Platelets for advanced drug delivery in cancer.

INTRODUCTION: Cancer-related drug expenses are rising with the increasing cancer incidence and cost may represent a severe challenge for drug access for patients with cancer. Consequently, strategies for increasing therapeutic efficacy of already available drugs may be essential for the future health-care system. AREAS COVERED: In this review, we have investigated the potential for the use of platelets as drug-delivery systems. We searched PubMed and Google Scholar to identify relevant papers written in English and published up to January 2023. Papers were included at the authors' discretion to reflect an overview of state of the art. EXPERT OPINION: It is known that cancer cells interact with platelets to gain functional advantages including immune evasion and metastasis development. This platelet-cancer interaction has been the inspiration for numerous platelet-based drug delivery systems using either drug-loaded or drug-bound platelets, or platelet membrane-containing hybrid vesicles combining platelet membranes with synthetic nanocarriers. Compared to treatment with free drug or synthetic drug vectors, these strategies may improve pharmacokinetics and selective cancer cell targeting. There are multiple studies showing improved therapeutic efficacy using animal models, however, no platelet-based drug delivery systems have been tested in humans, meaning the clinical relevance of this technology remains uncertain.

Freeze-dried platelets are a promising alternative in bleeding thrombocytopenic patients with hematological malignancies.

Thrombosomes are trehalose-stabilized, freeze-dried group O platelets with a 3-year shelf life. They can be stockpiled, rapidly reconstituted, and infused regardless of the recipient's blood type. Thrombosomes thus represent a potential alternative platelet transfusion strategy. The present study assessed the safety and potential early signals of efficacy of Thrombosomes in bleeding thrombocytopenic patients. We performed an open-label, phase 1 study of single doses of allogeneic Thrombosomes at three dose levels in three cohorts, each consisting of eight patients who had hematologic malignancies, thrombocytopenia, and bleeding. Adverse events, dose-limiting toxicities (DLTs), World Health Organization (WHO) bleeding scores, and hematology values were assessed. No DLTs were reported. The median age was 59 years (24-71). Most patients had AML (58%) or ALL (29%), followed by MDS (8%) and myeloproliferative neoplasm (4%). The WHO scores of 22 patients who were actively bleeding at a total of 27 sites at baseline either improved (n = 17 [63%]) or stabilized (n = 10 [37%]) through day 6. Twenty-four hours after infusion, 12 patients (50%) had a clinically significant platelet count increase. Of eight patients who received no platelet transfusions for 6 days after Thrombosomes infusion, 5 had a clinically significant increase in platelet count of ≥5000 platelets/µL and 2 had platelet count normalization. Thrombosomes doses up to 3.78 × 108 particles/kg demonstrated safety in 24 bleeding, thrombocytopenic patients with hematological malignancies. Thrombosomes may represent an alternative to conventional platelets to treat bleeding. A phase 2 clinical trial in a similar patient population is underway.

Platelet Microparticles Decrease Daunorubicin-Induced DNA Damage and Modulate Intrinsic Apoptosis in THP-1 Cells.

Platelets can modulate cancer through budding of platelet microparticles (PMPs) that can transfer a plethora of bioactive molecules to cancer cells upon internalization. In acute myelogenous leukemia (AML) this can induce chemoresistance, partially through a decrease in cell activity. Here we investigated if the internalization of PMPs protected the monocytic AML cell line, THP-1, from apoptosis by decreasing the initial cellular damage inflicted by treatment with daunorubicin, or via direct modulation of the apoptotic response. We examined whether PMPs could protect against apoptosis after treatment with a selection of inducers, primarily associated with either the intrinsic or the extrinsic apoptotic pathway, and protection was restricted to the agents targeting intrinsic apoptosis. Furthermore, levels of daunorubicin-induced DNA damage, assessed by measuring gH2AX, were reduced in both 2N and 4N cells after PMP co-incubation. Measuring different BCL2-family proteins before and after treatment with daunorubicin revealed that PMPs downregulated the pro-apoptotic PUMA protein. Thus, our findings indicated that PMPs may protect AML cells against apoptosis by reducing DNA damage both dependent and independent of cell cycle phase, and via direct modulation of the intrinsic apoptotic pathway by downregulating PUMA. These findings further support the clinical relevance of platelets and PMPs in AML.

Effect of leukoreduction and temperature on risk of bacterial growth in CPDA-1 whole blood: A study of Escherichia coli.

