W. Martin Howell

Interleukin-10 polymorphisms, cancer susceptibility and prognosis.

Interleukin-10 (IL-10) is a multifunctional cytokine with both immunosuppressive and anti-angiogenic functions and may have both tumour-promoting and -inhibiting properties. A large number of polymorphisms (primarily single nucleotide polymorphisms (SNPs)) have been identified in the IL-10 gene promoter. Convincing evidence that certain of these polymorphisms are associated with differential expression of IL-10 in vitro and in some cases in vivo has been obtained and a number of studies have investigated associations between IL-10 polymorphisms and cancer susceptibility/prognosis. The results from 22 studies in 13 different malignancies are reviewed. In 17 of these studies, positive associations between IL-10 genotype or haplotype and disease susceptibility and/or progression were reported. In some of these cancers genotypes associated with low IL-10 expression were a risk factor for disease or disease progression, while in others genotypes associated with high IL-10 expression were a risk factor. Published findings in breast cancer are as yet conflicting. Most, but not all of the studies reviewed are based on small sample sizes and a limited number of IL-10 polymorphisms. However, the preliminary data obtained thus far indicate that larger studies are required in a number of cancers, in order to confirm initial results, extend studies to include more detailed genotype/haplotype analysis and combine genotype and gene expression studies in the same subjects. Such studies will contribute significantly to our understanding of the biological role of IL-10 in cancer development.

EGF +61 gene polymorphism and susceptibility to and prognostic markers in cutaneous malignant melanoma.

CMM is the most serious cutaneous malignancy and is increasing in frequency among most Caucasian populations, where the most important risk factor is exposure to UV light. Relatively little is known of the genetic factors that mediate susceptibility to and prognosis in sporadic CMM, although a number of genes have been implicated. A striking association between EGF polymorphism and Breslow thickness of invasive CMM has been reported. We have sought confirmation of this finding in an independent study of 159 patients and 310 controls using TaqMan fluorescence‐based genotyping for EGF +61. In our study group, there were no significant differences in EGF genotype frequencies between patients and controls nor was EGF genotype associated with tumour growth phase, stage or mitotic count. However, correlation between EGF genotype and Breslow thickness showed a modestly significant increase in frequency of the EGF (G/G) genotype among tumours >3.5 mm thick (30.0% vs. 9.8%, p = 0.03). In summary, in our group, the EGF +61 polymorphism was not a risk factor for CMM susceptibility, but this polymorphism may play a role in disease progression.

Cancer cachexia is associated with the IL10 −1082 gene promoter polymorphism in patients with gastroesophageal malignancy

BACKGROUNDnThe genetic predisposition of the host to local or systemic inflammation may contribute to the effect of cancer cachexia.nnnOBJECTIVEnWe investigated the relation between cytokine polymorphisms (IL1B -511, IL6 -174, IL10 -1082, TNFA -308, and LTA +252) and markers of nutritional status among patients with gastroesophageal cancer to determine whether any such association was reflected by cytokine concentrations in the tumor or plasma compartments.nnnDESIGNnPatients (n = 203) with a diagnosis of gastroesophageal cancer underwent nutritional assessment (body mass index, anthropometric measures, dysphagia scoring, and estimation of dietary intake). Single nucleotide polymorphism genotyping was performed by TaqMan allelic discrimination genotyping. Serum cytokine and C-reactive protein concentrations were determined by enzyme-linked immunosorbent assay. Tumor tissue cytokine protein concentrations (n = 56) were determined by using the Cytometric Bead Array System.nnnRESULTSnIL10 GG and IL6 CC polymorphisms were associated with elevated serum C-reactive protein concentrations, and the IL6 CC genotype was also associated with elevated tumor tissue cytokine concentrations. At diagnosis, the IL10 GG, but not the IL6, genotype was linked with increased total weight loss: 4.9% for AA, 7.1% for AG, and 12.0% for GG (P = 0.007). Serum C-reactive protein concentrations correlated with increased weight loss (r = 0.24, P < 0.001). Compared with other genotypes, the IL10 GG genotype retained an independent association in determining the extent of weight loss on multivariate analysis (95% CI: 0.52, 3.43; P = 0.008). Possession of the GG allele was associated with a 2.3 times increased risk of developing cachexia (95% CI: 1.2, 4.3; P = 0.014).nnnCONCLUSIONnThese data suggest that the IL10 genotype of the host can influence the development of cachexia among patients with gastroesophageal malignancy.

