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Inhibition of the Complement Alternative Pathway by Chemically Modified DNA Aptamers That Bind with Picomolar Affinity to Factor B.

ABSTRACT: The complement system is a conserved component of innate immunity that fulfills diverse roles in defense and homeostasis. Inappropriate activation of complement contributes to many inflammatory diseases, however, which has led to a renewed emphasis on development of therapeutic complement inhibitors. Activation of complement component C3 is required for amplification of complement and is achieved through two multisubunit proteases called C3 convertases. Of these, the alternative pathway (AP) C3 convertase is responsible for a majority of the C3 activation products in vivo, which renders it an attractive target for inhibitor discovery. In this study, we report the identification and characterization of two related slow off-rate modified DNA aptamers (SOMAmer) reagents that inhibit formation of the AP C3 convertase by binding to the proprotease, factor B (FB). These aptamers, known as SL1102 (31 bases) and SL1103 (29 bases), contain uniform substitutions of 5-(N-2-naphthylethylcarboxyamide)-2'-deoxyuridine for deoxythymidine. SL1102 and SL1103 bind FB with K _d values of 49 and 88 pM, respectively, and inhibit activation of C3 and lysis of rabbit erythrocytes under AP-specific conditions. Cocrystal structures of SL1102 (3.4 Å) and SL1103 (3.1 Å) bound to human FB revealed that SL1102 and SL1103 recognize a site at the juncture of the CCP1, CCP3, and vWF domains of FB. Consistent with these structures and previously published information, these aptamers inhibited FB binding to C3b and blocked formation of the AP C3 convertase. Together, these results demonstrate potent AP inhibition by modified DNA aptamers and expand the pipeline of FB-binding molecules with favorable pharmacologic properties.

SUBMITTER: Xu X

PROVIDER: S-EPMC7851746 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Inhibition of the Complement Alternative Pathway by Chemically Modified DNA Aptamers That Bind with Picomolar Affinity to Factor B.

Xu Xin X Zhang Chi C Denton Dahlia T DT O'Connell Daniel D Drolet Daniel W DW Geisbrecht Brian V BV

Journal of immunology (Baltimore, Md. : 1950) 20210108 4

The complement system is a conserved component of innate immunity that fulfills diverse roles in defense and homeostasis. Inappropriate activation of complement contributes to many inflammatory diseases, however, which has led to a renewed emphasis on development of therapeutic complement inhibitors. Activation of complement component C3 is required for amplification of complement and is achieved through two multisubunit proteases called C3 convertases. Of these, the alternative pathway (AP) C3 ...[more]

PMID: 33419768

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Project description:The innate immune system plays a major role in the pathogenesis of nonalcoholic steatohepatitis (NASH). Recently we reported complement activation in human NASH. However, it remained unclear whether the alternative pathway of complement, which amplifies C3 activation and which is frequently associated with pathological complement activation leading to disease, was involved. Here, alternative pathway components were investigated in liver biopsies of obese subjects with healthy livers (n?=?10) or with NASH (n?=?12) using quantitative PCR, Western blotting, and immunofluorescence staining. Properdin accumulated in areas where neutrophils surrounded steatotic hepatocytes, and colocalized with the C3 activation product C3c. C3 activation status as expressed by the C3c/native C3 ratio was 2.6-fold higher (p<0.01) in subjects with NASH despite reduced native C3 concentrations (0.94±0.12 vs. 0.57±0.09; p<0.01). Hepatic properdin levels positively correlated with levels of C3c (rs?=?0.69; p<0.05) and C3c/C3 activation ratio (rs?=?0.59; p<0.05). C3c, C3 activation status (C3c/C3 ratio) and properdin levels increased with higher lobular inflammation scores as determined according to the Kleiner classification (C3c: p<0.01, C3c/C3 ratio: p<0.05, properdin: p<0.05). Hepatic mRNA expression of factor B and factor D did not differ between subjects with healthy livers and subjects with NASH (factor B: 1.00±0.19 vs. 0.71±0.07, p?=?0.26; factor D: 1.00±0.21 vs. 0.66±0.14, p?=?0.29;). Hepatic mRNA and protein levels of Decay Accelerating Factor tended to be increased in subjects with NASH (mRNA: 1.00±0.14 vs. 2.37±0.72; p?=?0.22; protein: 0.51±0.11 vs. 1.97±0.67; p?=?0.28). In contrast, factor H mRNA was downregulated in patients with NASH (1.00±0.09 vs. 0.71±0.06; p<0.05) and a similar trend was observed with hepatic protein levels (1.12±0.16 vs. 0.78±0.07; p?=?0.08). Collectively, these data suggest a role for alternative pathway activation in driving hepatic inflammation in NASH. Therefore, alternative pathway factors may be considered attractive targets for treating NASH by inhibiting complement activation.

Dataset Information

Inhibition of the Complement Alternative Pathway by Chemically Modified DNA Aptamers That Bind with Picomolar Affinity to Factor B.

Publications

Inhibition of the Complement Alternative Pathway by Chemically Modified DNA Aptamers That Bind with Picomolar Affinity to Factor B.

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