Clonal VDJ recombination of the immunoglobulin heavy chain gene by PCR in classical Hodgkin's disease.

Although Hodgkin's disease (HD) has been a subject of much investigation, fundamental questions remain unanswered regarding its lineage and clonality. The authors used a polymerase chain reaction (PCR) technique to investigate whether clonal Variable-Diversity-Joining recombination of the immunoglobulin heavy (IgH) chain gene, a phenomenon that characterizes clonal B-cell proliferations, exists in nodular sclerosing (NSHD) and mixed cellularity (MCHD) Hodgkin's disease (so-called "classical" Hodgkin's disease). The isolation of DNA from paraffin-embedded tissue sections allowed for direct correlation of PCR results with the cell populations that were analyzed. Thirty-two cases were studied. These included 12 cases in which the Reed-Sternberg (RS) cells expressed the B-cell antigen, CD20, and 10 cases that were classified as syncytial variant of NSHD (3 CD20+, 7 B-cell antigen negative). Overall, clonal patterns of VDJ PCR products were found in 14 of 32 (44%) cases. These clonal patterns were identified in 7 of 12 (58%) cases of CD20+ classical HD and in 7 of 20 (35%) cases of B-antigen-negative classical HD. Clonal patterns were found in 3 of 10 cases of syncytial variant of NSHD, including 2 of 3 (67%) CD20+ cases and 1 of 7 (14%) B-cell antigen-negative cases. The results of this study provide support that a subset of HD represents a clonal B-cell neoplasm, and indicate that clonal IgH VDJ sequences are more frequently found in CD20+ HD.

Virus-associated hemophagocytic syndrome characterized by clonal Epstein-Barr virus genome.

Virus-associated hemophagocytic syndromes are a heterogeneous group of disorders in which viral infection is associated with a proliferation of hemophagocytic histiocytes throughout the reticuloendothelial system. The authors report the case of a 24-year-old Vietnamese male who developed a hemophagocytic syndrome associated with Epstein-Barr virus (EBV) and who died following a rapidly progressive course. A proliferation of reactive-appearing lymphoid cells was associated with an extensive proliferation of erythrophagocytic histiocytes. Immunophenotypically, the lymphoid infiltrate consisted of CD56+ natural killer cells, predominantly CD8+ T-cells and rare B-cells (CD20+). Double-label immunohistochemical studies showed CD3+ T-cells and CD56+ natural killer cells to be distinct cell populations. Combined immunohistochemical-in situ hybridization studies localized EBV to CD43+, CD3-, CD68-, lymphoid-appearing cells, indicating the presence of EBV within natural killer cells. Southern hybridization analysis of EBV genomic termini revealed clonal EBV genome. However, there was no detectable immunoglobulin or T-cell receptor gene rearrangements. The findings indicate that this case of hemophagocytic syndrome represents a clonal proliferation of natural killer cells containing EBV and highlights the importance of the analysis of EBV genomic termini for determination of clonality in EBV-associated proliferations. It is possible that other cases of fulminant EBV-associated hemophagocytic syndromes represent clonal natural killer cell proliferations.

Clustered mutations in the second exon of the MYC gene in sporadic Burkitt's lymphoma.

The primary tumors from 15 untreated patients with Burkitt's lymphoma were analysed for abnormalities in the coding region of the MYC gene by single stranded conformational polymorphism (SSCP) analysis followed by DNA sequencing. Fourteen of the 15 tumors had one or more clonal mutations. Forty one mutations were found in the second exon; only one occurred in the third exon. Seven tumors had mutations that clustered in a region spanning amino acids 38-63. Four of these possessed mutations that altered prolines at positions 57 (3), 60 (1), and 63 (1). Seven tumors were mutated in the central portions of the second exon. These occurred at position 95 (2), position 115 (2), position 137 (1), and position 138 (3). Analysis of the published sequences from five lymphoma cell lines and one primary tumor showed a similar clustering of mutations, with all six having mutations in codons between positions 38-63. The regions where mutations occurred have been associated with a variety of properties, including transcriptional activation and cellular transformation. The number and location of mutations showed no correlation with either chromosome 8 or chromosome 14 breakpoints or with the Epstein-Barr virus positivity of the tumors. This unexpected, frequent occurrence of clustered mutations in the second exon of the MYC gene suggests a role for the mutated MYC proteins in the pathogenesis of Burkitt's lymphoma, possibly through altered interactions of this domain with other cellular factors.