Search

Aims The purpose of our study was to determine whether mesenchymal stem cells (MSCs) are an effective and safe therapeutic agent for the treatment of knee osteoarthritis (OA), owing to their cartilage regeneration potential. Methods We searched PubMed, Embase, and the Cochrane Library, with keywords including “knee osteoarthritis” and “mesenchymal stem cells”, up to June 2019. We selected randomized controlled trials (RCTs) that explored the use of MSCs to treat knee OA. The visual analogue scale (VAS), Western Ontario and McMaster University Osteoarthritis Index (WOMAC), adverse events, and the whole-organ MRI score (WORMS) were used as the primary evaluation tools in the studies. Our meta-analysis included a subgroup analysis of cell dose and cell source. Results Seven trials evaluating 256 patients were included in the meta-analysis. MSC treatment significantly improved the VAS (mean difference (MD), –13.24; 95% confidence intervals (CIs) –23.28 to –3.20, p = 0.010) and WOMAC (MD, –7.22; 95% CI –12.97 to –1.47, p = 0.010). The low-dose group with less than 30 million cells showed lower p-values for both the VAS and WOMAC. Adipose and umbilical cord–derived stem cells also had lower p-values for pain scores than those derived from bone marrow. Conclusion Overall, MSC-based cell therapy is a relatively safe treatment that holds great potential for OA, evidenced by a positive effect on pain and knee function. Using low-dose (25 million) and adipose-derived stem cells is likely to achieve better results, but further research is needed. Cite this article: Bone Joint Res 2020;9(10):719–728.

Mesenchymal stromal cells (MSCs) currently constitute the most frequently used cell type in advanced therapies with different purposes, most of which are related with inflammatory processes. Although the therapeutic efficacy of these cells has been clearly demonstrated in different disease animal models and in numerous human phase I/II clinical trials, only very few phase III trials using MSCs have demonstrated the expected potential therapeutic benefit. On the other hand, diverse controversial issues on the biology and clinical applications of MSCs, including their specific phenotype, the requirement of an inflammatory environment to induce immunosuppression, the relevance of the cell dose and their administration schedule, the cell delivery route (intravascular/systemic vs. local cell delivery), and the selected cell product (i.e., use of autologous vs. allogeneic MSCs, freshly cultured vs. frozen and thawed MSCs, MSCs vs. MSC-derived extracellular vesicles, etc.) persist. In the current review article, we have addressed these issues with special emphasis in the new approaches to improve the properties and functional capabilities of MSCs after distinct cell bioengineering strategies.

Hematopoietic cell transplants using stem cells from umbilical cord blood are used worldwide for the treatment of malignant and non-malignant disorders. Transplant procedures from this stem cell source have shown promising outcomes in successfully treating various hematologic, immunologic, malignant, and inherited metabolic disorders. Rapid availability of these stem cells is an important advantage over other unrelated donor transplants, especially in situations where waiting can adversely affect the prognosis. The umbilical cord blood is rich in CD34+ stem cells, though with a limited cell dose and usually takes longer to engraft. Limitations around this have been addressed by in vivo and ex vivo expansion techniques as well as enhanced engraftment kinetics. Development of adoptive immunotherapy using other components of umbilical cord blood such as regulatory T cells, virus-specific T cells, and natural killer cells has further transformed the field and enhanced the utility of umbilical cord blood unit.

Introduction Pediatric PBSC harvests pose specific challenges during apheresis and a knowledge of the same and variables affecting PBSC collection are very important in planning these procedures. In the present study safety profile of pediatric PBSC procedures and variables influencing the successful collection were analyzed. Method Pediatric PBSC harvest data for 3 years was reviewed for donor, procedural and product parameters and any specific challenges faced during the procedures. Successful PBSC collection was defined when CD34 dose obtained was ≥2 × 106 cells/Kg of recipients’ body weight. Results 85 PBSC collections performed on 46 children (age range 1.5–15 years) were included. Sixty-two procedures were on autologous donors and 23 on allogenic donors. The median CD34+ cell dose in the PBSC product per procedure was 2.12 × 106 cells/Kg for autologous procedures and 4.6 × 106 cells/Kg for allogenic procedures. Systemic adverse reaction was observed during only one procedure (0.01 %) and was managed conservatively. Successful dose was collected in 52 procedures (61.17 %) and was significantly associated with CD34+ count of more than 19.7/μL, monocyte count of more than 1.65 × 106/μL, allogenic collection and female gender (p = 0.00001, p = 0.011, p = 0.00052, and p = 0.0001, respectively). Conclusion PBSC collection is safe in pediatric age groups and pre-procedure CD34 count of ≥20/μL on the day of collection may result in successful collection of stem cell dose. It is important to identify factors associated with failed collection for appropriate counselling and justifying pre-emptive use of stem cell mobilizing agents.

Background and objectives Patient blood (more accurately: haemoglobin, Hb) management (PBM) aims to optimize endogenous Hb production and to minimize iatrogenic Hb loss while maintaining patient safety and optimal effectiveness of medical interventions. PBM was adopted as policy for patients by the World Health Organization (WHO), and, all the more, should be applied to healthy donors. Materials and methods Observational data from 489 bone marrow (BM) donors were retrospectively analysed, and principles of patient blood management were applied to healthy volunteer BM donations. Results and conclusion We managed to render BM aspiration safe for donors, notably completely avoiding the collection of autologous blood units and blood transfusions through iron management, establishment and curation of high-yield aspiration technique, limitation of collection volume to 1·5% of donor body weight and development of volume prediction algorithms for the requested cell dose.

