A Novel Direct Hemoglobin S Polymerization Inhibitor for the Treatment of Sickle Cell Disease - <i>In Vivo</i> Efficacy of Ilx-002 in Humanized Mice

Background - Current approved treatments for sickle cell disease (SCD), excluding bone marrow transplant and recent gene therapies, fail to adequately control hemolysis and prevent painful vaso-occlusive crises (VOCs). ILX-002 is a novel oral drug candidate that directly inhibits hemoglobin S (HbS) polymerization by mimicking the protective effect of a rare naturally occurring HbS variant called Hb Stanleyville II (HbSAsn78Lys). By binding to HbS and disrupting critical stabilizing contacts between HbS molecules on the surface of the αF-helix, ILX-002 directly blocks polymer formation without dramatically altering the oxygen affinity of hemoglobin. ILX-002 has the potential to become a once daily pill that is a functional cure for SCD. The objective of this study was to evaluate the impact of ILX-002 on key hematologic parameters in vivo in a humanized mouse model of SCD. Methods - Female humanized homozygous βS/βS Townes mice (Jackson Lab), 8-12 weeks of age, were allowed to acclimate for at least 5 days, and then randomized to one of five groups (n=5 per group). Animals received either vehicle or ILX-002 (0.5%, 1%, 1.5%, or 2% w/w) administered by non-forceful voluntary administration in chow for 27 days. On Day 27, mice from all groups underwent terminal blood collection for hematological and biochemical analyses and were humanely sacrificed. Spleens were also collected and weighed. Level of drug in the blood was determined by HPLC, and drug hemoglobin occupancy was assessed by cation exchange HPLC based on the relative area under the curve of drug-modified HbS and native HbS peaks. A comprehensive complete blood count with automated reticulocyte count was also performed (IDEXX Labs). Results - ILX-002 demonstrated dose-dependent improvements in all measured hematologic parameters. Notably, all mice achieving at least 40% drug occupancy (blood level approximately ≥ 500 µM) showed normalization of hemoglobin levels to at least 13 g/dL, similar to Townes AA and AS mice, compared to 7.7 g/dL for the vehicle group. The reticulocyte percentage decreased from 34.4% in the vehicle group to 14.2% in the high dose group, and average platelet count increased from 696 K/µL to 1182 K/µL. Spleen weights decreased on average by 41% from 1.1 grams in the vehicle group to 0.65 grams for animals with at least 40% drug occupancy. The treatment administered in chow was well tolerated by all groups. Conclusions - ILX-002 shows promise as a transformational oral therapy for SCD, potentially offering durable control of hemolysis and VOC prevention. Its unique mechanism of direct HbS polymerization inhibition, coupled with a minimal impact on hemoglobin oxygen equilibrium, suggests a favorable efficacy and safety profile. The dramatic disease-modifying effects observed in Townes SS mice at clinically achievable drug levels support ILX-002's potential to significantly improve patient outcomes. These compelling preclinical results warrant rapid advancement of ILX-002 into human trials.