University of Florida Ocala, Florida, United States
Background/Case Studies: Hematopoietic stem cell transplant has been the leading therapeutic for patients with hematological disorders. However, only 30 percent of patients find an HLA-match within their family, with even lower matching with unrelated donors1. Donated cord blood (CB) can fulfill the need for the remaining patients often from racially and ethnically diverse groups2. However, the donated units are limited in cell dosage, critical for engraftment time and overall survival. Indeed, methods for cellular manufacturing are increasingly being sought as a new solution. By validating our 7-day ex vivo platform using hPM to assess the proliferation and expansion of human CD34+ HPC(CB), we aim to develop a new biomanufacturing process.
Study
Design/Methods: As previously described, hPM was manufactured using placenta material sourced from the local hospital3. Purchased CD34+ cells were retained unmanipulated; cultured with either control media (StemSpan plus 100 ng/mL each SCF, TPO, and Flt3L); or hPM treatment (control plus prepared hPM) for 7-days before processing (n=5). Animal studies were carried out according to our institution’s guidelines using NSG mice (Strain #026622) (n=5/treatment). Peripheral blood analysis was performed at 12- and 16- week. Flow cytometry and CFUs assays were performed to assess HPCs activity (Fig 1). Cell count and viability was performed using trypan blue exclusion assay. Statistics were performed using one-way ANOVA or student t-test.
Results/Findings: This study showed a 25-fold increase in HPC CD34+ cells cultured ex vivo using hPM as a media constitution (Fig.1A). Our results indicated a 35-fold increase in CD34+CD45RA-CD38- HPCs (Fig. 1B), showing hPM cultured CD34+ cells capacity to remain phenotypically primitive. Preliminary assessment of human engraftment indicated nominal levels at 12- and 16- weeks, (10% and 18%; SD +/ 5.4, 5.6) respectively, compared to control media (13% and 35.2%; SD +/ 6.6, 21.2) (Fig 1D).
Conclusions: Interestingly, while hPM treated cells displayed exponential expansion, lower engraftment was observed. We surmise hPMs extrinsic factors impacted key HPCs migratory regulators. Indeed, our ongoing assessments of hPM’s impact on these critical regulators may lead to new therapeutic pipelines. These findings support further exploration of hPM-derived biologics for biomanufacturing. The innovative use of hPM for ex vivo expansion has unveiled a new sector of therapeutic applications for clinical cellular therapies.