Umbilical cord blood transplantation (UCBT) is a suitable alternative for patients with acute leukemia (AL) in need of an allograft and who lack an HLA-matched donor. Single-institution and registry studies have shown that, in both children and adults with AL, the outcome of UCBT is comparable to that of matched unrelated donor. At the same time, these studies have highlighted some limitations of UCBT, such as increased early mortality and delayed recovery of both hematopoietic and immune compartment, which hamper a more widespread adoption of this approach.In this review, we will analyze the current results of UCBT in children and adults with AL, including comparisons with other hematopoietic stem cell sources and transplant strategies. We will also discuss important factors to be considered when selecting UCB units, as well as future strategies to further improve the outcome of UCBT recipients.The utilization of UCBT for the treatment of AL patients has decreased in recent years. However, recent clinical data suggesting that UCBT might offer better results in patients with minimal residual disease, as well as innovative strategies to facilitate engraftment, reduce transplant-related mortality, and optimize anti-leukemic activity, may pave the way toward a second youth for use of UCB cells.

BACKGROUND The cell dose infused for cord blood transplantation strongly correlates with outcomes following transplantation. Post thaw recoveries can be improved by washing cord blood units (CBUs) in dextran/albumin. Early methods used a labor-intensive manual process. We have recently developed and validated an automated washing method. We now report our results of a study comparing cellular recoveries achieved after manual and automated wash, as well as the impact on engraftment following allogeneic transplantation. STUDY DESIGN AND METHODS CBUs distributed by the Carolinas Cord Blood Bank for clinical use at Duke University after manual or automated wash were included in this report. Precryopreservation total nucleated cell count, total CD34+, colony-forming units, recoveries, and sterility were analyzed by wash method. Patient age, cell dose/weight, diagnosis, conditioning regimen, immunosuppression, and time to neutrophil engraftment were also analyzed. RESULTS Manual and automated washed CBUs yielded similar total nucleated cell count and total CD34+ recoveries. Significantly higher colony-forming units recoveries were achieved after automated washing. Patients who received CBUs washed via an automated method experienced earlier neutrophil engraftment. CONCLUSION While manual and automated washing achieved similar post thaw cellular recoveries, automated washed CBUs demonstrated higher colony-forming unit recovery, which is an important predictor of potency and engraftment. Furthermore, we demonstrated that automated washing was associated with earlier neutrophil engraftment. Our findings favor the use of an automated wash method over a manual approach.

The effect of CD34+ cell dose in allogeneic hematopoietic stem cell transplantation (HSCT) on overall survival (OS) and incidence of acute and chronic graft-versus-host disease (GvHD) has not been established and few studies have been performed. Our single center analysis included 189 patients with hematological malignancies who received peripheral blood stem cell (PBSC) grafts from sibling donors. Myeloablative conditioning was used in 88 cases and 101 received reduced intensity conditioning. The median CD34+ cell dose was 5.6 × 106/kg (0.6–17.0). In the multivariate analysis, a CD34 cell dose of 6–7 × 106/kg was associated with better OS and lower transplant-related mortality (TRM), while a dose of 6.5 × 106/kg) correlated with less acute GVHD (aGVHD) II–IV. We conclude that the CD34 cell dose has an impact on the outcome of HSCT from sibling donor PBSCs.

Purpose Osteoarthritis (OA) represents a relevant social and economic burden worldwide. “Mesenchymal stem cells” or, as recently proposed, “medicinal signaling cells” (MSCs) have been recently introduced as injective treatments for OA with the aim of restoring joint homeostasis. The aim of this review is to provide the reader with the tools necessary to interpret the currently available clinical data, focusing on the MSC mechanisms of action which might help to clarify what we should expect from this treatment. Methods Clinical studies reporting MSC injections for the treatment of knee OA, either freshly isolated or culture-expanded cells, have been included and commented in relation to the supposed therapeutic effect that MSCs might exert giving their supposed mode of actions. Results The majority of the studies reports significant improvements in terms of pain and knee function compared to baseline values, up to 24 months of follow-up. Although these data support the expected therapeutic effect of this therapy giving the features of these cells, only 14% of the studies present a control group and more than one-third of them report the results on less than ten patients. Conclusions Despite the constant presence of positive and satisfactory results in the studies analyzed, the complexity of MSC metabolism and related therapeutic effects as well as the weakness of most of the studies do not allow withdrawing definitive conclusions about the superiority of one tissue source over another, as well as about the best cell dose and the long-term durability of the effects of these procedures. Given the high potential value of these therapies in the treatment of OA, further studies accurately designed, carefully defining the type of patients to be included and pursuing minimal standard requirements in terms of follow-up, number of patients, and types of measurements should be conducted to finally assess the efficacy of MSC-based injective treatments.

Although the CD34+ cell dose in allogeneic peripheral blood stem cell transplantation (PBSCT) is considered to be associated with transplantation outcomes, a lower acceptable threshold has not been defined. We retrospectively analyzed 2919 adult patients with hematologic malignancies who underwent related PBSCT in Japan between 2001 and 2014. According to the number of CD34+ cells in the graft, we categorized 2494 patients in the standard group (2 to 5 × 106 cells/kg), 377 patient in the low group (1 to 2 × 106 cells/kg), and 48 patients in the very low group (

Medicinal signaling cell (MSC)-based products represent emerging treatments in various therapeutic areas including cardiometabolic, inflammation, autoimmunity, orthopedics, wound healing and oncology. Exploring innovation beyond minimally manipulated plastic-adherent ex vivo expanded allogeneic MSCs enables product delineation. Product delineation is on the critical path to maximize clinical benefits and market access. An innovation framework is presented here along various innovation dimensions comprising composition-of-matter by means of positive cell surface markers, formulation varying for example the cell dose or the preservation mode and medium, manufacturing to adapt the secretome of MSCs to the condition of interest, the mode of delivery and corresponding delivery devices, as well as molecular engineering and biomarkers. The rationale of the innovation space thus described applies generally to all cell-based therapies.