BACKGROUND: Collection of non-leukoreduced citrate-phosphate-dextrose-adenine (CPDA-1) whole blood is performed in walking blood banks. Blood collected under field conditions may have increased risk of bacterial contamination. This study was conducted to examine the effects of WBC reduction and storage temperature on growth of Escherichia coli (ATCC® 25922™) in CPDA-1 whole blood. METHODS: CPDA-1 whole blood of 450 ml from 10 group O donors was inoculated with E. coli. Two hours after inoculation, the test bags were leukoreduced with a platelet-sparing filter. The control bags remained unfiltered. Each whole blood bag was then split into three smaller bags for further storage at 2-6°C, 20-24°C, or 33-37°C. Bacterial growth was quantified immediately, 2 and 3 h after inoculation, on days 1, 3, 7, and 14 for all storage temperatures, and on days 21 and 35 for storage at 2-6°C. RESULTS: Whole blood was inoculated with a median of 19.5 (range 12.0-32.0) colony-forming units per ml (CFU/ml) E. coli. After leukoreduction, a median of 3.3 CFU/ml (range 0.0-33.3) E. coli remained. In the control arm, the WBCs phagocytized E. coli within 24 h at 20-24°C and 33-37°C in 9 of 10 bags. During storage at 2-6°C, a slow self-sterilization occurred over time with and without leukoreduction. CONCLUSIONS: Storage at 20-24°C and 33-37°C for up to 24 h before leukoreduction reduces the risk of E. coli-contamination in CPDA-1 whole blood. Subsequent storage at 2-6°C will further reduce the growth of E. coli.

Platelet Microparticles Protect Acute Myelogenous Leukemia Cells against Daunorubicin-Induced Apoptosis.

The role of platelets in cancer development and progression is increasingly evident, and several platelet-cancer interactions have been discovered, including the uptake of platelet microparticles (PMPs) by cancer cells. PMPs inherit a myriad of proteins and small RNAs from the parental platelets, which in turn can be transferred to cancer cells following internalization. However, the exact effect this may have in acute myelogenous leukemia (AML) is unknown. In this study, we sought to investigate whether PMPs could transfer their contents to the THP-1 cell line and if this could change the biological behavior of the recipient cells. Using acridine orange stained PMPs, we demonstrated that PMPs were internalized by THP-1 cells, which resulted in increased levels of miR-125a, miR-125b, and miR-199. In addition, co-incubation with PMPs protected THP-1 and primary AML cells against daunorubicin-induced cell death. We also showed that PMPs impaired cell growth, partially inhibited cell cycle progression, decreased mitochondrial membrane potential, and induced differentiation toward macrophages in THP-1 cells. Our results suggest that this altering of cell phenotype, in combination with decrease in cell activity may offer resistance to daunorubicin-induced apoptosis, as serum starvation also yielded a lower frequency of dead and apoptotic cells when treated with daunorubicin.

Spontaneous Splenic Artery Rupture as the First Symptom of Systemic Amyloidosis.

Spontaneous splenic rupture is a life-threatening condition leading to a rapidly progressing hypovolemic shock due to intra-abdominal blood loss, with a mortality rate of about 10%. Spontaneous splenic rupture can be caused by widely different disorders including acute and chronic infections, neoplastic disorders, and inflammatory noninfectious disorders. In this case report, we present a 67-year-old male patient with hemorrhagic shock caused by an acute bleeding from the splenic artery. The patient was massively transfused with blood products and fluids and underwent laparotomy for hemostatic control and clinical stabilization. Multiorgan involvement by amyloid light-chain amyloidosis (AL-amyloidosis) caused by plasma cell dyscrasia, specifically with infiltration of the spleen artery, was found to be the underlying cause of his life-threatening bleeding. Based on this case, we discuss the features of serious spleen bleeding, massive transfusion therapy in the intensive care setting, and AL-amyloidosis pathophysiology and treatment.

Influence of platelet storage time on human platelet lysates and platelet lysate-expanded mesenchymal stromal cells for bone tissue engineering.

