Targeting cancer with a lupus autoantibody

Systemic lupus erythematosus (SLE) is distinct among autoimmune diseases because of its association with circulating autoantibodies reactive against host DNA. The precise role that anti-DNA antibodies play in SLE pathophysiology remains to be elucidated, and potential applications of lupus autoantibodies in cancer therapy have not previously been explored. We report the unexpected finding that a cell-penetrating lupus autoantibody, 3E10, has potential as a targeted therapy for DNA repair–deficient malignancies. We find that 3E10 preferentially binds DNA single-strand tails, inhibits key steps in DNA single-strand and double-strand break repair, and sensitizes cultured tumor cells and human tumor xenografts to DNA-damaging therapy, including doxorubicin and radiation. Moreover, we demonstrate that 3E10 alone is synthetically lethal to BRCA2-deficient human cancer cells and selectively sensitizes such cells to low-dose doxorubicin. Our results establish an approach to cancer therapy that we expect will be particularly applicable to BRCA2-related malignancies such as breast, ovarian, and prostate cancers. In addition, our findings raise the possibility that lupus autoantibodies may be partly responsible for the intrinsic deficiencies in DNA repair and the unexpectedly low rates of breast, ovarian, and prostate cancers observed in SLE patients. In summary, this study provides the basis for the potential use of a lupus anti-DNA antibody in cancer therapy and identifies lupus autoantibodies as a potentially rich source of therapeutic agents.

The diet of the dingo (Canis lupus dingo and hybrids) in north-eastern Australia: a supplement to the paper of Brook and Kutt (2011)

Dingoes and other wild dogs (Canis lupus dingo and hybrids) are generalist predators that consume a wide variety of different prey species within their range. Little is known, however, of the diets of dingoes in north-eastern Australia where the potential for impacts by dingoes exists. Recently new information has been provided on the diets of dingoes from several sites in Queensland, Australia, significantly adding to the body of published knowledge on ecosystems within this region. Further information on the diet of dingoes in north-eastern Australia is added from 1460 scats collected from five sites, representing tropical savannahs, tropical offshore islands (and a matched mainland area), dry sclerophyll forests and peri-urban areas on the fringe of Townsville. Macropods, possums and bandicoots were found to be common prey for dingoes in these areas. Evidence suggested that the frequency of prey remains in scats can be an unreliable indicator of predation risk to potential prey and it was found that novel and unexpected prey species appear in dingo diets as preferred prey become unavailable. The results support the generalisation that dingoes prefer medium- to large-sized native prey species when available but also highlight the capacity for dingoes to exploit populations of both large and small prey species that might not initially be considered at risk from predation based solely on data on scats.

Genetic studies on Finnish lupus erythematosus patients with cutaneous manifestations

Lupus erythematosus (LE) is a chronic, heterogeneous autoimmune disorder with abnormal immune responses, including production of autoantibodies and immune complexes. Clinical presentations of the disease range from mild cutaneous manifestations to a more generalised systemic involvement of internal organs. Cutaneous (CLE) forms are further subclassified into discoid LE (DLE), subacute cutaneous LE (SCLE) and acute cutaneous lupus erythematosus (ACLE), and may later progress to systemic disease (SLE). Both genetic and environmental factors contribute to the disease, although the precise aetiology is still elusive. Furthermore, complex gene-gene or gene-environment interactions may result in different subphenotypes of lupus. The genetic background of CLE is poorly known and only a few genes are confirmed, while the number of robust genetic associations in SLE exceeds 30. The aim of this thesis was to characterise the recruited patients clinically, and identify genetic variants conferring susceptibility to cutaneous variants of LE. Given that cutaneous and systemic disease may share underlying genetic factors, putative CLE candidate genes for genotyping were selected among those showing strong evidence of association in SLE. The correlation between relevant clinical manifestations and risk genotypes was investigated in order to find specific subphenotype associations. In addition, epistatic interactions in SLE were studied. Finally, the role of tissue degrading matrix metalloproteinases (MMP) in LE tissue injury was explored. These studies were conducted in Finnish case-control and family cohort, and Swedish case-control cohort. The clinical picture of the patients in terms of cutaneous, haematological and immunological findings resembled that described in the contemporary literature. However, the proportion of daily smokers was very high supporting the role of smoking in disease aetiology. The results confirmed that, even though clinically distinct entities, CLE and SLE share predisposing genetic factors. For the first time it was shown that known SLE susceptibility genes IRF5 and TYK2 also increase the risk of CLE. A tendency toward gene-gene interaction between these genes was found in SLE. As a remarkable novel finding, it was observed that ITGAM polymorphisms associated even more strongly to DLE than SLE, and the risk estimates were substantially higher than those reported for SLE. Several other recently identified SLE susceptibility genes showed signs of good or modest association especially in DLE. Subphenotype analyses indicated possible associations to clinical features, but marginally significant results reflected lack of sufficient power for these studies. Thorough immunohistochemical analyses of several MMPs demonstrated a role in epidermal changes and dermal tissue remodelling in diseased skin, and suggested that targeted action using selective MMP inhibitors may reduce lupus-induced damage in inflamed tissues. In conclusion, the results provide an insight into the genetics of CLE and demonstrate that genetic predisposition is at least in part shared between cutaneous and systemic variants of LE. This doctoral study has contributed IRF5, TYK2, ITGAM and several other novel genes to the so far short list of genes implicated in CLE susceptibility. Detailed examination of the function of these genes in CLE pathogenesis warrants further studies. Furthermore, the results support the need of subphenotype analysis with sample sizes large enough to reveal possible specific disease associations in order to better understand the heterogeneous nature and clinical specificities of the disease. Comprehensive analysis of clinical data suggests that smoking is an environmental triggering factor.