Background: Bispecific anti-CD20, anti-CD19 (LV20.19) CAR T-cells may improve outcomes in relapsed, refractory (R/R) non-Hodgkin lymphoma (NHL) by limiting relapse due to single antigen downregulation. We recently reported outcomes of a phase I trial of LV20.19 CAR T-cells expanded in IL-2 in R/R NHL & CLL (Shah Nat Med 2020). Preclinical models demonstrated CAR T-cells expanded in IL-7 & IL-15 (IL7+15) had improved persistence and anti-tumor efficacy compared to IL-2 (Xu Blood 2014). It is unknown how the length of manufacturing modulates the composition and efficacy of the CAR-T product in the clinical setting. To optimize our CAR-T product, we opened a phase I/II trial of LV20.19 CAR T-cells expanded in IL-7+15 manufactured under variable lengths of time (8 vs 12 days) in R/R NHL (NCT04186520). We describe manufacturing and immunophenotypic differences for LV20.19 CAR T-cells expanded in IL7+15 and manufactured for 8- or 12-days versus (vs) cells expanded in IL-2 and manufactured for a fixed 14-days. Methods: LV20.19 CAR T-cells were manufactured onsite with the CliniMACS Prodigy device. Immunophenotyping was conducted for CAR-T products using 8-color flow cytometry. A panel of markers to assess T-cell differentiation status and an immune checkpoint protein panel (LAG3, TIM3, & PD1) were used to analyze IL7+15 expanded CAR T-cells. Testing during the manufacturing process was done to determine: %CD3 T-cells, CD4:CD8 ratios (total T cells & CAR T-cells), #CD3 T-cells, fold CD3 T-cell expansion, %CAR+ T-cells, #CAR T-cells, and % viability at days 0, 7 & 8 or day 12 for IL7+15 products, and days 0, 8 & 14 for IL-2 products. Variables were analyzed with parametric or non-parametric tests where appropriate. Analysis was limited to those treated at 2.5x10 6 cells/kg. A p-value Results: There were 21 patients from the IL7+15 LV20.19 trial and 12 from the LV20.19 IL-2 trial included in the analyses. In the IL7+15 trial, there were 13 patients included in the 8-day manufacturing arm and 8 in the 12-day arm. Manufacturing differences of products expanded in IL7+15 vs IL-2 LV20.19 CAR T-cells expanded in IL7+15 had over 2 times greater fold CD3 expansion by day 7 than IL-2 expanded cells at day 8 (23.4± 7.7 vs 10.4 ± 5.7) (figure 1). Fold CD3 expansion was also higher on day 12 (n=8) for IL7+15 expanded cells at 45.2 ± 11.3 vs 33.9 ± 9.3 on day 14 for IL-2 expanded cells. Although starting numbers of CD3 T-cells were similar between arms, the number of CAR T-cells at day 7 was over 3-fold higher for IL7+15 vs IL-2 expanded cells at day 8 (5.19 ± 2.94e8 vs 1.51 ± 1.48e8)(table 1). Immunophenotypic differences of products expanded in IL7+15 manufactured for 8 vs 12 days LV20.19 CARs harvested at day 8 had a higher frequency of CAR T-cells with a central-memory (CM) phenotype (CD4 cells: 14.0% vs 1.1%, p=0.001; CD8 cells: 21.1% vs 2.1%, p=0.009) (figure 2). The 8-day arm had a lower frequency of CAR T-cells with a terminally differentiated effector-memory CD45RA+ (EMRA) phenotype (CD4 cells: 21.2% vs 3.4%, p=0.06; CD8 cells: 33.0% vs 13.9%, p=0.003). There were no significant differences in the frequency of naïve, T-stem cell memory (Tscm), or effector memory (EM) phenotypes between arms. However, T-cells with a naïve phenotype were virtually absent from the day 12 products but still present in day 8 products. Expression of immune checkpoint proteins was similar between 8- and 12-day arms. Conclusions: Manufacturing LV20.19 CAR T-cells with IL7+15 leads to improved CD3 fold expansion and greater absolute numbers of CAR T-cells vs IL-2. IL7+15-expanded LV20.19 CAR T-cells manufactured for a shorter duration (8 days) have a higher frequency of cells with a CM phenotype and less terminally differentiated cells than those manufactured for 12 days. Central-memory T-cells have been found to have superior in vivo persistence than EM T-cells (Berger, J Clin Invest 2008), and CAR-T products generated from CM T-cells have greater antitumor efficacy than those from EM T-cells (Sommermeyer, Leukemia 2016). Given the composition of CAR-T products has been shown to impact anti-tumor efficacy, this suggests that manufacturing bispecific CAR T-cells in IL7+15 for a shorter duration could lead to greater clinical efficacy without negatively impacting attainment of cell dose. Clinical outcomes of 8- vs 12-day manufactured IL-7+15 expanded LV20.19 CAR T-cells will be reported separately. Figure 1 Figure 1. Disclosures Schneider: Employee of Lentigen Technology, a Miltenyi Biotec Company: Current Employment. Hari: Oncopeptides: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Adaptive Biotech: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding, Speakers Bureau; Millenium: Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding, Speakers Bureau; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding, Speakers Bureau; Karyopharm: Consultancy; GSK: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding, Speakers Bureau; Celgene-BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other, Research Funding, Speakers Bureau; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Johnson: Miltenyi Biotec: Research Funding. Shah: Umoja: Consultancy; Miltenyi Biotec: Consultancy, Honoraria, Research Funding; Kite: Consultancy; Incyte: Consultancy; Epizyme: Consultancy; Legend: Consultancy; Lily: Consultancy, Honoraria, Research Funding.

Autologous hematopoietic stem cell transplantation (ASCT) is an established treatment for multiple myeloma (MM), yet the impact of transplanted CD34+ cell dose remains unresolved, especially in patients over the age of 65 years. Data was collected from 207 consecutive ASCT patients to determine the relationship between CD34+ infusion count and short-term and long-term platelet recovery. For MM patients under the age of 65 years (n=155), CD34+ dosage correlates with time to platelet engraftment (p

