W. M. Howell

Influence of vascular endothelial growth factor single nucleotide polymorphisms on tumour development in cutaneous malignant melanoma.

Vascular endothelial growth factor (VEGF) is a potent regulator of vasculogenesis and tumour angiogenesis. We have investigated whether the VEGF −2578, −1154, +405 and +936 SNPs and associated haplotypes confer susceptibility to and/or influence prognosis in cutaneous malignant melanoma (CMM) skin cancer. A total of 152 CMM patients and 266 controls were genotyped for VEGF promoter SNPs by ARMS-PCR. Strong linkage disequilibrium between the −2578, −1154 and +405 SNPs was detected (association, ρ = 0.488–0.965), but not between these SNPs and SNP +936 (association, ρ = 0.004–0.130). No SNPs or three SNP haplotypes (−2578, −1154, +405) were significantly associated with CMM, although a number of non-significant trends were observed. However, the VEGF −1154 AA genotype and −2578, −1154, +405 CAC haplotype were both significantly associated with less advanced (Stage 1) disease (P = 0.03). In addition, the VEGF −1154 AA genotype was associated with thinner primary vertical growth phase tumours (P = 0.002), while VEGF −1154 GG was associated with thicker primary tumours (P = 0.02). These preliminary results indicate that VEGF genotype may influence tumour growth in CMM, possibly via the effects of differential VEGF expression on tumour angiogenesis.

IL-10 promoter polymorphisms influence tumour development in cutaneous malignant melanoma

Cutaneous malignant melanoma (CMM) is the most serious cutaneous malignancy. CMM patients often develop an immune response to their tumours. Conflicting evidence suggests that IL-10 may contribute to tumour escape from the immune response, but may also have an anti-tumour effect. To distinguish between these models and to determine whether genotypes associated with differential IL-10 expression confer susceptibility to and/or influence prognosis in CMM, 165 CMM patients and 158 controls were genotyped for IL-10 promoter SNPs by ARMS-PCR. The IL-10 −1082 AA low expression genotype was increased in incidence among CMM patients (P = 0.04). In addition, IL-10 genotypes showed significant associations with three of four prognostic indicators examined; IL-10 −1082 GG (P = 0.02) and −1082, −819 and −592 compound high expression (P = 0.03) genotypes were associated with horizontal (non-invasive) tumour growth; IL-10 −1082 AA low expression genotype was associated with more advanced (Stage II–IV) disease (P = 0.04); finally, the IL-10 −1082 AA (P = 0.005) and compound low expression (P = 0.009) genotypes were significantly increased in frequency among patients with thicker primary Vertical growth phase tumours. These results indicate that genotypes associated with high levels of IL-10 expression in vitro are protective in CMM, while low expression genotypes are a risk factor for more advanced/poorer prognosis disease and may confer susceptibility to CMM. Although the influence of IL-10 on melanoma development is likely to be complex, these results support recent findings that IL-10 has an anti-tumour effect in CMM, possibly via inhibition of angiogenesis.

Human leukocyte antigens and cancer: is it in our genes?

Human leukocyte antigens (HLAs) are widely expressed cell surface molecules which present antigenic peptides to T‐lymphocytes, thus modulating the immune response. The efficiency of peptide presentation by HLAs is dependent on the extreme polymorphism in the HLA‐encoding loci within the major histocompatibility complex (MHC). HLA polymorphism is known to alter disease susceptibility and progression in a range of predominantly inflammatory conditions, many of which are T‐lymphocyte‐mediated. More recently, the importance of alterations in HLA expression and polymorphisms within HLA‐encoding loci has emerged in the development of malignancy. This review concentrates on the role of HLA polymorphism in malignant disease, with discussion of the major cancers in which HLA associations have become clear, as well as the potential mechanisms by which HLA polymorphisms may act as important factors, or cofactors, in the pathogenesis of malignant disease. In addition, the role of certain non‐HLA genes within the MHC in the pathogenesis of malignancy is also considered briefly. Copyright

Variable X‐chromosome DNA methylation patterns detected with probe M27β in a series of lymphoid and myeloid malignancies

