Automated closed volume reduction process for apheresis stem cell grafts: From development to clinical implementation.

BACKGROUND: Collection of HPC by apheresis (HPC-A) can sometimes result in higher collection volumes, increasing the dimethyl sulfoxide (DMSO) volume infused into patients and the space requirements in liquid nitrogen freezers. Volume reduction prior to the addition of cryoprotectant is an efficient means to reduce the DMSO load infused into patients and to optimize freezer storage space. STUDY DESIGN AND METHODS: To implement a closed semi-automated volume reduction process, a method was developed to produce leukocyte-rich mock apheresis products using buffy coats derived from whole blood collections. The mock HPC products were then used to measure the efficiency and reliability of the semi-automated process over a range of volumes and cell concentrations. The resulting data was used to support the implementation of the process with concurrent monitoring. RESULTS: A closed, semi-automated volume reduction process resulted in recoveries of over 93% and 91% of white blood cells and CD34+ cells with no significant loss of product viability or potency. Mean doses of CD34+ and CFU infused per kilogram recipient body weight were 4.0 ± 1.1 × 106 /kg and 4.2 ± 1.7 × 105 /kg, resulting in no delays in median time to neutrophil and platelet engraftment, significant increase in adverse reaction or nonconformances. DISCUSSION: The effectiveness outcomes of the first Canadian experience in the implementation of a closed semi-automated volume reduction system in the processing of HPC-A products for autologous transplant have met the predetermined acceptance criteria, supporting its use in a stem cell manufacturing laboratory compliant with good manufacturing practice regulations.

A Bayesian adjustment for multiplicative measurement errors for a calibration problem with application to a stem cell study.

We develop a Bayesian approach to a calibration problem with one interested covariate subject to multiplicative measurement errors. Our work is motivated by a stem cell study with the objective of establishing the recommended minimum doses for stem cell engraftment after a blood transplant. When determining a safe stem cell dose based on the prefreeze samples, the postcryopreservation recovery rate enters in the model as a multiplicative measurement error term, as shown in the model. We examine the impact of ignoring measurement errors in terms of asymptotic bias in the regression coefficient. According to the general structure of data available in practice, we propose a two-stage Bayesian method to perform model estimation via R2WinBUGS (Sturtz, Ligges, and Gelman, 2005, Journal of Statistical Software 12, 1-16). We illustrate this method by the aforementioned motivating example. The results of this study allow routine peripheral blood stem cell processing laboratories to establish recommended minimum stem cell doses for transplant and develop a systematic approach for further deciding whether the postthaw analysis is warranted.

Cryopreserved mobilized autologous blood progenitors stored for more than 2 years successfully support blood count recovery after high-dose chemotherapy.

BACKGROUND AIMS: The ability of hematopoietic progenitor cells-apheresis (HPC-A) that have been stored for many years after cryopreservation to reconstitute hematopoiesis following high-dose chemo/radiotherapy has not been well-documented. METHODS: In this retrospective study, eight Canadian centers contributed data from 53 autologous stem cell transplants (ASCT) performed using HPC-A that had undergone long-term storage (>2 years, range 2-7 years) and 120 ASCT using HPC-A stored for <6 months (short-term storage). RESULTS: The doses of nucleated and CD34(+) cells per kilogram recipient weight were similar between the short- (mean ± SD, 4.7 ± 4.9 × 10(8) and 6.8 ± 4.3 × 10(6), respectively) and long- (4.0 ± 4.9 × 10(8) and 6.1 ± 3.4 × 10(6), respectively) term storage groups. The median days to neutrophils (absolute neutrophil count; ANC) >0.5 × 10(9)/L (median 11 days for both short- and long-term storage) and platelets >20 × 10(9)/L (median 12 and 11 for short- and long-term storage, respectively) post-ASCT were not significantly different between the two groups. When ASCT performed with <5 × 10(6)/kg CD34(+) cells was compared there was also no difference in ANC or platelet recovery (median 12 days for both after short-term storage, and 12 and 11 days, respectively, after long-term storage). Fourteen HPC-A products stored for >5 years also showed similar count recoveries as the entire long-term storage group (median 11 days for both ANC and platelets). CONCLUSIONS: Cryopreserved HPC-A can be stored for at least 5 years with no apparent loss in their ability to support hematopoietic reconstitution after high-dose chemotherapy.

CD34+ cell responsiveness to stromal cell-derived factor-1alpha underlies rate of engraftment after peripheral blood stem cell transplantation.

BACKGROUND: Stromal cell-derived factor (SDF)-1, a chemokine produced in the bone marrow (BM), is essential for the homing of hematopoietic stem/progenitor cells (HSPCs) to the BM after transplantation. This study examines whether there is a correlation between the in vitro chemotaxis of CD34+ HSPC toward an SDF-1 gradient and in vivo hematopoietic engraftment. STUDY DESIGN AND METHODS: Thirty-five patients underwent granulocyte-colony-stimulating factor HSPC collection and autologous transplant with a median dose of 7.7 (range, 3.9-41.5) x 10(6) CD34+ cells per kg body weight. The chemotactic index (CI) of CD34+ cells isolated from leukapheresis products collected from these patients was calculated as the ratio of the percentages of cells migrating toward an SDF-1 gradient to cells migrating to media alone. Expression of the SDF-1 receptor CXCR4 on CD34+ cells was measured by flow cytometry. RESULTS: Spontaneous cell migration (range, 3.1 +/- 0.6 to 26.5 +/- 7.7%) and SDF-1-directed chemotaxis (11.1 +/- 0.7 to 54.9 +/- 8.3%) of CD34+ cells did not correlate with time to neutrophil engraftment, which occurred at a median of 10 days (range, 8-16 days). Nonparametric tests showed a negative correlation (r = -0.434) between CI and CD34+ cell dose such that neutrophil recovery occurred within the same period in patients transplanted with a lower dose of CD34+ cells but having a high CI as in those transplanted with a higher dose of CD34+ cells but having a low CI. Moreover, CI correlated (r = 0.8) with surface CXCR4 expression on CD34+ cells. CONCLUSION: In patients transplanted with a relatively lower CD34+ cell dose who achieved fast engraftment, a higher responsiveness to SDF-1 and high CI could have compensated for the lower cell dose. However, to apply the CI as a prognostic factor of the rate of engraftment requires validation in a larger number of patients.