Although both Bcl-2/Bax ratio and Mcl-1 have been identified to be of clinical relevance in B-cell chronic lymphocytic leukemia (CLL), there is controversy regarding their role; further, their relative importance is not well delineated. Expression of Bcl-2, Bax, the Bcl-2/Bax ratio, and Mcl-1 in 51 consecutive previously untreated CLL patients and 16 controls was determined by Western blotting. Only 37 patients were treated, all with chlorambucil and prednisone initially. Six patients achieved complete response (CR), 14 were non-responders (NR), and 17 had a partial response (PR), as defined by NCI criteria. There was considerable inter-patient variability in protein expression and overlap with healthy volunteers (P > 0.05). All patients with CR had low Mcl-1 levels compared to the PR + NR group (0.07 ± 0.02 vs. 0.14 ± 0.07, P = 0.043). Higher Mcl-1 expression as determined by dichotomizing the data was associated with a failure to achieve CR (P = 0.021). The Bcl-2/Bax ratio was significantly associated with treatment response only when CR and PR were considered together (0.89 ± 0.53 [CR + PR] vs. 3.38 ± 4.47 [NR], P = 0.0118). There was no association with Rai stage. Low Mcl-1 appears to be a requirement for CR, while low Bcl-2/Bax ratio is indicative of some response to conventional treatment. Am.

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Sequence similarity between the clonal bands in HT and subsequently developed thyroid lymphoma is supportive of the argument that primary thyroid lymphoma may evolve from HT.

Helicobacter pylori (Hp)-associated gastritis is a risk factor for gastric mucosa-associated lymphoid tissue (MALT) lymphoma. Clonal B-cell populations are present in both reactive and neoplastic MALT tissue, thus limiting their usefulness in the evaluation of gastric lymphoid in®ltrates in endoscopic biopsy specimens. The aim of this study was to identify the presence of clonal B-cell populations in Hp-gastritis with MALT and to assess their usefulness in distinguishing reactive from malignant in®ltrates. Routinely ®xed paraf®n-embedded blocks from 20 patients with Hp-gastritis with lymphoid hyperplasia were analysed for B-cell clonality by a semi-nested polymerase chain reaction (PCR) using FRIII/LJH and FRIII/VLJH primers for ampli®cation of the VDJ region of the immunoglobulin heavy chain gene. The histopathological ®ndings were evaluated according to a previously published scoring system. Immunohistochemistry was performed by the labelled streptavidin±biotin technique using the following primary antibodies: CD45, CD45RO, CD3, CD20, and cytokeratin. The histopathological ®ndings were diagnostic of Hp-chronic active gastritis (grade 2, n=17; grade 3, n=3). Scattered intraepithelial B-cells were present in all cases and non-destructive lymphoepithelial lesions in one grade 3 case. Ampli®able DNA was obtained from all samples. Clonal bands were observed in ten (7/17 grade 2 and 3/3 grade 3 lesions) and polyclonal smears in ten cases (all grade 2). The clonal bands were often (n=6) associated with a background polyclonal smear and were not reproducible from deeper sections (n=10) or another paraf®n block (n=1), while the clonal bands in control lowgrade MALT lymphomas were not associated with a background smear and were reproducible from deeper sections. None of the patients has developed lymphoma to date (follow-up 21±44 months). In conclusion, B-cell clonal bands are common in H. pylori-gastritis with lymphoid hyperplasia. The irreproducibility of these bands is a useful feature in favouring a reactive process.

The MCL-1 promoter insertion may identify a high-risk group of CD38-negative CLL patients.

Earlier we had reported a guanine to adenosine substitution at position 125 (G125A) in the BAX promoter, and its association with higher stage of the disease and failure to achieve complete response to treatment in chronic lymphocytic leukemia (CLL) patients. The aim of this study was to test the hypothesis that G125A leads to a reduction in the transcription of the BAX gene and that this is a direct cause of altered BAX mRNA and protein expression. In lymphocytes of CLL patients, BAX mRNA expression was determined by RNase protection assay and Bax protein was detected by immunoblotting. The presence of G125A in the BAX promoter was associated with lower BAX mRNA (P ¼ 0.004) and protein (P ¼ 0.024) levels. In transient expression assays using wild-type and mutant BAX promoter sequences linked to Luciferase as a reporter, the G125A polymorphism reduced expression of the BAX promoter by 2.6-fold. These studies suggest a mechanism for the biological effect of the G125A.

Background: A novel single nucleotide polymorphism (SNP), G(−248)A, in the 5′ untranslated region of the BAX promoter and its association with reduced protein expression, progression beyond Rai stage 0, and treatment resistance in chronic lymphocytic leukaemia (CLL) has been reported previously. Aim: To develop a restriction enzyme analysis (REA) based method for routine detection of BAX promoter SNP in a clinical laboratory. Methods: The BAX promoter was analysed in duplicate by REA and sequencing in 90 samples (from 45 patients with CLL, 43 controls, and two cell lines). The promoter region was amplified, digested with restriction endonucleases (Aci I and Tau I), and separated by gel electrophoresis. Results: After digestion, the normal GG genotype samples produced three distinct bands. The homozygous AA replacement abolished the cleavage site, resulting in a single band. Although the heterozygous samples produced three bands, the two smaller visible bands were reduced in intensity (> 50%). The test characteristics of Aci I REA were better than those of Tau I REA, in terms of sensitivity (100% v 77.8%), specificity (98.6% v 92.3%), positive predictive value (95.03% v 87.4%), and negative predictive value (100% v 85.83%). Conclusions: REA using Aci I is a highly sensitive and specific method for detecting the BAX G(-248)A SNP in CLL.

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