Introduction: The majority of kidney transplant (Tx) loss is related to development of donor-specific anti-HLA antibodies (HLA-DSA) and antibody-mediated rejection (ABMR). However, HLA-DSAs cannot explain all cases of ABMR, nor the differences observed in the outcome of kidney recipients with circulating DSAs endowed with similar biologic characteristics. Antibodies (Abs) directed to non-HLA antigenic targets may also have a role in ABMR and graft loss, and we could demonstrate how Abs to glutathione S-transferase theta 1 (GSTT1) protein were independent predictors of graft loss, and contributed to HLA-DSA-mediated damage, in our non-sensitized pediatric renal recipients. However, other groups have shown a negative impact of other non-HLA antigens on organ survival.  
Methods: We retrospectively analyzed humoral immune response to 6 non-HLA antigens, angiotensin II type 1 receptor (AT1R),Vinculin (VCL), Vimentin (VIM), Fibroblast Growth Factor-2 (FGF-2), Vascular Endothelial Growth Factor α (VEGFα), Epidermal growth factor-like 3 (EDIL-3), along with development of de novo (dn)HLA-DSAs and GSTT1-Abs, in a cohort of 146 pediatric nonsensitized recipients of first kidney allograft, to analyze their role in ABMR and graft loss. A multiplex bead assay was employed to assess non-HLA Abs on plasma samples collected before Tx, three-monthly in the first post-transplant year, and annually during a median 123 months follow-up.
Results: Among the 146 pediatric Tx recipients, 50 developed HLA-DSAs post-Tx, that persisted positive for the most part throughout the follow-up, and 132 developed 1 or more non-HLA Abs among those tested, above the positivity threshold. Of the latter group, 16 were found with GSTT1Abs at a MFI above the 75% quartile (Q4), 51 with 1 or more of the other non-HLA Abs (AT1R/VCL/VIM/FGF2/VEGFα/ EDIL3) at Q4, only 7 concomitantly positive for GSTT1. ABMR was diagnosed in 31 patients at a median follow-up of 5.0 years. Twenty-nine of the 31 patients had circulating dnDSAs, while two dnDSA-negative recipients were found positive for circulating anti-GSTT1 Abs but no other non-HLA Abs among those tested. Fifteen patients lost their graft at a median time of 6.7 years (range 2.3-14.4). Among the immunological parameters investigated, the presence of dnDSAs and of high MFI-level GSTT1 Abs, but not the presence of Q4 MFI level AT1R, VCL, VIM, FGF2, VEGFα, and EDIL3 Abs, were associated with graft loss. Cumulative incidence of graft survival in patients with positivity for GSTT1Abs was 42%, compared with 79% in GSTT1Abs-negative recipients and 93% in patients with positive AT1R,VCL,VIM, FGF2, VEGFα, and EDIL3 Q4 MFI level Abs (log-rank: p<0.000).     
Conclusions: According to our findings, the detection of antibodies against GSTT1 antigen, but not other non-HLA targets such as AT1R, VCL, VIM, FGF2, VEGFα, and EDIL3, increase immunologic risk and impact on graft survival in our cohort of non-sensitized pediatric renal recipients.