In this study the X chromosome probe M27β was used to investigate DNA methylation at the DXS255 locus and hence X inactivation status and determination of tumour clonality in blood, bone marrow and biopsy tissue involved with morphologically and phenotypically defined lymphoid and myeloid disease from 14 female patients along with uninvolved bone marrow from two control individuals. Thirteen out of 16 individuals (81%) were restriction fragment length polymorphism (RFLP) heterozygous for DXSZ 55. DNA methylation status could not be assessed in the three DXS255 homozygous individuals. In eight DXS255 heterozygous individuals clonality was clearly demonstrated using M27β and in six of these cases independent analysis using T cell receptor (TcR) and immunoglobulin (Ig) gene probes confirmed the presence of clonal tumour cell populations. In the two controls, polyclonality was inferred from M27β probe analysis. In the remaining three cases (all acute lymphoblastic leukaemia (ALL)) both DXSZ55 X chromosome sequences appeared to be methylated. Clonality in these cases was demonstrated by TcR or Ig monoclonal gene rearrangements. These data demonstrate the value of the M27β probe for determining tumour clonality in a number of cases with lymphoid and myeloid disease but indicate that there may not always be a complete correlation between DNA methylation, X inactivation status and tumour clonality in certain lymphoid neoplasms, restricting the use of this probe in clonality studies. Correlations between DNA methylation, X inactivation status and stage of normal and neoplastic T and B cell development require further investigation.

Investigation of specimen mislabelling in paraffin-embedded tissue using a rapid, allele-specific, PCR-based HLA class II typing method.

Mislabelling of surgical specimens can seriously affect the accuracy of histopathology reports. We describe two cases in which clinically suspected mislabelling was investigated by polymerase chain reaction (PCR)‐based HLA DRB and DQB tissue typing of paraffin biopsy‐derived DNA, using sequence specific primers (PCR‐SSP HLA typing). In the first case, two patients underwent surgery for breast carcinoma. A subcutaneous lymph node containing metastatic carcinoma was received with the breast specimen from one patient, but was clinically considered more likely to originate from the other patient who underwent breast surgery on the same day. In the second case, histological examination of retained products of conception from a young woman revealed adenocarcinoma, but a repeat curettage specimen consisted of secretory phase endometrium. In case 1, PCR‐SSP HLA typing confirmed the clinical suspicion that the subcutaneous lymph node received with tissue from one patient originated from the other patient. This result converted the first patient from lymph node positive breast carcinoma to lymph node negative disease. In case 2, there was no evidence from PCR‐SSP HLA typing that the endometrial samples had originated from different patients. PCR‐SSP HLA typing requires fewer steps than methods based on PCR amplification followed by oligonucleotide probing (PCR‐SSOP HLA typing), and relies on the amplification of shorter sequences of DNA. Therefore, this technique can produce more rapid results than PCR‐SSOP HLA typing, and is ideally suited to typing partially degraded DNA derived from formalin‐fixed and paraffin‐embedded tissue.

HLA DPB1 ALLELES AND SUSCEPTIBILITY TO RHEUMATOID ARTHRITIS

HLA‐DPB1 genotypic and phenotypic frequencies were investigated in a series of 35 adult rheumatoid arthritis (RA) patients and 42 controls. No significant associations between DPB1 alleles and susceptibility to RA were demonstrated, although some non‐significant differences in DPB1*0301 and 0401 allele frequencies between patients and controls were observed.

Histologic, immunophenotypic and genotypic analyses of bone marrow trephines from patients with non-Hodgkin's lymphoma

Marrow involvement in 20 patients with non-Hodgkins lymphoma (NHL) were studied by histology, immunophenotypic and genotypic methods. Eighteen of these trephines were histologically involved with recognizable lymphomatous infiltrates and five of these were the primary disease site. In the remaining two cases (with histologically involved lymph nodes) the trephines were uninvolved with tumour. Three B-cell cases expressing surface immunoglobulin (sIg) and/or CD37 and one case not analysed phenotypically showed Ig gene rearrangements. The two remaining cases with B NHL showed no gene rearrangements, however, in one of these the trephine was histologically uninvolved with tumour. Twelve out of 14 T-cell cases were characterized by variable or absent expression of one or more T-cell antigens from the tumour population, one case was negative for all T-cell antigens and the remaining case was not histologically involved with tumour. All three lymphoblastic lymphomas and only 4/11 peripheral T-cell lymphomas (PTCL) cases revealed T-cell receptor (TcR) gene rearrangements. One of the latter cases also exhibited Ig JH gene rearrangements. This study demonstrates the usefulness of bone marrow trephines (BMT) in histologic, phenotypic and genotypic analyses. However, although genotypic data confirm clonality in B NHL and the lymphoblastic lymphomas there was genotypic heterogeneity within the PTCL group.