Symposium on ’Nutrition in the post-genomic era’ Plenary session 4: Genetic variation and diet-related disease

Genes whose products play a critical role in regulation of the immune response include the human leucocyte antigen (HLA) and cytokine families of genes. The HLA genes are the most polymorphic found in the human genome, and the bulk of this polymorphism results in functional differences in expressed HLA molecules, resulting in inter-individual differences in presentation of peptide antigens to T-cells. In addition, a considerable number of cytokine-associated gene polymorphisms have been identified, the bulk of which occur in the upstream promoter sequences of these genes, which in many cases results in differential in vitro expression of the respective pro- or anti-inflammatory gene product. Particular HLA polymorphisms result in well-defined associations with a large number of immunologically-mediated diseases, including some diseases with known dietary risk factors. For example, individuals of HLA-DQA1*0501, DQB1*0201 genotype have a greater than 200-fold increased risk of developing intolerance to dietary wheat gluten (coeliac disease), and additional HLA-related factors may influence the development of malignant lymphoma within pre-existing coeliac disease. Similarly, HLA-DRB1 alleles sharing a common sequence motif constitute the primary known genetic risk factor for rheumatoid arthritis. The influence of polymorphisms associated with differential cytokine expression on disease susceptibility is currently of much interest. Most attention has been focused on associations with susceptibility to benign immunologically-mediated diseases, including a number of gut diseases. However, recent work from our laboratory indicates that cytokine polymorphisms may influence susceptibility to and prognosis in a number of different cancers, including malignant melanoma skin cancer and solid tumours which may be influenced by diet, such as prostate cancer (collaboration with the CRC/BPG UK Familial Prostate Cancer study). In addition, preliminary work suggests that dietary modulation of expression levels of certain cytokines in healthy human subjects may be genotype dependent.

Rheumatoid arthritis T cells produce Th1 cytokines in response to stimulation with a novel trispecific antibody directed against CD2, CD3, and CD28

Rheumatoid arthritis (RA) T cells respond poorly to conventional mitogens. We have examined the proliferative and cytokine responses of T cells to a synthetic trispeci®c antibody (Tsab) directed against CD2, CD3, and CD28. In 11 subjects RA T cells proliferated more, and secreted signi®cantly more IL-2, in response to Tsab than did control peripheral blood (PB) cells. Very high levels of IL-2 were produced by 2 patients with aggressive disease. Measurement of intracellular IL-2, IFN-c, IL-4, and IL-5 by ̄ow cytometry showed a Th1 pattern of cytokine production in 13 RA and 9 control subjects. We conclude that RA T cells are not irreversibly inactivated, and that spatial arrangement of stimulating molecules may be important in eliciting maximal responses.Rheumatoid arthritis (RA) T cells respond poorly to conventional mitogens. We have examined the proliferative and cytokine responses of T cells to a synthetic trispecific antibody (Tsab) directed against CD2, CD3, and CD28. In 11 subjects RA T cells proliferated more, and secreted significantly more IL-2, in response to Tsab than did control peripheral blood (PB) cells. Very high levels of IL-2 were produced by 2 patients with aggressive disease. Measurement of intracellular IL-2, IFN-gamma, IL-4, and IL-5 by flow cytometry showed a Th1 pattern of cytokine production in 13 RA and 9 control subjects. We conclude that RA T cells are not irreversibly inactivated, and that spatial arrangement of stimulating molecules may be important in eliciting maximal responses.

The HLA System: An Update and Relevance to Patient‐Donor Matching Strategies in Clinical Transplantation

In recent years the development of recombinant DNA and sequencing techniques has led to a greatly increased understanding of the genetic complexity, structure and function of the human major histocompatibility complex. This system may be subdivided into the ‘classical HLA (human leukocyte antigen) class I and II transplantation antigens and novel HLA and non‐HLA genes, involved in antigen processing and presentation to T cells. Parallel technological developments in HLA DNA typing in the clinical laboratory have resulted in a more precise awareness of the role of HLA matching for the classical HLA antigens in bone marrow and solid organ transplantation, while alternative strategies and techniques for donor selection are currently under evaluation. This review offers a current perspective on the genetics, structure and function of the HLA system, its relevance to clinical transplantation and future prospects for improvements in donor selection.