Manufacture of red blood cells (RBCs) from progenitors has been proposed as a method to reduce reliance on donors. Such a process would need to be extremely efficient for economic viability given a relatively low value product and high 2E12 cell dose. Therefore, the aim of these studies was to define the productivity of an industry standard stirred-tank bioreactor and determine engineering limitations of commercial RBC production. Cord blood derived CD34+ cells were cultured under erythroid differentiation conditions in a stirred micro-bioreactor (ambr™). Enucleated cells of 80% purity could be created under optimal physical conditions: pH 7.5, 50% oxygen, without gas-sparging (which damaged cells) and with mechanical agitation (which directly increased enucleation). O2 consumption was low (~5x10(-8) µg/cell.hr) theoretically enabling erythroblast densities in excess of 5x10(8) /ml in commercial bioreactors and sub-10 L/unit production volumes. The bioreactor process achieved a 24% and 42% reduction in media volume and culture time respectively relative to unoptimized flask processing. However, media exchange limited productivity to 1 unit of erythroblasts per 500 L of media. Systematic replacement of media constituents, as well as screening for inhibitory levels of ammonia, lactate and key cytokines did not identify a reason for this limitation. We conclude that the properties of erythroblasts are such that the conventional constraints on cell manufacturing efficiency, such as mass transfer and metabolic demand, should not prevent high intensity production; furthermore this could be achieved in industry standard equipment. However, identification and removal of an inhibitory mediator is required to enable these economies to be realized.

Background: Peripheral blood hematopoietic stem cell mobilization for autologous hematopoietic stem cell transplantation (auto-HSCT) in multiple myeloma (MM) can be achieved with either growth factors (GF) alone (filgrastim +/- plerixafor), or with chemotherapy (GF + chemo). When utilized, the chemotherapy regimens include single-agent cyclophosphamide (Cy), or combination regimens, including cyclophosphamide, vincristine or bortezomib, doxorubicin, dexamethasone (CVAD/CBAD) at our center. The optimal mobilization strategy, however, has yet to be established. Methods: In this single center retrospective analysis, we identified 1,006 patients who received auto-HCT for MM between 2009 and 2015. This time-period was chosen to include patients who received auto-HCT after the availability of plerixafor. Patients were divided into 4 groups: G (filgrastim alone), G+P (filgrastim + plerixafor), Cy, and CVAD/CBAD. Plerixafor was mainly used "just-in-time", and not as planned therapy in accordance with our Departmental guidelines. Primary endpoints were CD34+ cell dose/kg collected, days to collect the target CD34+ cell dose, time to neutrophil engraftment (first of three consecutive days of peripheral blood neutrophil count of >500 x 106/L), packed red blood cell (PBRC) and platelet transfusion requirement, duration of hospitalization, progression-free survival (PFS), and overall survival (OS). Results: Patient characteristics are summarized in Table 1. There were 654 patients mobilized with G, 203 with G + P, 80 with Cy, and 69 with CVAD/CBAD. Patients mobilized with CVAD/CBAD were younger compared to the other three groups, were less likely to have achieved VGPR to induction, and more likely to have received a more intense preparative regimen (Table 1). Patients who received G alone, G+P, Cy, and CVAD/CBAD collected a median of 4.1 (0.7-12.2), 4.0 (1.8-11.1), 5.2 (2.2-19.2), and 5.6 (2.5-26.6) x106 CD34+ cells/kg [p Conclusion: Approximately 85% of MM patients underwent PBSC mobilization with GF only (G or G+P). GF + chemo (Cy, CVAD/CBAD) was primarily used in patients with suboptimal response to induction, and allowed successful PBSC collection in this high-risk group. GF + chemo-based mobilization was associated with a higher CD34+ cell dose collection, without improving the time to neutrophil or platelet engraftment, PRBC or platelet transfusion requirement, or the duration of hospitalization. Disclosures Bashir: Purdue: Other: Advisory Board; StemLine: Research Funding; Acrotech: Research Funding; Takeda: Other: Advisory Board, Research Funding; Celgene: Research Funding; Amgen: Other: Advisory Board; KITE: Other: Advisory Board. Nieto:Secura Bio: Other: Grant Support; Astra Zeneca: Other: Grant Support; Novartis: Other: Grant Support; Affimed: Consultancy, Other: Grant Support. Hosing:NKARTA Inc.: Consultancy. Popat:Bayer: Research Funding; Novartis: Research Funding. Lee:Sanofi: Consultancy; GlaxoSmithKline: Consultancy, Research Funding; Genentech: Consultancy; Amgen: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Genentech: Consultancy; Regeneron: Research Funding; Daiichi Sankyo: Research Funding; Takeda: Consultancy, Research Funding; Janssen: Consultancy, Research Funding. Patel:Bristol Myers Squibb: Consultancy, Research Funding; Oncopeptides: Consultancy; Poseida: Research Funding; Janssen: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Cellectis: Research Funding; Precision Biosciences: Research Funding; Takeda: Consultancy, Research Funding; Nektar: Consultancy, Research Funding. Manasanch:Merck: Research Funding; Novartis: Research Funding; Quest Diagnostics: Research Funding; Adaptive Biotechnologies: Honoraria; GSK: Honoraria; Sanofi: Honoraria; BMS: Honoraria; Takeda: Honoraria; JW Pharma: Research Funding; Sanofi: Research Funding. Thomas:BMS: Research Funding; Ascentage: Membership on an entity's Board of Directors or advisory committees, Research Funding; X4 Pharma: Research Funding; Xencor: Research Funding; Pharmacyclics: Other: Advisory Boards; Genentech: Research Funding. Kaufman:Karyopharm: Honoraria; Bristol Myers Squibb: Research Funding; Janssen: Research Funding. Orlowski:Amgen, Inc., AstraZeneca, BMS, Celgene, EcoR1 Capital LLC, Forma Therapeutics, Genzyme, GSK Biologicals, Ionis Pharmaceuticals, Inc., Janssen Biotech, Juno Therapeutics, Kite Pharma, Legend Biotech USA, Molecular Partners, Regeneron Pharmaceuticals, Inc.,: Honoraria, Membership on an entity's Board of Directors or advisory committees; Laboratory research funding from BioTheryX, and clinical research funding from CARsgen Therapeutics, Celgene, Exelixis, Janssen Biotech, Sanofi-Aventis, Takeda Pharmaceuticals North America, Inc.: Research Funding; Founder of Asylia Therapeutics, Inc., with associated patents and an equity interest, though this technology does not bear on the current submission.: Current equity holder in private company, Patents & Royalties; Sanofi-Aventis, Servier, Takeda Pharmaceuticals North America, Inc.: Honoraria, Membership on an entity's Board of Directors or advisory committees; STATinMED Research: Consultancy. Champlin:Actinium: Consultancy; Johnson and Johnson: Consultancy; Omeros: Consultancy; Cytonus: Consultancy; DKMS America: Membership on an entity's Board of Directors or advisory committees; Genzyme: Speakers Bureau; Takeda: Patents & Royalties. Qazilbash:Bioclinica: Consultancy; Amgen: Research Funding; Janssen: Research Funding; Angiocrine: Research Funding; Bioline: Research Funding.

