Further Validation of Flow Adhesion of Whole Blood to P-Selectin (FA-WB-PSEL) to Assess Targeted P-Selectin Therapies in Sickle Cell Disease (SCD)

Sickle cell disease (SCD) results from a mono-genetic mutation in the hemoglobin (Hb) gene that promotes frequent sickling-unsickling events which causes destabilization of RBC membranes and cell lysis, elevated plasma free Hb, and subsequent oxidative stress, inflammation, reticulocytosis, abnormal RBC adhesive interactions and, RBC trapping in small blood vessels. Great strides have been witnessed in recent years with the approval of five SCD-modifying therapies targeting distinct mechanisms nearly a half century following the first use of hydroxyurea (HU) to reduce the frequency of VOEs by inducing fetal hemoglobin (HbF) levels in sickle RBCs.

Lack of reliable, blood-based biomarkers to objectively define VOEs in SCD burdens providers to prescribe therapies heavily based on subjective, patient-reported pain crises. We developed RBC health biomarkers to objectively define SCD clinical state at baseline, during crises, and while on therapy. P-selectin is a major contributor to micro-vascular occlusion and was the 1^st direct therapeutic target to inhibit vaso-occlusive adhesive events that precede VOEs in SCD using Adakveo, a humanized monoclonal antibody against P-selectin. The objective of this study was to confirm the value of our flow adhesion of whole blood to P-selectin (FA-WB-PSEL) biomarker in distinguishing Adakveo responders from non-responders and monitoring patient response to therapy in a real-world clinical setting.

This study is a retrospective assessment of FA-WB-PSEL in SCD patients from 5 US clinics between 2018-2024 on/off Adakveo therapy. Biomarkers were ordered as standard, clinical send-out (CSO) tests and an IRB-approved waiver of informed consent permitted access to patient medical records. We compared FA-WB-PSEL levels in blood samples from Adakveo-treated and non-treated patients and, individual SCD patients that received at least 3 biomarker evaluations within 1-year pre- and 1-year post-Adakveo treatment.

We received 5507 clinical samples from 994 SCD patients; 1160 samples were collected during treatment from 61 patients as reported by clinicians on sample requisition forms and, 14 patients with confirmed Adakveo therapy start dates. Non-treated samples (n=4347) were significantly more adherent than Adakveo samples (n=1160; 56.4±52 to 51.7±45, p=0.0001) and the proportion of FA-WB-PSEL levels measuring above the critical threshold (50 cells/mm²) was significantly reduced from 42.9% to 39.5% (p=0.039). Similarly, Adakveo significantly reduced the adhesive phenotype of WB samples obtained from 61 SCD patients (n_Adakveo=1014; n_non-treated=1159; 61±54 cells/mm² to 52±45 cells/mm², p=0.0071) as well as critical biomarker levels (46.6% to 40%, p=0.0021) and, the proportion of patients with severe pain (pain score >=7 in a 0-10 scale) decreased significantly from 92.2% to 85.7% (p=0.0000). There were 36 of 61 patients with FA-WB-PSEL levels within normal range (<50 cells/mm²); FA-WB-PSEL levels and pain scores were not affected by treatment. However, 25 patients with critical FA-WB-PSEL levels demonstrated significant decreases in adhesion (76±25 to 49±19 cells/mm², p<0.05).

Provider-confirmed therapy start dates identified 14 unique SCD patients with significantly reduced FA-WB-PSEL levels post-treatment (mean_pre=69.97±10.01 cells/mm², mean_post=45.93±5.14 cells/mm², p=0.0494) as previously reported and, non-treated FA-WB-PSEL levels strongly correlated with patient response to therapy (r=-0.7877, p=0.0013). Also, SCD patients (n=10) exhibiting high P-selectin activity (>50 cells/mm²) in the pre-treatment period were significantly more responsive to Adakveo 1-year post-treatment (mean_pre=89.35±7.31 cells/mm², mean_post=50.61±5.05 cells/mm², p=0.001) and biomarker levels stabilize post-treatment (SD_pre=60.27±4.59 cells/mm², SD_post=34.19±5.45 cells/mm², p=0.001) whereas, the low P-selectin activity cohort was unaffected (data not shown).

These data further support the use of our FA-WB-PSEL biomarker to distinguish Adakveo responders from non-responders and monitor patient response to P-selectin targeted therapies in SCD and beyond. Ongoing studies are underway to validate FA-WB-PSEL as a surrogate endpoint for VOEs in clinical trials and in the real-world clinical setting.