Multi-component cord blood banking: a proof-of-concept international exercise.

BACKGROUND: Most public cord blood (CB) banks currently discard more than 80% of umbilical CB units not suitable for hemopoietic stem cell transplant due to low stem cell count. Although CB platelets, plasma, and red blood cells have been used for experimental allogeneic applications in wound healing, corneal ulcer treatment, and neonatal transfusion, no standard procedures for their preparation have been defined internationally. MATERIALS AND METHODS: A network of 12 public CB banks in Spain, Italy, Greece, the UK, and Singapore developed a protocol to validate a procedure for the routine production of CB platelet concentrate (CB-PC), CB platelet-poor plasma (CB-PPP), and CB leukoreduced red blood cells (CB-LR-RBC) using locally available equipment and the commercial BioNest ABC and EF medical devices. CB units with >50 mL volume (excluding anticoagulant) and ≥150×109/L platelets were double centrifuged to obtain CB-PC, CB-PPP, and CB-RBC. The CB-RBC were diluted with saline-adenine-glucose-mannitol (SAGM), leukoreduced by filtration, stored at 2-6°C, and tested for hemolysis and potassium (K+) release over 15 days, with gamma irradiation performed on day 14. A set of acceptance criteria was pre-defined. This was for CB-PC: volume ≥5 mL and platelet count 800-1,200×109/L; for CB-PPP: platelet count <50×109/L; and for CB-LR-RBC: volume ≥20 mL, hematocrit 55-65%, residual leukocytes <0.2×106/unit, and hemolysis ≤0.8%. RESULTS: Eight CB banks completed the validation exercise. Compliance with acceptance criteria was 99% for minimum volume and 86.1% for platelet count in CB-PC, and 90% for platelet count in CB-PPP. Compliance in CB-LR-RBC was 85.7% for minimum volume, 98.9% for residual leukocytes, and 90% for hematocrit. Compliance for hemolysis ≤0.8% decreased from 89.0 to 63.2% from day 0 to 15. K+ release increased from 3.0±1.8 to 25.0±7.0 mmol/L from day 0 to 15, respectively. DISCUSSION: The MultiCord12 protocol was a useful tool to develop preliminary standardization of CB-PC, CB-PPP, and CB-LR-RBC.

Cord blood and amniotic membrane extract eye drop preparations display immune-suppressive and regenerative properties.

Diseases and injuries that compromise the ocular surface cause considerable patient distress and have long term consequences for their quality of life. Treatment modalities that can address the delicate balance of tissue regeneration, inflammation and maintenance of corneal transparency are therefore needed. We have recently formulated two novel eye drops from placental tissues: cord blood platelet lysate (CBED) and amniotic membrane extract eye drops (AMED), which can be used to treat severe ocular disorders. Here we characterise these two preparations by measuring: (a) growth factors (GF) and cytokines composition, (b) promotion of human corneal epithelial cell (HCEC) growth and (c) effects on immune cells in a lymphocyte culture assay. Finally, their bioavailability was assayed in an ex vivo porcine corneal model. We show that both preparations contain GF and cytokines that were able to promote the in vitro growth of HCEC and support repair in an in vitro scratch test. When assessed in a lymphocyte culture, both favoured immune suppression reducing the cellular expression of NKG2D and CD107a as well as the production of interferon gamma (IFN-γ) in natural killer, NKT and T cells. Regarding bioavailability, CBED active molecules were found mainly in the pre-corneal fraction with some penetration into the corneal fraction, in an ex vivo model. In summary, both placental-derived allogeneic preparations, CBED and AMED, display regenerative and immunomodulatory capabilities. These results will help define mechanisms of action and the best indications and doses of each product for use in a particular patient and support the development of off-the-shelf therapies for ocular surface pathologies in which wound healing defects and inflammatory events are contributing factors.

Feasibility of umbilical cord blood as a source of red blood cell transfusion in preterm infants.