Background & Aim The Barkey Plasmatherm (Barkey GmbH & Co. KG) dry heating device (BPDHD) is designed to thaw frozen products intended for infusion. This study compared cryopreserved apheresis hematopoietic stem/progenitor cell (HS/PC) product CD34 + cell dose recovery, viability and sterility using the BPDHD and a traditional 37°C water bath (Isotemp; Fisher Scientific). Methods, Results & Conclusion METHODS Cryopreserved HS/PC apheresis products were used to assess CD34 recovery, viability and sterility. We selected two products from a single processing and freezing from three independent collections. A paired bag from each of the three collections and freezing procedures was thawed using either the BPDRD or Isotemp water bath. Each thawed product was tested for CD34+ cell dose recovery, CD34+ cell 7AAD viability, sterility by bacterial culture, and colony forming unit (CFU) growth (Stem Cell Technologies). We also performed an independent 3-month post-transplant follow up to compare the time to neutrophil and platelet engraftment in patients who received products thawed using the BPDHD vs products thawed using the Isotemp water bath RESULTS Three products were thawed using BPDHD and 3 products were thawed using a water bath for a total of 6 products thawed and tested. Table 1 summarizes the post-thaw testing performed on each product. All post-thaw testing met our established release criteria (Table 2). We compared the engraftment outcomes for the first 15 patients who received products thawed with the BPDHD to transplant recipients who received products thawed in the Isotemp water bath. We evaluated days to > 500 neutrophils/uL for three consecutive days, and days to platelet count > 20 × 109/L without platelet transfusion support for 7 days. We found no difference between the two thawing methods (Table 3.) CONCLUSION A 37oC water bath is the most commonly used device to thaw cellular therapy products in the United States. This method requires manually submerging the product directly into temperature-controlled water and massaging the bag to ensure a complete thaw, and the water must be changed after each use. The BPDHD is a dry heating system with water-filled cushions and the water is changed annually. Moreover, the water never touches the product, which in principle will reduce the possibility of contamination. The BPDHD takes away the manual manipulation of the product making thawing more standardized, which may translate into improved laboratory efficiency.

Purpose: The aim of this study is to build a computational model to investigate the cell dose and cell DNA damage distribution of a multicellular tissue system under the irradiation.Materials and methods: In this work, we developed a computational model for quantifying cell dose and double strand break (DSB) number in a multicellular system by simulating the radiation transport in 2D and 3D cell culture. The model was based on an open-source radiation transport package, Geant4 with Geant4-DNA physics. First, the computational multicellular system was created using a developed program, CelllMaker. Second, the radiation transport simulation for cells was conducted using Geant4 package with the Geant4-DNA physics to obtain the cellular dose and cellular DSB yield.Results: Using the method described in this work, it is possible to obtain the cellular dose and DNA damage simultaneously. The developed model provides a solution for quantifying the cellular dose and cellular DNA damage which are not easily determined in a radiobiological experiment.Conclusions: With limited validation data for the model, this preliminary study provides a roadmap for building a comprehensive toolkit for simulating cellular dose and DNA damage of multicellular tissue systems.

Despite decades of intensive attention directed to creation of genetically altered cells (e.g., as in development of chimeric antigen receptor (CAR) T-cells) and/or to achieve requisite in vitro accumulation of desired immunologic effectors (e.g., elaboration of virus-specific T cells, expansion of NK cells, differentiation of dendritic cells, isolation, and propagation of Tregs, etc.), there has been essentially no interest in the most fundamental of all hurdles: assuring tissue-specific delivery of administered therapeutic cells to sites where they are needed. With regards to use of CAR T-cells, the absence of information on the efficacy of cell delivery is striking, especially in light of the clear association between administered cell dose and adverse events, and the obvious fact that pertinent cell acquisition/expansion costs would be dramatically curtailed with more efficient delivery of the administered cell bolus. Herein, based on information garnered from studies of human leukocytes and adult stem cells, the logic underlying the use of cell surface glycoengineering to enforce E-selectin ligand expression will be conveyed in the context of how this approach offers strategies to enhance delivery of CAR T-cells to marrow and to tumor beds. This application of glycoscience principles and techniques with intention to optimize cell therapeutics is a prime example of the emerging field of “translational glycobiology.”

The main mechanisms of action resulting in a long-term cure, but also in many life-threatening side effects after HSCT, are mediated by the rapidly reconstituting immune repertoire, which depends on the conditioning regimen, cell dose and graft composition, as well as the type of immune suppression. However, knowledge of these mechanisms is limited, due to many variations in clinical programs, including the specific type of transplantation procedure, as well as a lack of standardized immune monitoring after HSCT.

Haematological engraftment was assessed in 804 autologous transplants. Neutrophil recovery occurred in over 99% within 14 d but platelet recovery was delayed beyond this time in 14·8%. Time to recovery was dependent on the progenitor cell dose infused. The minimum CD34+ cell threshold adopted in this study (2 × 106 /kg) was safe although recovery was faster with a dose >5 × 106 /kg. CD34+ cell doses of between 1 and 2 × 106 /kg were also acceptable if either the granulocyte-macrophage colony-forming cell dose exceeded 2 × 105 /kg or this dose was due to splitting a higher yield harvest. Prompt neutrophil recovery affords important quality assurance for laboratory processing.