IgG4 Autoantibodies to DNA in Systemic Lupus erythematosus Patients

The presence of DNA-specific IgG4 antibodies was demonstrated in the sera of patients with systemic lupus erythematosus (SLE) by a microtiter solid-phase radioimmunoassay. A patient with distal inter-phalangeal swelling and extensive ulcers in the oral cavity, seronegative for anti-DNA antibodies of the IgG isotype, was found to have anti-DNA autoantibodies exclusively of the IgG4 subclass. These autoantibodies directed against the dsDNA conformation cross-reacted with chondroitin sulfate, dermatan sulfate and heparin.

Serological and Genetic Evidence for Altered Complement System Functionality in Systemic Lupus Erythematosus: Findings of the GAPAID Consortium

Systemic lupus erythematosus is a chronic autoimmune disease with multifactorial ethiopathogenesis. The complement system is involved in both the early and late stages of disease development and organ damage. To better understand autoantibody mediated complement consumption we examined ex vivo immune complex formation on autoantigen arrays. We recruited patients with SLE (n = 211), with other systemic autoimmune diseases (n = 65) and non-autoimmune control subjects (n = 149). Standard clinical and laboratory data were collected and serum complement levels were determined. The genotype of SNP rs1143679 in the ITGAM gene was also determined. Ex vivo formation of immune complexes, with respect to IgM, IgG, complement C4 and C3 binding, was examined using a functional immunoassay on autoantigen microarray comprising nucleic acids, proteins and lipids. Complement consumption of nucleic acids increased upon binding of IgM and IgG even when serum complement levels were decreased due to consumption in SLE patients. A negative correlation between serum complement levels and ex vivo complement deposition on nucleic acid autoantigens is demonstrated. On the contrary, complement deposition on tested protein and lipid autoantigens showed positive correlation with C4 levels. Genetic analysis revealed that the non-synonymous variant rs1143679 in complement receptor type 3 is associated with an increased production of anti-dsDNA IgG antibodies. Notwithstanding, homozygous carriers of the previously reported susceptible allele (AA) had lower levels of dsDNA specific IgM among SLE patients. Both the non-synonymous variant rs1143679 and the high ratio of nucleic acid specific IgG/IgM were associated with multiple organ involvement. In summary, secondary complement deficiency in SLE does not impair opsonization of nucleic-acid-containing autoantigens but does affect other antigens and potentially other complement dependent processes. Dysfunction of the receptor recognizing complement opsonized immune complexes promotes the development of class-switched autoantibodies targeting nucleic acids.

Polymerase chain reaction based C4AQ0 and C4BQ0 genotyping: association with systemic lupus erythematosus in southwest Han Chinese

Objective: To investigate the association of complement C4 null genes (C4QO, including C4AQO and C4BQO) and C2 gene with systemic lupus erythematosus (SLE) in southwest Han Chinese; 136 patients with SLE and 174 matched controls were genotyped. Methods: C4 null genes were determined by a polymerase chain reaction (PCR) procedure with sequence specific primers (PCR-SSP). The 2 bp insertion in exon 29, which was previously identified in non-Chinese populations and caused defective C4A genes, was directly typed by sequencing the whole exon 29 using exon specific primers. The exon 6 of complement C2 was also sequenced in both the patients and controls. Results: The frequency of homozygous C4AQO allele was 12.5% (17/136) in patients with SLE compared with 1.1% (2/174) in controls (p<0.001, odds ratio (OR)=12.286, 95% confidence interval (95% CI) 2.786 to 54.170). There was no significant difference for homozygous C4BQO allele between patients with SLE and controls (p=0.699). Patients with the C4AQO gene had an increased risk of acquiring renal disorder, serositis, and anti-dsDNA antibodies compared with those without C4AQO (for renal disorder, p=0.018, OR=8.951, 95% Cl 1.132 to 70.804; for serositis, p=0.011, OR 4.891, 95% CI 1.574 to 15.198; for anti-dsDNA, p=0.004, OR 7.630, 95%Cl 1.636 to 35.584). None of the patients or controls had the 2 bp insertion in exon 29 of the C4 gene. The type I C2 deficiency was not detected in the 3 10 samples. Conclusion: It is suggested that deficiency of C4A (not due to a 2 bp insertion in exon 29), but not C4B or C2, may be a risk factor for acquiring SLE in south west Han Chinese; this results in increased risk of renal disorder, serositis, and anti-dsDNA antibodies in patients with SLE. Racial differences seem to be relevant in susceptibility to SLE.

Lupus Eritematoso Sistémico

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Farmacêuticas