BACKGROUND: Preterm infants born earlier than 32 weeks of gestational age (GA) often need red blood cell (RBC) transfusions, which have been associated with an increased incidence of complications of prematurity, due to changes in tissue oxygenation. Transfusion of umbilical cord blood (UCB) could be beneficial for this group. The aims of this study were: (i) to determine the RBC transfusion needs in infants <32 weeks in Hospital Clinic of Barcelona; (ii) to identify the target GA group that would benefit most from UCB transfusion; and (iii) to assess the current availability of UCB as a potential source of RBC transfusion for these premature infants in our tertiary referral blood bank. MATERIAL AND METHODS: A retrospective observational study was performed on infants born at <32 weeks GA, divided into two groups: (i) extremely low gestational age neonates (ELGAN) (from 230 to 276 weeks) and (ii) very preterm neonates (VPN) (from 280 to 316 weeks). Their complications and transfusion rates were compared. Processing and availability of UCB samples in the reference blood bank were assessed. RESULTS: Overall, 1,651 infants <32 weeks GA were admitted in the study period. While 12.5% of VPN received at least one RBC transfusion, the percentage increased to 60% among the ELGAN. Retinopathy of prematurity and bronchopulmonary dysplasia were diagnosed more frequently in the ELGAN group (p<0.001) than in the VPN group. The annual average volume of RBC transfusion in our study group was 1.35 L (95% CI: 1.07-1.64). The reference blood bank was able to produce 16 L (95% CI: 14-18) of UCB-RBC per year. CONCLUSION: Considering the data obtained about RBC transfusion needs and morbidities, the ELGAN group has been identified as the target group that would benefit most from UCB-RBC transfusions. We have demonstrated that our blood bank is able to produce enough RBC from UCB. Randomised control trials are warranted to study the potential benefits of UCB compared to adult blood for RBC transfusions.

Clinical evaluation of allogeneic eye drops from cord blood platelet lysate.

BACKGROUND: Current treatments for several corneal lesions show limited efficacy. Here we report the clinical evaluation of the efficacy of a novel eye drop preparation produced in a public cord blood (CB) bank. MATERIALS AND METHODS: In a multicentre, retrospective, consecutive case study we evaluated 33 patients (46 eyes) unresponsive to conventional treatments who required urgent intervention. The patients were given allogeneic eye drops obtained from cord blood platelet lysate (CBED) to treat severe ocular surface lesions under a compassionate use protocol. The CBED were prepared from CB units donated for haematopoietic stem cell transplantation that did not contain the minimum stem cell dose required for this use. Patients were grouped by acute conditions (neurotrophic ulcers: group I; other corneal ulcers: group II; corneal burns: group III), and chronic conditions (ocular graft-versus-host disease: group IV; severe dry eye syndrome: group V). The patients received one or two drops of the product to the affected eye four to six times per day for 19 days. A further 19-day cycle of treatment could be repeated according to the initial clinical response. RESULTS: Patients received a median of 19 CBED vials (interquartile range 19-57, range 19-442) to complete the therapy. Group I-II-III patients showed full and partial ulcer recovery in 25 (78%) and six (19%) eyes respectively. One eye (3%) did not respond to treatment. For groups IV-V improvement was reported for 12 (85%) eyes and lesions worsened on treatment in both eyes (15%) of one patient. No severe adverse events were directly attributed to CBED. DISCUSSION: Promptly available CBED resulted in a well-tolerated allogeneic treatment that showed evidence of efficacy in this cohort of patients. These positive results support further studies on CBED from platelet lysate as a novel product of CB banks. A prospective clinical trial in neurotrophic keratitis (NCT03084861) is ongoing to confirm these preliminary data.

Cord Blood Platelet Rich Plasma Derivatives for Clinical Applications in Non-transfusion Medicine.

