Faculty Bibliography

In order to delineate the specific morphological and immunophenotypic features of B-cell lymphoproliferative disorders associated with trisomy 12, 172 sequential unselected cases of CD19+CD5+ B-cell disorders, primarily affecting the peripheral blood and bone marrow, were studied. Trisomy 12 was found in 24 cases (13.9%), with all cases morphologically classified as either CLL-PL or CLL-mixed by FAB criteria. Trisomy 12 was not found in any cases of typical CLL. Trisomy 12 cases demonstrated a significant higher expression of CD11a (P<0.0001) and CD20 (P<0.0006) when compared to cases with the equivalent morphology and immunophenotype, but without the chromosomal abnormality. Trisomy 12 cases also demonstrated a higher frequency of FMC7, CD38 expression and moderate to strong surface immunoglobulin staining. However, no correlation was detected between the percentages of trisomy 12 cells and cells expressing CD11a, CD38, FMC7 or sIg mean fluorescent intensity. Cells from trisomy 12 positive cases were sorted according to their CD11a expression using fluorescent activated cell sorting. There was a significant increase in the percentage of trisomy 12 cells within the CD11a+ sorted fraction compared to the unsorted population (P < 0.05), implying that trisomy 12 is associated with increased expression of CD11a. With the highly specific morphological and immunophenotypic features demonstrated by trisomy 12 cases in this study, it is highly likely that these cases constitute a specific group of B-cell lymphoproliferative disorders.

Autologous transplantation in CML has been a focus of interest over the last few years. Determining the indications, optimal timing and method for this procedure remains controversial. One approach has been the mobilisation of Philadelphia chromosome negative (Ph-) peripheral blood stem cells following high-dose chemotherapy as a method of purging the graft. We have described a mobilisation regimen of 7 days of hydroxyurea followed by G-CSF and have shown it to be substantially less toxic than other methods. We now report further experience with this technique in a total of 18 patients and the outcome of transplantation in seven patients using cells so-derived. Following mobilisation, approximately a third of patients had 100% Ph-collections and half had less than 50% Ph+ collections. All patients were 100% Ph+ prior to mobilisation. Six out of seven transplanted patients showed sustained engraftment and two of these patients became 18 and 34% Ph+ 3 months post-transplant. Five patients remain alive and well 13 to 25 months post-autograft. In conclusion, we have developed a well-tolerated regimen for Ph- PBSC mobilisation and have demonstrated that such cells are capable of sustained engraftment and of producing significant cytogenetic responses.

The diagnosis of haematological malignancies has undergone major conceptual changes in the past decade. Accurate diagnosis now depends on a combination of morphological examination, cell marker studies, and molecular biological techniques. Specialist haematopathology diagnostic services are best provided in an integrated laboratory, able to carry out the full range of techniques and staffed by pathologists and scientists with expertise in the investigation of leukaemia and lymphoma.

The reduced levels of normal immunoglobulin in patients with myeloma may be due to suppression of normal B-cell differentiation. However, reports on the numbers of B cells vary, with some finding decreases consistent with immunoparesis, and others reporting expansions of phenotypically aberrant cells. We have therefore assessed the phenotype and levels of B lymphocytes in patients at presentation (n = 23), in plateau or complete remission (PB n = 42, BM n = 18), and in relapse (PB n = 17, BM n = 14), in comparison to normal individuals (n = 10). Phenotypic analysis was performed using five-parameter flow cytometry, with CD14 used to exclude monocytes where necessary. We found no evidence of a phenotypically distinctive blood or marrow B-cell population in patients with myeloma, nor of an increase in the levels of any B-cell subset. Numbers of blood CD19+ 38+ normal plasma cell precursors were significantly reduced in presentation/relapse patients, but not in patients in plateau/remission. Total CD19+ cells were significantly reduced only in patients with circulating myeloma cells, detected by IgH-PCR. In the marrow, CD19+ B cells expressing CD5, CD10, CD34, CD38, CD45(low) and Syndecan-1 were significantly decreased at presentation/relapse, but not in patients in plateau/remission. The majority of these antigens are expressed by normal B-cell progenitors, indicating that myeloma also affects the early stages of B-cell development. The suppression of progenitor cells was not restricted to B-lymphoid differentiation, as total CD34+ cells were also significantly reduced in the marrow of myeloma patients at presentation. These results indicate that, if neoplastic B cells are present in myeloma, they are low in number and have a phenotype similar to their normal counterparts. Furthermore, there is a reversible suppression of CD19+ B lymphocytes that correlates inversely with disease stage, and specifically affects the early and late stages of normal B-cell differentiation.

Non-Hodgkin's lymphoma is not commonly a familial condition. This is believed to be the first two cases of primary pancreatic lymphoma within a single family. The two cases, a brother and sister, both presented in their 60s and were diagnosed histologically as having high grade B cell lymphoma affecting the pancreas, an uncommon primary site. Both responded well to treatment with chemotherapy and were in remission at the time of writing. On further investigation it was found that their mother also presented with a malignant lymphoma of cervical nodes 30 years earlier and subsequently died of the disease.

Acute promyelocytic leukaemia (APL) is characterized cytogenetically by t(15;17)(q22:q21) which results in the production of a PML/RAR alpha fusion protein. Detection of the translocation or the fusion gene product is required for objective diagnosis of APL. This can be accomplished by conventional cytogenetic methods, fluorescence in situ hybridization or RT-PCR. Such techniques are time consuming and not universally available. The intracellular distribution of the PML protein in promyelocytes is characteristically altered in APL and this can be detected by immunocytochemistry. We have assessed two immunocytochemical methods, immunofluorescence and alkaline phosphatase-anti-alkaline phosphatase staining (APAAP), with regard to sensitivity, specificity and rapidity of diagnosis. 85 patients with AML including 15 cases of APL were studied. Immunofluorescence PML detection was concordant with RT-PCR for t(15:17) in 14/15 (93.3%) cases with no false positives. The negative APL case in our series was a patient with a 5' PML breakpoint who did not express the reciprocal t(17;15) fusion product. APAAP was concordant in only 6/13 (46%) APL cases with one false positive. In conclusion, immunofluorescent localization of PML using 5E10 monoclonal antibody is a rapid, sensitive and specific diagnostic tool for APL.