Validation of an enforced change in cryopreservation method after hydroxyethylstarch (HES) withdrawal: Engraftment kinetics of 1007 autograft procedures

      Quality assurance of HPC collection, storage, cryopreservation, thawing and ultimately potency of the cells for an autograft program is defined by engraftment kinetics. From 1995 the cryoprotectant method at UCLH used final concentrations of HES, DMSO and human serum albumin (HSA) of 5%, 5% and 4% respectively (HES method). The withdrawal of HES for clinical use in June 2013 compelled change and an equal dilution of the product with 20%DMSO in 4.5% HSA (10%DMSO method) was selected. Thaw CFU-GM yields of ten paired PBSC samples cryopreserved by both methods were in close accord with a median (range) of 72 (37–160%) versus 72 (43–154%) for 10%DMSO and HES respectively, R2 = 0.904 (P = 0.69) and the 10% DMSO method was adopted. Subsequent engraftment times appeared satisfactory on a case by case basis but subsequent haematological recovery of 1007 consecutive autograft procedures using HES (n = 623) or the replacement 10%DMSO method (n = 384) permitted final validation. The series (Aug 2007 to Sep 2016) included 33 tandem autografts and applies to 974 patients. The distribution of diagnostic groups between cryoprotectants was similar (Table I). Engraftment was censored for 7 patients who died before neutrophil recovery and for a further 9 patients for platelet recovery (7 early deaths, 2 lost to follow up). The median (range) CD34 dose infused was 3.0 (0.7–27) × 106/kg and 3.0 (0.5–46) × 106/kg for 10% DMSO and HES respectively (p = NS) and no significant differences were seen in cumulative recovery curves for days to neutrophils 0.5 × 109/l or days to an unsupported platelet count of 20 × 109/l (Figure 1). The median (range) days to neutrophil recovery was 11 (3–20) days vs 11 (6–23) days and for platelet recovery 12 (0–50) days vs 11 (0–67) days for 10%DMSO and HES respectively. Neutrophil recovery occurred within 14 days in over 99% of all patients and between 15–28 days in only 7 (0.7%). Platelet recovery occurred in 84% of all patients and was delayed (>28 days) in 35 (3.5%). The CD34 dose infused was the most significant factor affecting neutrophil and platelet recovery independently of cryoprotectant and the proportion of patients with early recovery (10 days or less) significantly increased at each higher CD34 dose (P < 0.05 to <0.0001) (Figure 1) delayed platelet recovery did not occur above a CD34 dose of 5 × 106/kg. These data suggest that close monitoring and audit of engraftment data is essential to assure continued control of clinical and laboratory performance.
      Table IPatient demographics and cryoprotectant use.
      Cryoprotectant: 10%DMSO HES
      Patient no. 379 595
      Diagnosis n(%) n(%)
      Myeloma 185(49%) 282(47%)
      NHL 125(33%) 220(37%)
      Hodgkin 44(12%) 59(10%)
      Solid Tumour 22(6%) 32(5%)
      Acute Leukaemia 3(0.8%) 2(0.3%)
      Male 244(66%) 363(61%)
      Female 135(36%) 232(39%)
      Age Median (range) 56(13–75) yrs 55(13–71) yrs
      Figure 1
      Figure 1Cumulative recovery curves of 1000 autograft procedures. A) and B) ANC to 0.5 × 109/l and unsupported platelet recovery to 20 × 109/l according to cryoprotectant. C) and D) increasing proportions of patients with early recovery (10 days or less) with higher CD34 doses infused.
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