Variants in the genes encoding TNF-α, IL-10, and GSTP1 influence the effect of α-tocopherol on inflammatory cell responses in healthy men

BACKGROUNDnDespite evidence of antioxidant effects of vitamin E in vitro and in animal studies, large, randomized clinical trials have not substantiated a benefit of vitamin E in reducing inflammation in humans. An individuals genetic background may affect the response to α-tocopherol supplementation, but this has rarely been investigated.nnnOBJECTIVEnThe aim of this study was to explore the role of genetic polymorphisms on changes in LPS-stimulated inflammatory cytokine production from peripheral blood mononuclear cells (PBMCs) after α-tocopherol supplementation.nnnDESIGNnA total of 160 healthy, middle-aged male volunteers (mean age: 52.7 y) were given dietary supplements of either 75 IU (low dose; n = 57) or 600 IU (high dose; n = 103) α-tocopherol/d for 6 wk. The production of TNF-α and IL-1β, -6, and -10 by PBMCs after LPS stimulation was measured at baseline and after 6 wk. Polymorphisms in 15 genes involved in inflammation or responses to oxidative stress were characterized in the subjects.nnnRESULTSnThe ability of α-tocopherol to affect TNF-α production by LPS-stimulated PBMCs was influenced by the TNFA -238 polymorphism (P = 0.016). The ability of α-tocopherol to affect IL-6 production was influenced by the GSTP1 313 polymorphism (P = 0.019). The ability of α-tocopherol to affect IL-1β production was influenced by the IL10 -592 and -1082 polymorphisms (P = 0.025 and P = 0.016, respectively).nnnCONCLUSIONSnIn healthy control subjects, the effect of α-tocopherol supplementation on the production of inflammatory cytokines appears to be dependent on an individuals genotype. These genotype-specific differences may help explain some of the discordant results in studies that used vitamin E.

Symposium on 'Nutrition in the post-genomic era' Plenary session 4: Genetic variation and diet-related disease Gene polymorphisms, inflammatory diseases and cancer

Genes whose products play a critical role in regulation of the immune response include the human leucocyte antigen (HLA) and cytokine families of genes. The HLA genes are the most polymorphic found in the human genome, and the bulk of this polymorphism results in functional differences in expressed HLA molecules, resulting in inter-individual differences in presentation of peptide antigens to T-cells. In addition, a considerable number of cytokine-associated gene polymorphisms have been identified, the bulk of which occur in the upstream promoter sequences of these genes, which in many cases results in differential in vitro expression of the respective pro- or anti-inflammatory gene product. Particular HLA polymorphisms result in well-defined associations with a large number of immunologically-mediated diseases, including some diseases with known dietary risk factors. For example, individuals of HLA-DQA1*0501, DQB1*0201 genotype have a greater than 200-fold increased risk of developing intolerance to dietary wheat gluten (coeliac disease), and additional HLA-related factors may influence the development of malignant lymphoma within pre-existing coeliac disease. Similarly, HLA-DRB1 alleles sharing a common sequence motif constitute the primary known genetic risk factor for rheumatoid arthritis. The influence of polymorphisms associated with differential cytokine expression on disease susceptibility is currently of much interest. Most attention has been focused on associations with susceptibility to benign immunologically-mediated diseases, including a number of gut diseases. However, recent work from our laboratory indicates that cytokine polymorphisms may influence susceptibility to and prognosis in a number of different cancers, including malignant melanoma skin cancer and solid tumours which may be influenced by diet, such as prostate cancer (collaboration with the CRC/BPG UK Familial Prostate Cancer study). In addition, preliminary work suggests that dietary modulation of expression levels of certain cytokines in healthy human subjects may be genotype dependent.

The Special Case of HLA Genes: Detection and Resolution of Multiple Polymorphic Sites in a Single Gene

The major histocompatibility complex (MHC) has been the most intensively studied region of the human genome during the past three decades (Bodmer 1995) and it has also been widely studied in other mammalian species, including primates (Trowsdale 1995). These studies suggest that the MHC is probably the area of the mammalian genome that is most densely populated with functional genes. In humans, the MHCis principally known as the region that includes genes encoding the human leukocyte antigens (HLA), which playa crucial role in regulating the immune response in health and disease. These HLA genes are the most polymorphic found in humans and, since most polymorphisms encode functional variants, this polymorphism results in inter-individual variation in the immune response. As a result of this, HLA polymorphism has long been recognised as a critical factor in determining allograft acceptance and rejection mechanisms in both renal and bone marrow transplantation (Opelz et al. 1993; Madrigal et al. 1997b), while particular HLA genotypes are associated with a large number of benign and malignant immunologically mediated diseases (Thorsby 1997; Bateman and Howell 1999). Although the MHC also encodes a number of other genes of immunological significance (e. g. the tumour necrosis factor genes), this chapter will focus solely upon the so-called “classical” HLA class I and class II antigens.