BIOLOGICALS TARGETING CYTOKINES INDUCING CD4 + T CELL SUBSET DIFFERENTIATION FOR THE TREATMENT OF EXPERIMENTAL ANTI-MPO GN H

PY GAN^1,2, A CHAN¹, J DICK^1,3, MA ALIKHAN¹, JD OOI¹, AR KITCHING^1,3, SR HOLDSWORTH^1,2,3

¹Department of Medicine, Centre for Inflammatory Diseases, Monash University, Clayton, Victoria; ²Department of Immunology, Monash Health, Monash Medical Centre, Clayton, Victoria; ³Medical Centre, Clayton, Victoria
Department of Nephrology, Monash Health, Monash 

Aim: To test the hypothesis that biological neutralization of the dominant T helper (T_H) cell subset inducing cytokine attenuates anti-MPO GN.

Background: Both CD4⁺ T_H17 and T_H1 cells, have been demonstrated to be pathogenic in anti-MPO GN. Definition of the dominant T_H subset would allow targeted biological treatment.

Methods: Anti-MPO autoimmunity was induced by MPO immunisation and GN triggered using anti-GBM Ig. T_H subsets in anti-MPO GN was defined by their cytokine profile in draining lymph nodes (dLN) and glomerular effector responses. T_H subsets was assessed early after induction of anti-MPO GN on day 20 (d20) and when disease was fully established (d32).

Results: In WT mice, anti-MPO GN progressively intensifies from d20 to d32 with increasing frequency of segmental necrosis (10±1vs17±2%, P<0.01) and albuminuria (1.22±0.2vs2.0±0.2mg/24hr, P<0.05). Analysis of T_H cytokines in dLN showed key T_H17 cytokines, IL-17A (2.8±0.1vs1.6±0.3ng/ml, P<0.05) and IL-6 were maximal at d20 and subsides by d32 while T_H1 dominant cytokines, IFNγ (19.9±8.1vs52.9±12.3pg/ml, P<0.05) and TNF-α were maximum at d32. Gene analysis of isolated kidney CD4⁺ T cells demonstrated that expression of T_H17 genes (IL-17A, CCR6, TGF-β) are more upregulated early while upregulation of T_H1 genes (IFNγ, CXCR3, STAT4) increases by d32. This suggests T_H17 dominates GN early and T_H1 late.

Treatment with monoclonal antibodies (mAb) to the T_H17 inducing cytokine, IL-23p19, attenuated GN (albuminuria; 105.3±28.2vsIgG2b controls; 325.4±60.5µg/24hr, P<0.01) on d20 but not on d32. Whereas mAb to T_H1 inducing cytokine, IL-12p35, was protective at d32 (albuminuria, 788.2±323.3vs IgG2a controls; 1797±229.9µg/24hr, P<0.05).

Conclusion: In anti-MPO GN, T_H subset dominance is biphasic, T_H17 early then T_H1 late. Anti-cytokine biological treatment is effective only when it targets the dominant T_H cytokines inducing anti-MPO GN.