Inconsistent results have been reported regarding the influence of graft composition on the incidence of graft versus host disease (GVHD), disease control and survival after reduced-intensity conditioning (RIC) allogeneic peripheral blood stem cell transplantation (allo-PBSCT). These discrepancies may be at least in part explained by the differences in disease categories, disease status at transplant, donor type and conditioning. The current retrospective EBMT registry study aimed to analyze the impact of CD3+ and CD34+ cells dose on the outcome of RIC allo-PBSCT in patients with acute myelogenous leukemia (AML) in first complete remission, allografted from HLA-matched unrelated donors (10 of 10 match). We included 203 adults. In univariate analysis, patients transplanted with the highest CD3+ and CD34+ doses (above the third quartile cut-off point values, >347 x 10^6/kg and >8.25 x 10^6 /kg, respectively) had an increased incidence of grade III-IV acute (a) GVHD (20% vs. 6%, P = .003 and 18% vs. 7%, P = .02, respectively). There was no association between cellular composition of grafts and transplant-related mortality, AML relapse, incidence of chronic GVHD and survival. Neither engraftment itself nor the kinetics of engraftment were affected by the cell dose. In multivariate analysis, CD3+ and CD34+ doses were the only adverse predicting factors for grade III-IV aGVHD (HR = 3.6; 95%CI: 1.45-9.96, P = .006 and 2.65 (1.07-6.57), P = .04, respectively). These results suggest that careful assessing the CD3+ and CD34+ graft content and tailoring the cell dose infused may help in reducing severe acute GVHD risk without negative impact on the other transplantation outcomes. ispartof: Oncotarget vol:7 issue:19 pages:27255-27266 ispartof: location:United States status: published

BACKGROUND Although fat grafting is an increasingly popular practice, suboptimal volume retention remains an obstacle. Graft enrichment with the stromal vascular fraction has gained attention as a method of increasing graft retention. However, few studies have assessed the fate and impact of transplanted stromal vascular fraction on fat grafts. In vivo imaging techniques can be used to help determine the influence stromal vascular fraction has on transplanted fat. METHODS Stromal vascular fraction was labeled with 1,1'-dioctadecyl-3,3,3',3'-tetramethylindotricarbocyanine iodide (DiR), a near-infrared dye, and tracked in vivo. Proliferation and differentiation of labeled cells were assessed to confirm that labeling did not adversely affect cellular function. Different doses of labeled stromal vascular fraction were tracked within fat grafts over time using the in vivo imaging system. RESULTS No significant differences in differentiation and proliferation were observed in labeled versus unlabeled cells (p > 0.05). A pilot study confirmed that stromal vascular fraction fluorescence was localized to fat grafts and different cell doses could be distinguished. A larger-scale in vivo study revealed that stromal vascular fraction fluorescence was statistically significant (p < 0.05) between different cell dose groups and this significance was maintained in higher doses (3 × 10(6) and 2 × 10(6) cells/ml of fat graft) for up to 41 days in vivo. CONCLUSIONS DiR labeling allowed the authors to differentiate between cell doses and confirm localization. This article supports the use of DiR labeling in conjunction with in vivo imaging as a tool for imaging stromal vascular fraction within fat grafts.

Background Adoptive cell therapy (ACT) with T cell receptor (TCR) gene modified peripheral blood T cells creates large pools of tumor reactive T cells. Based on preclinical validation we have selected a high affinity MART-1-specific TCR for TCR gene therapy in metastatic melanoma (MM). Utilizing a novel GMP production protocol we performed a phase I trial to asses feasibility, safety and efficacy of TCR gene therapy in pts with MM. Methods HLA-A2*0201+ pts with irresectable stage IIIc/IV melanoma, expressing MART-1 and MHC class I, with no standard treatment options were included. Autologous T cells were isolated via apheresis and transduced with a MP-71 retroviral vector encoding the 1D3HMCys MART-1 TCR and expanded ex vivo in presence of IL-7 and IL-15. Non-myeloablative chemotherapy was given prior to one i.v. infusion of MART-1 TCR transduced T cells in a dose escalating manner after evaluation of adverse events (AEs). Feasibility, safety (CTCAE 4.0) and ORR (RECIST 1.1) were assessed. Results Twelve heavily pretreated metastatic cutaneous (n = 7) and uveal (n = 5) melanoma (mUM) pts were treated with MART-1 TCR transduced T cells across 4 dose cohorts. Transduction efficiency was 42-76%. Viability of the cell product was 92.9-98.5%. Pt 1 received 4.56 x 109 MART-1 TCR transduced T cells but died 9 days post infusion due to multiple organ failure. Subsequent pts received 5 x 107 (n = 3; cohort (c) 2), 25 x 107 (n = 2; c3) and 10 x 107 (n = 6; c4) cells. On-target AEs were dose-dependent and included dermatitis (10/11) max grade 3, uveitis (3/11) max grade 2 and ototoxicity (4/11) max grade 3, highest in cohort 3. Four pts (n = 2 c3; n = 2 c4) showed signs of cytokine release syndrome and 3 pts required tocilizumab. Objective PR by RECIST 1.1 was seen in 2 pts (17%), with a DOR of 4.2 (mUM, c4) and 8.4 (c3) months. Persistence of transduced T cells in peripheral blood was correlated with infused cell dose. Conclusions Treatment with MART-1 TCR transduced peripheral blood T cells expanded in presence of IL-7 and IL-15 led to severe dose-dependent toxicity with a maximum tolerated dose of 10 x 107 transduced cells. Despite observed responses, toxicity limited further development and use of this MART-1 TCR cellular therapy in MM patients. NCT02654821. Clinical trial identification NCT02654821. Legal entity responsible for the study Netherlands Cancer Institute/Antoni van Leeuwenhoek hospital (NKI-AVL). Funding KWF Kankerbestrijding. Disclosure J.H. Beijnen: Shareholder / Stockholder / Stock options, Full / Part-time employment: Modra Pharmaceuticals BV . J.V. van Thienen: Honoraria (institution), Advisory / Consultancy: Pfizer ; Honoraria (institution), Advisory / Consultancy: Novartis. C.U. Blank: Honoraria (self), Advisory / Consultancy, Research grant / Funding (institution): BMS; Honoraria (self), Advisory / Consultancy: MSD; Honoraria (self), Advisory / Consultancy: Roche; Honoraria (self), Advisory / Consultancy, Research grant / Funding (institution): Novartis; Honoraria (self), Advisory / Consultancy: Lilly; Honoraria (self), Advisory / Consultancy: Pfizer; Honoraria (self), Advisory / Consultancy: GSK; Honoraria (self), Advisory / Consultancy: GenMab; Honoraria (self), Advisory / Consultancy: Pierre Fabre ; Research grant / Funding (institution): NanoString. T.N. Schumacher: Advisory / Consultancy: Adaptive Biotechnologies; Advisory / Consultancy, Shareholder / Stockholder / Stock options: AIMM Therapeutics; Advisory / Consultancy, Shareholder / Stockholder / Stock options: Allogene Therapeutics; Advisory / Consultancy: Amgen; Advisory / Consultancy, Shareholder / Stockholder / Stock options: Merus; Advisory / Consultancy, Shareholder / Stockholder / Stock options: Neon Therapeutics; Advisory / Consultancy: Scenic Biotech; Advisory / Consultancy: Third Rock Ventures; Research grant / Funding (institution): Merck; Research grant / Funding (institution): Bristol-Myers Squibb; Research grant / Funding (institution): Merck KGaA; Shareholder / Stockholder / Stock options: Neogene Therapeutics; Shareholder / Stockholder / Stock options: Scenic Biotech. J.B.A.G. Haanen: Honoraria (institution), Advisory / Consultancy, Research grant / Funding (institution): BMS; Honoraria (institution), Advisory / Consultancy, Research grant / Funding (institution): MSD; Honoraria (institution), Advisory / Consultancy: Pfizer; Honoraria (institution), Advisory / Consultancy: AZ/MedImmune; Honoraria (institution), Advisory / Consultancy: Roche/Genentech; Honoraria (institution), Advisory / Consultancy: Ipsen; Honoraria (institution), Advisory / Consultancy: Bayer; Honoraria (institution), Advisory / Consultancy: Immunocore; Honoraria (institution), Advisory / Consultancy, Research grant / Funding (institution): Novartis; Honoraria (institution), Advisory / Consultancy: Seattle Genetics; Honoraria (institution), Advisory / Consultancy, Research grant / Funding (institution): Neon Therapeutics; Honoraria (institution), Advisory / Consultancy: Celsius Therapeutics; Honoraria (institution), Advisory / Consultancy: Gadet ; Honoraria (institution), Advisory / Consultancy: GSK. All other authors have declared no conflicts of interest.