BACKGROUND: Human platelet lysate (HPL) is emerging as the preferred xeno-free supplement for the expansion of mesenchymal stromal cells (MSCs) for bone tissue engineering (BTE) applications. Due to a growing demand, the need for standardization and scaling-up of HPL has been highlighted. However, the optimal storage time of the source material, i.e., outdated platelet concentrates (PCs), remains to be determined. The present study aimed to determine the optimal storage time of PCs in terms of the cytokine content and biological efficacy of HPL. METHODS: Donor-matched bone marrow (BMSCs) and adipose-derived MSCs (ASCs) expanded in HPL or fetal bovine serum (FBS) were characterized based on in vitro proliferation, immunophenotype, and multi-lineage differentiation. Osteogenic differentiation was assessed at early (gene expression), intermediate [alkaline phosphatase (ALP) activity], and terminal stages (mineralization). Using a multiplex immunoassay, the cytokine contents of HPLs produced from PCs stored for 1-9 months were screened and a preliminary threshold of 4 months was identified. Next, HPLs were produced from PCs stored for controlled durations of 0, 1, 2, 3, and 4 months, and their efficacy was compared in terms of cytokine content and BMSCs' proliferation and osteogenic differentiation. RESULTS: BMSCs and ASCs in both HPL and FBS demonstrated a characteristic immunophenotype and multi-lineage differentiation; osteogenic differentiation of BMSCs and ASCs was significantly enhanced in HPL vs. FBS. Multiplex network analysis of HPL revealed several interacting growth factors, chemokines, and inflammatory cytokines. Notably, stem cell growth factor (SCGF) was detected in high concentrations. A majority of cytokines were elevated in HPLs produced from PCs stored for ≤ 4 months vs. > 4 months. However, no further differences in PC storage times between 0 and 4 months were identified in terms of HPLs' cytokine content or their effects on the proliferation, ALP activity, and mineralization of BMSCs from multiple donors. CONCLUSIONS: MSCs expanded in HPL demonstrate enhanced osteogenic differentiation, albeit with considerable donor variation. HPLs produced from outdated PCs stored for up to 4 months efficiently supported the proliferation and osteogenic differentiation of MSCs. These findings may facilitate the standardization and scaling-up of HPL from outdated PCs for BTE applications.

A Pilot Trial of Platelets Stored Cold versus at Room Temperature for Complex Cardiothoracic Surgery.

BACKGROUND: This pilot trial focused on feasibility and safety to provide preliminary data to evaluate the hemostatic potential of cold-stored platelets (2° to 6°C) compared with standard room temperature-stored platelets (20° to 24°C) in adult patients undergoing complex cardiothoracic surgery. This study aimed to assess feasibility and to provide information for future pivotal trials. METHODS: A single center two-stage exploratory pilot study was performed on adult patients undergoing elective or semiurgent complex cardiothoracic surgery. In stage I, a two-armed randomized trial, platelets stored up to 7 days in the cold were compared with those stored at room temperature. In the subsequent single-arm stage II, cold storage time was extended to 8 to 14 days. The primary outcome was clinical effect measured by chest drain output. Secondary outcomes were platelet function measured by multiple electrode impedance aggregometry, total blood usage, immediate and long-term (28 days) adverse events, length of stay in intensive care, and mortality. RESULTS: In stage I, the median chest drain output was 720 ml (quartiles 485 to 1,170, n = 25) in patients transfused with room temperature-stored platelets and 645 ml (quartiles 460 to 800, n = 25) in patients transfused with cold-stored platelets. No significant difference was observed. The difference in medians between the room temperature- and cold-stored up to 7 days arm was 75 ml (95% CI, -220, 425). In stage II, the median chest drain output was 690 ml (500 to 1,880, n = 15). The difference in medians between the room temperature arm and the nonconcurrent cold-stored 8 to 14 days arm was 30 ml (95% CI, -1,040, 355). Platelet aggregation in vitro increased after transfusion in both the room temperature- and cold-stored platelet study arms. Total blood usage, number of adverse events, length of stay in intensive care, and mortality were comparable among patients receiving cold-stored and room temperature-stored platelets. CONCLUSIONS: This pilot trial supports the feasibility of platelets stored cold for up to 14 days and provides critical guidance for future pivotal trials in high-risk cardiothoracic bleeding patients.

Multiple electrode aggregometry and thromboelastography in thrombocytopenic patients with haematological malignancies.

