The Role of Red Blood Cell Vascular Adhesion Biomarkers in Understanding Sickle Cell Disease Associated Chronic Pain

Background: Sickle cell disease (SCD) is characterized by repeated episodes of vaso-occlusion (VOC) that can lead to a host of complications including acute and chronic pain. Prior studies have demonstrated acute changes in RBC health biomarker flow adhesion of whole blood to VCAM-1 (FA-WB-VCAM) and P-Selectin (FA-WB-Psel). Adhesive indices are significantly elevated during a proadhesive VOC. We examined whether individuals with SCD with and without chronic pain will exhibit differences in RBC health biomarkers between steady state and acute VOC pain.

Methods: This was a 6-month, longitudinal, case-control cohort pilot study of FA-WB-VCAM and FA-WB-Psel indices collected in youth with SCD at Children's National Hospital in Washington, DC. SCD chronic pain patients (as defined per AAPT diagnostic criteria) were classified as “cases,” and SCD patients without chronic pain were “controls.” Controls were age (± 2 years), and genotype matched to cases. Both FA-WB-VCAM and FA-WB-Psel were collected at baseline and during acute VOC with assays performed by Functional Fluidics Inc. in Detroit, MI. Patient-reported outcomes were captured as pain scores and through completion of the centralized pain index survey to assess for centralized pain features of chronic pain. Statistical analysis included descriptive statistics and categorical analysis by t-test (GraphPad, v9, 2021). P value <0.05 was statistically significant.

Results: We identified 10 pediatric patients, 4 with chronic pain “cases” and 6 without chronic pain “controls” with FA-WB-VCAM and FA-WB-Psel assays performed at baseline and during an acute VOC episode. The average age of the cohort was 17.5 years with 70% females (n=7) and 30% males (n=3). Eighty percent (n=8) of the cohort were HgbSS and 20% (n=2) HgbSC genotype. Twenty-five percent (n=1) of the chronic pain patients were on disease-modifying therapy (DMT) with hydroxyurea (HU) at study initiation [all patients had been offered ≥ 1 FDA-approved DMT and previously discontinued DMT use due to intolerable side effects or personal choice], whereas 83% (n=5) of the control patients were on DMT with 50% (n=3) being on dual DMT therapy (HU + voxelotor or HU + crizanlizumab). The chronic pain cohort had a higher average number of ED visits/year than the control cohort (14 ± 7 vs. 4 ± 2 ED visits/year, p=0.11). The average baseline pain score for the chronic pain cohort was 7.25 versus 1.16 in the control cohort (p= 0.069). Centralized pain index survey responses were similar between both groups [median (IQR): 11.5 (7.25 - 15) vs. 9 (7-16), p=0.72)]. Figures 1A and 1B show a comparison of FA-WB-VCAM and FA-WB-Psel at baseline versus acute VOC for each chronic pain (P) “case” with their respective “controls (C).” The average baseline FA-WB-VCAM (P: 555.8 ± 605.3 vs. C: 435.3 ± 217 cells/mm2, p=0.905) and the average baseline FA-WB-PSel (P: 83.5 ± 64.5 vs. C: 57.33 ± 45.4, p=0.833) were similar between the cohort of chronic pain versus control subjects. Acute VOC FA-WB-VCAM (P: 722 ± 366.6 vs. C: 553.5 ± 309.6 cells/mm2, p=0.667) and FA-WB-PSel (P: 116.3 ± 66.6 vs. C: 98.7 ±96 cells/mm2, p=0.777) adhesion values were similar between cases and controls although adhesion values trended higher than baseline for both groups. All the cases had acute VOC adhesion indices above clinically established critical thresholds [Figure 1A and 1B]. The FA-WB-VCAM adhesion values increased from baseline in 75% (n=3) of the chronic pain patients (#3, #14, #16) during acute VOC. All the chronic pain patients (#3, #7, #14, #16) showed increased FA-WB-Psel adhesion from baseline during acute VOC [Figure 1A and 1B]. Fifty percent (n=3) of the control patients (#10, #12, #28) exhibited increased FA-WB-VCAM adhesion during acute VOC, and 60% (n=4) of the controls (#4, #5, #10, #28) exhibited increased FA-WB-Psel adhesion during acute VOC.

Conclusions: We showed changes in RBC health vascular adhesion markers from baseline to acute VOC in patients with and without chronic pain. FA-WB-VCAM and FA-WB-Psel trended higher in the chronic pain cohort and all acute VOC values for the chronic pain cohort were above established critical thresholds. FA-WB-Psel adhesion increased from baseline with the onset of acute VOC episodes for most of the chronic pain cohort. Mechanistically, there is a need to understand differences in acute versus chronic pain, which may provide further insight into pro-adhesive versus non-vascular pain mechanisms.