During the first two years of this grant we characterize structure, rearrangement and transcription of immunoglobulin heavy chain variable (VH) and constant (CH) segments in B cell chronic lymphocytic leukemia (CLL). Our original hypothesis of biased usage of the VHV family within CLL kindred was found to apply to the population at large. About 30% of all CLLs rearrange and somatically mutate only one (VH251) of the three VHV genes. Germline transcripts of VH251 are noted at high frequency in CLL and ALL cells. No correlation of VHV and VL usage was noted. We have determined the entire sequence across the Cmu-Cdelta locus in CLL DNA and have analyzed transcription at stages of B cell development. We found unexpectedly high levels of the secreted form of delta mRNA (deltas) which may derive from a uniquely efficient recombinational mechanism to delete Cmu. Finally, in analyzing two cases of aggressive CLL in children, we found a common chromosomal translocation [t(2,14)(2p13;14q32)] with breakpoints within 40 bp of one another in an uncharacterized region centromeric to Ckappa. This 2p13 region is hypomethylated and transcribed in the tumors and in CD5-positive B cells. Two other pediatric tumors show a 2p13 intrachromosomal deletion about 25 kbp telomeric to the t(2;14) breaks. We seek to extend these observations in the next funding period through four specific aims: (1) Sequence the rearranged VHV genes of an extended panel of CLLs and of CD5+ and CD5- normal B cells to determine whether selection may be operative in the transformation process. (2) Analyze the structure and stage-specific expression of VHV germline transcripts in normal and malignant cells as correlated with the biased rearrangement of the VHV genes. (3) Determine the mechanism for increased deltas levels in normal and malignant B cells. (4) Study the potential developmental regulation of the 2p13 chromosomal locus that shows deletion and translocation in CLL.
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