Background/Purpose: DNASE1L3 is a unique secreted DNase that is capable of degrading DNA complexed with proteins and/or encapsulated in membranes, such as chromatin within microparticles from apoptotic cells. The role of DNASE1L3 in the pathogenesis of SLE is supported by the nearly universal association of DNASE1L3 null mutations with familial SLE, the linkage of a hypomorphic variant to sporadic SLE, and the development of anti-DNA response and SLE-like disease in DNASE1L3-deficient mice. Accordingly, this study was initiated to delineate the role of microparticle-associated antigens sensitive to DNASE1L3 digestion as targets of antibody reactivity in human SLE.

Methods: Antibodies to DNA on microparticles were measured using an in vitro assay. Jurkat human T cells were treated with staurosporine to induce apoptosis, and microparticles were isolated. After mock-treatment or treatment with recombinant DNASE1L3, the microparticles were subsequently incubated with sera, washed, and stained with fluorescently labeled anti-human IgG prior to evaluation using flow cytometry. The ratio of binding values with or without DNASE1L3 pre-treatment indicates the binding to DNASE1L3-sensitive antigens (including DNA and/or DNA-associated proteins) versus other antigens exposed on microparticles.

Results: Evaluation of sera (76 from 66 SLE patients, 19 controls, 34 anti-Ro positive neonatal lupus mothers) revealed three types of reactivity to microparticles: i) no binding; ii) binding insensitive to DNASE1L3; and iii) binding that is sensitive to DNASE1L3. Notably, the only two patients heterozygous for the DNASE1L3 (R206C) hypomorphic variant belonged to this latter group. Samples from 30 (45%) of the SLE patients showed DNASE1L3-sensitive binding while this reactivity was absent in all controls. Although hypomorphic allelic variants are restricted to European ancestry, sensitive binding was observed in all ethnic/racial groups. SLE patients with sensitive vs insensitive binding had a significantly higher frequency of past/present renal disease (87% vs 44%, p=0.0007). Moreover, at the time of blood sampling, sensitive binding was associated with the presence of >0.5 uPCR (61% vs 30%, p=0.012) and low complement levels (78% vs 53%, p=0.031). Overall, sensitive binders were more active with SLEDAI score ≥8 in 47% vs 21% (p=0.007). Supporting that DNASE1L3 can disrupt pre-formed complexes of IgG with antigen on microparticles, binding to the native microparticles could be abrogated by subsequent addition of the enzyme. Several subjects within the group of high titer anti-Ro positive mothers (asymptomatic, UAS, SS, SLE but no history of nephritis) showed strong binding to native microparticles, but this binding was not DNASE1L3-sensitive.

Conclusion: These data support that reactivity to physiological DNA-associated antigens from apoptotic cells is relevant in lupus nephritis and suggest DNASE1L3 as a novel pharmacological approach to forestall organ injury.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/dysfunction-of-the-dnase1l3-pathway-and-antigen-accumulation-in-lupus-nephritis/