fail to eliminate quiescent leukemic stem cells residing in the protective bone marrow niche. Leukemia stem cells (LSC) are able to drive disease relapse and may eventually contribute to the emergence of TKI resistant blast crisis (BC) CML, which is the final phase in the evolution of CML with rapid progression and short survival. CD44 is an adhesion molecule that promotes retention in the niche through adhesion to extracellular matrix components, such as hyaluronic acid and osteopontin. It plays an important role in wound healing and cell migration as well as in tumor invasion and metastasis. Through alternative mRNA splicing several CD44 isoforms exist, some of which are frequently overexpressed by cancer stem cells, including LSCs. The CD44 variant expression pattern on human blast crisis CML LSC, however, had not been elucidated. In this study we aimed to investigate the CD44 transcript variant expression of human blast crisis CML LSC.

We performed whole transcriptome RNA sequencing of FACS sorted CML LSCs (Lin-CD34+CD38+) from chronic phase (CP)(n=8) and blast crisis (BC)(n=8), CML patients as well as the normal counterpart from cord blood (CB) (n=3) and adult peripheral blood (NPB)(n=3). A number of CD44 transcript variants were detected: v3, v4 (CD44s), v5, v6, v7, v8 plus additional variants. Earlier data suggest a total of 16 different isoforms of CD44. We found a higher overall variant gene expression of CD44 in BC compared to CP. A higher expression of CD44 transcript variant 3 and CD44 transcript variant 5 was detected in both CP and BC compared to CB and NPB. Specific CD44 transcript variant expression patterns distinguished BC progenitors from CP samples. Using splice isoform specific PCR, we were able to confirm isoform variants that were upregulated in the BC CML samples. We also compared the expression of the CD44 transcript variants in young versus old bone marrow in order to exclude that LSC-specific isoform variant expression was expressed in aged patients. We could not detect such a correlation.

These observations suggest that unique CD44 isoform expression patterns predict progression from CP to BC as well as the generation of TKI resistant LSCs and may be used as biomarkers of response to LSC targeted therapy.

Jamieson:J&J, Roche: Research Funding; Sanofi: Membership on an entity’s Board of Directors or advisory committees.