Distribution of de novo Donor-Specific Antibody Subclasses Quantified by Mass Spectrometry: High IgG3 Proportion Is Associated With Antibody-Mediated Rejection Occurrence and Severity

Abstract

Donor-specific antibodies (DSAs) are the main risk factor for antibody-mediated rejection (ABMR) and graft loss but could have variable pathogenicity according to their IgG subclass composition. Luminex-based test might lack sensitivity for the detection of IgG subclasses and this test does not allow quantifying the relative abundance of each IgG subclass. We investigated the precise repartition of each DSA subclass and their role in ABMR occurrence and severity, using an innovative mass spectrometry-based method. Between 2014 and 2018, we enrolled 69 patients who developed de novo DSA (n = 29 without ABMR, and n = 40 with ABMR) in two transplant centers. All IgG subclasses were detected in every samples tested: 62.7% were IgG1, 26.6% were IgG2, 6.6% were IgG3, and 4.2% were IgG4. The IgG3 proportion was significantly higher in the ABMR+ compared to the ABMR-group (8.4% vs. 5.6%, p = 0.003). The proportion of IgG1, IgG2, and IgG4 of DSA was similar between the two groups. Higher IgG3 level was associated with higher C4d deposition, higher microvascular inflammation scores, and glomerular filtration rate decline >25%. IgG3 proportion was not correlated with DSA MFI. Multivariate analysis showed that proteinuria and high level of IgG3 DSA were the only two factors independently associated with ABMR. In conclusion, de novo DSA are always composed of the four IgG subclasses, but in different proportions. High IgG3 proportion is associated with ABMR occurrence and severity and with poorer outcome, independently of DSA MFI.

IgG3 donor–specific antibodies with a proinflammatory glycosylation profile may be associated with the risk of antibody-mediated rejection after kidney transplantation

“…7,8,30 Using a more sensitive mass spectrometry-based method, the four IgG subclasses were detected for all DSA and a higher proportion of IgG3 subclass was associated with the occurrence and severity of ABMR, confirming our recently results reported. 32 These data suggest that the IgG3 subclass is particularly involved in the physiopathology of ABMR.…”

“…Relative quantification of the IgG subclasses was performed as previously described. 32 Peptides and proteins were identified using the ProteinPilot software v.5.0 (Sciex). Each IgG subclass was quantified using two unique proteotypic peptide sequences (Table S1).…”

“…IgG3 iDSA were isolated from the total IgG3 after binding on donorspecific single-antigen beads (Luminex FlowPRA ® , One Lamda, Inc) according to a method previously described. 32 First, the HLA beads coated with the donor-specific HLA antigen, the dnDSA target, were sorted by flow cytometry from a batch containing eight different HLA antigen beads. The donor-specific HLA beads were then incubated for 1 h with the purified total IgG3 before being washed three times with ammonium bicarbonate buffer.…”

Assessment of donor specific IgG Anti-HLA subclasses before and after kidney transplantation in Upper Egypt: A Single Center Study

“…40 In contrary to our findings, Pernin et al, 2020, assessed DSAs IgG subclasses in 69 KT recipients by using mass spectrometry after DSA isolation and that allowing their relative quantification. 15 They reported that all IgG subclasses were present in post-TX de novo DSA as IgG1 was predominant (62.2% of total IgG), IgG2 (23.1%), IgG3 (7.8%) and IgG4 (4.7%). Thus, IgG1, IgG3 and IgG4 fractions were high in alloimmune response than in total IgG and AMR occurrence with poor graft outcome, correlated with high IgG3 DSA percentage.…”

“…Some of them reported that presence of IgG3 is associated with poor graft outcome. 14,15 In the past decades, the short-term graft survival in KT recipients was improved, with no noticeable improvement in long-term graft survival. 16,17 The present work aimed to analyze the distribution of anti-HLA class I and class II PRA, IgG, and its subclasses in the pre and post KT period and their association with acute AMR and graft survival.…”