Background: In PNH, a somatic mutation in the PIGA gene of one or a few hematopoietic stem cells generates a clone of abnormal erythrocytes which lack two key alternative pathway (AP) regulatory proteins, CD55 and CD59, leading to uncontrolled complement activation on affected erythrocytes (RBCs) and membrane attack complex (MAC)-mediated lysis.

Factor D (FD), a serine protease, catalyzes the cleavage of complement factor B into Ba and Bb, which allows for the formation of the AP C3 convertase. By inhibiting FD, danicopan, an oral small molecule FD inhibitor, blocks C3 convertase formation, the control point for AP activation as well as the amplification of all pathways. This leads to inhibition of C3 cleavage, C3 fragment deposition, terminal pathway activation and MAC formation. Therefore, FD is a promising target in diseases of excess activation of the AP, such as PNH.

Aims: Correlation between complement activity biomarkers and clinical/laboratory efficacy of danicopan, administered as monotherapy, in the treatment of PNH. Given that danicopan is potential first in class AP inhibitor for the treatment of PNH, it is important to investigate changes and relationships among relevant biomarkers of the AP to better understand the mechanism of action of danicopan in vivo.

Methods: Danicopan starting doses ranged from 100-150 mg Q8H, with subsequent dose escalation based on clinical and biochemical response to doses as high as 200 mg Q8H. Danicopan plasma concentrations were determined by liquid chromatography/tandem mass spectrometry (LC/MS-MS). Danicopan pharmacodynamics (PD) were determined by measuring serum AP activity (AP Wieslab). Plasma Bb concentration, serum FD concentration, serum complement C3 and C4 concentrations, and serum classical pathway (CP) activity were also measured. C3 fragment deposition on RBCs was measured by flow cytometry with FITC conjugated anti- human C3d antibody.

Results: Clinical results were previously presented, Table 1. Pharmacokinetic (PK) and PD analysis of danicopan concentration and AP activity measured ex vivo showed a clear correlation: ≤10% AP activity was seen for 6 hours following dosing with some recovery in AP activity prior to the next danicopan dose, Figure 1. Mean baseline Bb levels in this untreated PNH population were elevated relative to the reference range, demonstrating ongoing AP activity. Bb levels decreased into normal range with danicopan, demonstrating decreased in vivo C3 convertase formation, Figure 2. A strong positive correlation existed between Bb concentration and LDH level, demonstrating Bb as a reliable biomarker of AP activation in vivo during therapeutic complement inhibition in PNH. Strong correlations were also observed between danicopan concentration with Bb concentration (positive), AP activity (negative) and LDH level (positive), validating the role of danicopan in the changes of these endpoints. As expected, serum FD concentration was unchanged throughout treatment as was CP activity, which was measured with an in vitro assay designed to exclude any AP contribution. Notably, in contrast with C5 inhibition, C3 fragment deposition was not observed (<0.1 % of RBCs) with danicopan. The slight increase in serum C3 concentration is likely due to blockade of constitutive C3 consumption which may be elevated in PNH due to impaired control of complement activation on RBC surface.

Conclusion: Previously reported proof of concept study with oral danicopan has demonstrated that upstream complement inhibition at the level of FD can prevent MAC-mediated intravascular hemolysis even in the absence of …