BACKGROUND: In thrombocytopenic patients better assessment of bleeding risk than that provided by platelet count alone is required. Multiplate® aggregometry and thromboelastography (TEG) could be used, but information on their role in such patients is limited. The primary aim of this study was to investigate the feasibility of Multiplate® analyses in patients with haematological malignancies. A secondary aim was to explore whether a multiple logistic regression model combining Multiplate®, TEG, clinical and laboratory variables was associated with risk of bleeding. MATERIALS AND METHODS: This was an exploratory, prospective observational study of thrombocytopenic patients with haematological malignancies. Total platelet count (TPC), white blood cell count, C-reactive protein (CRP) level, temperature and bleeding status were recorded daily. TEG and Multiplate® analyses with four agonists were performed on weekdays. RESULTS: Ten patients were enrolled into the study. The median number of days in a study period was 21. Bleeding was observed on 64 of 298 study days. TPC <20×109/L and <10×109/L occurred on 119 and 25 days, respectively. When TPC was <33×109/L, many samples showed no aggregation, regardless of bleeding status. Despite this, the odds of World Health Organization (WHO) grade 2 bleeding decreased significantly as aggregation increased and Multiplate® had a negative predictive value (NPV) of 96% and a positive predictive value (PPV) of 19% for significant bleeding. In the multiple logistic regression model collagen-activated Multiplate® aggregation, TEG angle, TEG reaction time and CRP significantly affected the odds of WHO grade 2 bleeding. The combined model had a NPV of 99% and a PPV of 19%. DISCUSSION: Our findings suggest that the markers of platelet function and haemostasis provided by Multiplate® aggregometry and TEG may add information to support prediction of bleeding, although platelet count still remains the most accessible analysis for routine testing.

Hemostatic efficacy of pathogen-inactivated vs untreated platelets: a randomized controlled trial.

Pathogen inactivation of platelet concentrates reduces the risk for blood-borne infections. However, its effect on platelet function and hemostatic efficacy of transfusion is unclear. We conducted a randomized noninferiority trial comparing the efficacy of pathogen-inactivated platelets using riboflavin and UV B illumination technology (intervention) compared with standard plasma-stored platelets (control) for the prevention of bleeding in patients with hematologic malignancies and thrombocytopenia. The primary outcome parameter was the proportion of transfusion-treatment periods in which the patient had grade 2 or higher bleeding, as defined by World Health Organization criteria. Between November 2010 and April 2016, 469 unique patients were randomized to 567 transfusion-treatment periods (283 in the control arm, 284 in the intervention arm). There was a 3% absolute difference in grade 2 or higher bleeding in the intention-to-treat analysis: 51% of the transfusion-treatment periods in the control arm and 54% in the intervention arm (95% confidence interval [CI], -6 to 11; P = .012 for noninferiority). However, in the per-protocol analysis, the difference in grade 2 or higher bleeding was 8%: 44% in the control arm and 52% in the intervention arm (95% CI -2 to 18; P = .19 for noninferiority). Transfusion increment parameters were ∼50% lower in the intervention arm. There was no difference in the proportion of patients developing HLA class I alloantibodies. In conclusion, the noninferiority criterion for pathogen-inactivated platelets was met in the intention-to-treat analysis. This finding was not demonstrated in the per-protocol analysis. This trial was registered at The Netherlands National Trial Registry as #NTR2106 and at www.clinicaltrials.gov as #NCT02783313.

Immediate effects of blood donation on physical and cognitive performance-A randomized controlled double-blinded trial.

BACKGROUND: The success of implementing damage control resuscitation principles pre-hospital has been at the expense of several logistic burdens including the requirements for resupply, and the question of donor safety during the development of whole blood programs. Previous studies have reported effects on physical performance after blood donation; however, none have investigated the effects of blood donation on cognitive performance. METHOD: We describe a prospective double-blinded, randomized, controlled study comprised of a battery of tests: three cognitive tests, and VO2max testing on a cycle ergometer. Testing was performed 7 days before blinded donation (baseline day), immediately after donation (Day 0), and 7 days (Day 7) after donation. The inclusion criteria included being active blood donors at the Haukeland University Hospital blood bank, where eligibility requirements were met on the testing days, and providing informed consent. Participants were randomized to either the experimental (n = 26) or control group (n = 31). Control group participants underwent a 'mock donation" in which a phlebotomy needle was placed but blood was not withdrawn. RESULTS: In the experimental group, mean ± SEM VO2max declined 6% from 41.35 ± 1.7 mLO2/(min·kg) at baseline to 39.0 ± 1.6 mLO2/(min·kg) on Day 0 and increased to 40.51 ± 1.5 mLO2/(min·kg) on Day 7. Comparable values in the control group were 42.1 ± 1.8 mLO2/(min·kg) at baseline, 41.6 ± 1.8 mLO2/(min·kg)) on Day 1 (1% decline from baseline), and 41.8 ± 1.8 mLO2/(min·kg) on Day 7.Comparing scores of all three cognitive tests on Day 0 and Day 7 showed no significant differences (p > 0.05). CONCLUSION: Our main findings are that executive cognitive and physical performances were well maintained after whole blood donation in healthy blood donors. The findings inform postdonation guidance on when donors may be required to return to duty. LEVEL OF EVIDENCE: Randomized, controlled, double-blinded prospective trial study, level 1.