Introduction Optimal CB unit selection incorporates TNC & CD34+ cell dose & 8-allele donor-recipient HLA-match. How graft characteristics for these parameters vary by patient (pt) race/ ethnicity, however, is unknown. Methods We analyzed cryopreserved TNC & CD34+ cell content, the cell dose incorporating pt weight, & HLA-match of infused & backup (reserved but not shipped) units by pt ancestry in CB transplant (CBT) recipients transplanted 7/2013-6/2018. Units were chosen based on banking practices, TNC & CD34+ dose & 4-6/6 & 8-allele HLA-match. Patient (pt) racial/ ethnic origins were prospectively obtained & grouped per Barker J. et al, BBMT 2010. Results The characteristics of 544 units chosen for 144 diverse CBT recipients by pt ancestry are shown (Table 1). 286 units were infused (142 doubles, 2 singles) & pts had a total of 258 backup units (0-2/pt). The median TNC content of units for Europeans was higher than for Non-Europeans (216 vs 197, p = 0.002). Units for NW Europeans had the highest median TNC content with lowest in units for Africans & White Hispanics. Units chosen for Europeans also had a higher median CD34+ cell content than Non-Europeans (160 vs 132, p = 0.007). Units for NW Europeans had the highest & units for Africans the lowest median CD34+ content. Median TNC/kg was similar in units for European & Non-European pts (2.7 vs 2.5, p = 0.11) & differences between groups were not marked. However, there was a trend for units for Europeans having a higher median CD34+ dose than units for Non-Europeans (1.9 vs 1.7, p = 0.057) with disparity between groups. Although the median 8 allele HLA-match was 5/8 for units chosen for both European & Non-European pts, units identified for non-Europeans were more mismatched (p = 0.017). When the 286 transplanted units (considered the best) were analyzed, the median TNC & CD34+ contents of units were higher for Europeans than non-Europeans (232 vs 216, p = 0.022, & 181 vs 152, p = 0.009). As shown in Table 2, while there were no overall differences in the median TNC & CD34+ doses of units for Europeans & non-Europeans, differences in cell content combined with differences in weights resulted in TNC & CD34+ dose disparities within ancestry groups. For example, NW Europeans (high weight, high CD34+ content) had the highest CD34+ doses; lower content in Asian pts was partially compensated for by low weight; African pts (high weight, low content) had the lowest doses. Finally, there was marked differences in HLA-match distribution by ancestry group: eg only 23% of units for NW Europeans were 3-4/8 vs 40% for S. Europeans, 46% for white Hispanics & 51% for Africans. Conclusions CB extends transplant access to minorities but differences in cellular content translates to many ancestry groups pts having access to only lower dosed units. There are also marked racial/ ethnic disparities in HLA-match. This data supports continued funding of banks to further increase the CB inventory.

During more recent years only few studies have analyzed the effect of total nucleated cell (TNC) and CD34(+) cell dose in allogeneic hematopoietic stem cell transplantation (HSCT). A single-center analysis included 544 patients, 227 with a sibling donor and 317 with an unrelated donor. Most patients (n = 292) were treated with myeloablative conditioning, whereas the remaining patients (n = 252) received reduced-intensity conditioning. Bone marrow (BM) (n = 121) and peripheral blood stem cell (PBSC) grafts (n = 423) were analyzed separately. Median TNC and CD34(+) cell dose was 3.2 × 10(8)/kg versus 11.6 × 10(8)/kg in BM and 3.9 × 10(6)/kg versus 8.1 × 10(6)/kg in PBSC. In the BM group we found a higher TNC and CD34(+) cell dose was associated with a faster neutrophil engraftment (P.001 and P = .02). In the PBSC group we found patients given a very high (≥11 × 10(6)/kg) CD34(+) cell dose had decreased rates of survival (P = .001) and increased relapse (P = .02). A high CD34(+) cell dose correlated with faster platelet engraftment (P.01). In HSCT using PBSCs, the CD34(+) cell doses should be kept below 11 × 10(6)/kg but over 2.5 × 10(6)/kg.