Cord blood platelet rich plasma (CB-PRP) derivatives have been investigated as potential therapeutic agents for the treatment of diverse conditions including ocular surface disease and skin ulcers. We have developed processes for the formulation of several CB-PRP preparations, which have different composition and attributes. Here we describe the molecular characteristics of these preparations and we make recommendations as to their most appropriate clinical application based on functional and immunomodulatory profiles. We show that incubation of adult peripheral blood mononuclear cells (PBMCs) with all three preparations dramatically reduced the production of INFγ and the expression of NKG2D and CD107a in NK, NKT, and T cells thus diminishing their activation, we propose that the likely mechanism is the high levels of soluble NKG2D ligands present in plasma. Of the three preparations we investigated, CB platelet lysate (PL) and platelet releaseate (PR) have higher concentrations of trophic and pro-angiogenic factors, CB platelet poor plasma (PPP) has the lowest concentration of all analytes measured. Based on these finding we propose that CB-PR is the most suitable raw material for skin wound patches, while CB-PL and PPP can be used to prepare eye drops for severe ocular surface pathologies and inflammatory conditions such as corneal ulcers or severe dry eye disease, respectively.

Cord blood-derived platelet concentrates as starting material for new therapeutic blood components prepared in a public cord blood bank: from product development to clinical application.

BACKGROUND: There are many advantages to using cord blood (CB) as a source of therapeutic platelet and plasma derivatives for regenerative medicine. These include availability, universal use, young donor source, and virally safe biological material, rich in tissue regenerative factors. MATERIALS AND METHODS: We aimed to validate a bioprocess design for the production of cord blood-derived platelet concentrates (CBPC) in a public Cord Blood Bank (CBB). CBPC was defined as a product of 10±5 mL, 1,000±200×109/L total platelets, free of erythrocytes and leukocytes. A total of 300 CB units were centrifuged in two steps to enrich for platelets, in compliance with Good Manufacturing Practice. The samples were tested for the degree of platelet activation present, and the levels of growth factor were analysed to evaluate their potential function. CBPC were then activated after thawing with 10% calcium gluconate to generate platelet gels (CBPG) to treat patients with diabetic foot ulcers. RESULTS: After processing, 84% of the products fulfilled the acceptance criteria. Final products contained 1,017±149×106 platelets/mL in 10±3mL of plasma. Platelet recovery was 50±9%. The methods described here ensure depletion of white and red blood cells down to a residual concentration of 0.2±0.1×106/mL and 0.03±0.02×106/mL, respectively. Platelets showed low levels of activation during processing, but were significantly activated after thawing, as indicated by an increase in CD62p expression. The growth factors EGF, VEGF, bFGF, PDGF AB/BB and TGF-ß1 were at concentrations of 1,706±123 pg/mL; 1,602±227 pg/mL; 314±26 pg/mL; 30±1.5 ng/mL; 24±2 ng/mL (mean±standard error of mean), respectively. For clinical evaluation, a total of 21 CBPG were applied in 3 patients, with no reported adverse events and improvement of ulcers in all of them. DISCUSSION: We designed and validated a highly reproducible, closed system method to manufacture high quality CBPC suitable for clinical applications using CB units not suitable for transplantation in a public CBB.

Rapid review: next generation of cord blood banks; transplantation and beyond.

Cord blood (CB) is a medicinal product of human origin with unique cellular properties such as the presence of multipotent stem cells, naive immune cells, and fetal blood components. CB transplantation provides high rate of donor chimerism, and a good balance of graft-versus-host (GVH) and graft-versus-leukemia (GVL) effects. Use of CB for transplantation has decreased in recent years as haplo-identical stem cell transplants have achieved similar short-term clinical outcomes. For most patients, however, the optimal stem cell source remains unclear. CB inventories can be used as a starting material to develop new cellular medicines, and units with low cellular content can be converted to produce blood components like platelet-rich plasma and red blood cell (RBC) units for special indications.