Abstract
Genetic heterogeneity though common in tumors has been rarely documented in cell lines. To examine how often B-lymphoma cell lines are comprised of subclones, we performed immunoglobulin (IG) heavy chain hypermutation analysis. Revealing that subclones are not rare in B-cell lymphoma cell lines, 6/49 IG hypermutated cell lines (12%) consisted of subclones with individual IG mutations. Subclones were also identified in 2/284 leukemia/lymphoma cell lines exhibiting bimodal CD marker expression. We successfully isolated 10 subclones from four cell lines (HG3, SU-DHL-5, TMD-8, U-2932). Whole exome sequencing was performed to molecularly characterize these subclones. We describe in detail the clonal structure of cell line HG3, derived from chronic lymphocytic leukemia. HG3 consists of three subclones each bearing clone-specific aberrations, gene expression and DNA methylation patterns. While donor patient leukemic cells were CD5+, two of three HG3 subclones had independently lost this marker. CD5 on HG3 cells was regulated by epigenetic/transcriptional mechanisms rather than by alternative splicing as reported hitherto. In conclusion, we show that the presence of subclones in cell lines carrying individual mutations and characterized by sets of differentially expressed genes is not uncommon. We show also that these subclones can be useful isogenic models for regulatory and functional studies.
Highlights
Tumors evolve under selective pressures, including the inhibitory influences of anti-cancer drugs
The bimodal expression of various B-cell markers on U-2932 allowed flow-sorting of the subclones which – underlining their usefulness - let to the discovery that BCL6 can drive expression of germinal center markers in diffuse large B cell lymphoma (DLBCL) [5]
We set out to examine how often cell lines consist of subclones
Summary
Tumors evolve under selective pressures, including the inhibitory influences of anti-cancer drugs. Clonal evolution is an important topic in cancer research because it underlies development of heterogeneous, molecularly related tumors in one patient and may explain why relapsed samples are often genetically distinct from primary diagnostic clones [1,2,3]. The diffuse large B cell lymphoma (DLBCL) cell line U-2932 is an example of a cell line comprising subclones, with differential expression of > 100 genes, including the germinal center oncogenes BCL6 and MYC [4]. The bimodal expression of various B-cell markers on U-2932 allowed flow-sorting of the subclones which – underlining their usefulness - let to the discovery that BCL6 can drive expression of germinal center markers in DLBCL [5]
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