Double umbilical cord blood transplantation (UCBT) was developed as a strategy to circumvent the cell dose limitation of single UCBT with a concomitant potential benefit of lowering the rate of leukemia relapse. Sustained hematopoiesis after double UCBT usually is derived from a single donor unit, as only a few patients have been reported to display stable mixed-unit chimerism for varying periods of time. Explanations for the 1 unit dominance, predictors for identifying unit superiority, and persistence of long-term mixed-unit chimerism remain elusive. Review of published literature revealed only 11 of 280 patients (4%) with mixed-unit chimerism for at least 1 year after transplantation, with 3 patients receiving reduced-intensity conditioning regimens. Mixed-unit chimerism was more likely if both units were closely HLA matched to each other. Outcome data for patients with stable mixed-unit chimerism, for the most part, were scarcely reported. Analysis of the small sample size revealed a potential advantage of stable mixed-unit chimerism on enhancing the graft-versus-leukemia effect; however, definitive conclusions cannot be made on the effect of mixed-unit chimerism on the rates of graft-versus-host disease. Therefore, gathering outcome data prospectively in larger clinical series will help answer the question of whether stable mixed-unit chimerism is either beneficial and, therefore, should be strived for, detrimental and, thus, needs to be eliminated, or if it is of no clinical consequence.

It has been shown that it is possible to predict the CD 34+ hematopoietic progenitor cell dose from collection procedures on TerumoBCT COBE Spectra® cell separator platform using simple variables available at the start of the procedure. In this article, we demonstrate that this can be done simply and reliably using TerumoBCT Spectra Optia® ("Optia") cell separator platform with a very close correlation between predicted and actual results (correlation coefficient 0.956). This knowledge can be used to optimize apheresis sessions and to minimize harmful effects and costs. In addition, we have shown differences in collection efficiency between healthy donors and cancer patients undergoing autologous donation. Finally, we have shown a small but significant improvement in collection efficiency for the Optia platform compared with the COBE Spectra platform.

Peripheral blood stem cells have largely replaced harvested bone marrow stem cells both in the autologous and allogeneic settings. Advantages of peripherally harvested cells include higher stem cell dose, more rapid engraftment, reduced donor/patient discomfort, and better graft-versus-leukemia effect in the allogeneic setting. Within the apheresis machine, whole blood is separated into its components by centrifugation, and the red cell-depleted, stem cell-rich buffy coat is extracted for use as a stem cell product, simultaneously returning the other blood components back to the donor. Prediction of procedure length is based on the required cell dose target but still remains challenging. Moreover, the number of apheresis procedures needed should be as few as possible in order to reduce costs and patient/donor discomfort and to increase safety. The volume of blood which needs to be processed in order to collect an adequate number of stem cells depends on several factors such as method of stem cell mobilization, vascular access, and collection efficiency.

We retrospectively analyzed a total of 188 autologous stem cell transplantations (ASCT) performed in patients with multiple myeloma (MM), at Grenoble Hospital, between January 2010 and April 2017, in order to evaluate clinical activity and identify factors affecting hematopoietic recovery. In the univariate analysis, age, gender, MM type, disease status, CD34+ cell doses before or after cryopreservation, transfusion requirement were not statistically significant, on time to neutrophil engraftment. Only 2 factors, patients not included in clinical assays (p=0,009) and mobilization modalities with G-CSF alone (p=0,034) are statistically significant and prognostic factors for early engraftment. Our analyses confirm previous findings that patients may be mobilized with G-CSF alone, so we propose this mobilization regimen for patients not included in clinical assay. We also demonstrate that a CD34+ cell dose > 5 x 106 before cryopreservation, is required to reduce transfusion requirement (p=0,006). Our study showed a linear correlation between hematopoietic recovery and CD34+ cell dose infused after cryopreservation but more studies are required to confirm this point.; Nous avons réalisé une étude rétrospective de revue de sortie d’aplasie, constituée de 188 patients atteints de myélome multiple (MM) autogreffés au CHU de GRENOBLE, depuis 7 ans, afin de permettre une évaluation de l’activité, d’identifier les facteurs pouvant influencer cette sortie d’aplasie et améliorer les pratiques cliniques. En analyse univariée, l’âge à la greffe, le sexe, le type de MM, le statut de la maladie, la richesse du greffon en CD34+/kg avant et après congélation, le nombre de produits sanguins transfusés et le statut transfusionnel du patient avant l’autogreffe n’ont pas d’impact. Deux variables, les patients non inclus dans un protocole de recherche clinique (p=0,009) et la mobilisation en état hématologique stable (p=0,034) sont significatives et sont des facteurs de bon pronostic pour une sortie précoce d’aplasie. Le point commun est l’utilisation protocolaire quasi exclusive de l’Endoxan® de mobilisation. Au vu de nos résultats et des données de la littérature, pour les patients traités ‘‘hors protocole’’, nous proposons de réaliser le recueil en état hématologique stable avec mobilisation par G-CSF seul. Nous avons aussi montré que la richesse du greffon avant congélation > 5 x 106 cellules CD34+/kg présente un avantage significatif en terme de besoin transfusionnel (p=0,006). Notre étude fait apparaitre un lien entre la qualité de reconstitution hématopoïétique et le nombre de cellules CD34+ injectées après décongélation, mais ce point mérite des études complémentaires. Le plan d’analyse proposé permet un suivi multiparamétrique du mois suivant l’intensification, pouvant contribuer à une meilleure maîtrise de l’activité de greffe autologue.