Clonal evolution in a primary cutaneous follicle center B cell lymphoma revealed by single cell analysis in sequential biopsies

B cell neoplasias descending from germinal center cells harbor the hallmark of intraclonal diversity resulting from ongoing mutation in the variable p arts of their immunoglobulin-encoding genes. To characterize a primary cuta neous follicle center B cell lymphoma in more detail, we analyzed the respe ctive V-H and V-L genes in single cells mobilized from four sequential biop sies, three taken from the skin and one obtained after internal disseminati on from a retrobulbar infiltrate. The lymphoma cells were found to contain V5-51/D6-12/JH5b (heavy chain) and A27/Jkappa2 (light chain) gene rearrange ments detected on both the genomic and the transcriptional level. To provid e an accurate mutation analysis, the specific V-H gene counterpart (V5-51UK ) was cloned from the patient's germline. Analyzing 226 single cells, we fo und: (i) complete nucleotide identity when V-H and V-L genes of lymphoma ce lls from one particular biopsy were compared among each other; (ii) intracl onal diversity due to ongoing mutation comparing the sequences obtained fro m sequential biopsies; (iii) both V-H and V-L genes to be highly mutated. D educing from the sequence data, we propose a scenario of the clonal evoluti on of the B cell tumor in this patient. From the molecular-biological point of view, this primary cutaneous follicle center B cell lymphoma shows the features of a germinal center cell lymphoma. To draw this conclusion from s ingle cell PCR data, however, a sample of sequential biopsies had to be ana lyzed.