The healthy donor profile of immunoregulatory soluble mediators is altered by stem cell mobilization and apheresis.

BACKGROUND: Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and thereafter harvested by leukapheresis are commonly used for allogeneic stem cell transplantation. METHODS: Plasma levels of 38 soluble mediators (cytokines, soluble adhesion molecules, proteases, protease inhibitors) were analyzed in samples derived from healthy stem cell donors before G-CSF treatment and after 4 days, both immediately before and after leukapheresis. RESULTS: Donors could be classified into two main subsets based on their plasma mediator profile before G-CSF treatment. Seventeen of 36 detectable mediators were significantly altered by G-CSF; generally an increase in mediator levels was seen, including pro-inflammatory cytokines, soluble adhesion molecules and proteases. Several leukocyte- and platelet-released mediators were increased during apheresis. Both plasma and graft mediator profiles were thus altered and showed correlations to graft concentrations of leukocytes and platelets; these concentrations were influenced by the apheresis device used. Finally, the mediator profile of the allotransplant recipients was altered by graft infusion, and based on their day +1 post-transplantation plasma profile our recipients could be divided into two major subsets that differed in overall survival. DISCUSSION: G-CSF alters the short-term plasma mediator profile of healthy stem cell donors. These effects together with the leukocyte and platelet levels in the graft determine the mediator profile of the stem cell grafts. Graft infusion also alters the systemic mediator profile of the recipients, but further studies are required to clarify whether such graft-induced alterations have a prognostic impact.

Preparation of leukoreduced whole blood for transfusion in austere environments; effects of forced filtration, storage agitation, and high temperatures on hemostatic function.

BACKGROUND: Damage control resuscitation principles advocate the use of blood to treat traumatic hemorrhage. Hemorrhage is a leading cause of preventable death on the battlefield, but making blood components available far forward presents logistical challenges due to shelf life and storage requirements. Whole blood simplifies logistics and enables collection in the field but can cause leukocyte-related transfusion reactions. A field-adapted leukoreduction system must be fast and safe, and storage of whole blood should preserve hemostatic function. METHODS: Blood was collected using Imuflex WB-SP and leukoreduced at 0, 150, or 300 mm Hg. Additional bags were stored at 4°C for 21 days unagitated, mixed daily, agitated or head-over-heel rotated, at 22°C for 3 days, or 32°C for 2 hours. Hematology, coagulation, CD62P/CD42b, thromboelastography (TEG)/thromboelastometry (ROTEM), and Multiplate was performed. RESULTS: Filtration time was 35 ± 1, 14 ± 0, and 9 ± 0 minutes at 0, 150, and 300 mm Hg, respectively. One of 10 units at 150 mm Hg and 4 of 11 at 300 mm Hg had residual whole blood cells greater than 5.0 × 10 per unit. One of 11 at 300 mm Hg had platelet recovery of less than 80%. Hemolysis was less than 0.2%. Filtration decreased thromboelastography/thromboelastometry and Multiplate aggregation response. Stored at 4°C, α and MA/MCF moderately decreased regardless of mixing. Significant loss of aggregation response and increased CD62P expression was seen by Day 10. By Day 3, storage at 22°C caused loss of most aggregation. Two-hour storage at 32°C did not significantly affect hemostatic capacity. CONCLUSION: Forced filtration reduced leukoreduction time, but increased residual whole blood cells reduced hemostatic function. Aggregation response deteriorated early in storage, while viscoelastic assays decreased more gradually. Mixing showed no benefits. LEVEL OF EVIDENCE: Diagnostic study, level IV.

Staff officers as blood suppliers: Effects of repeated donations and autologous reinfusions of untransfused units.

BACKGROUND: Limited blood inventory and resupply chains in combat settings can result in preventable deaths from traumatic hemorrhage. One way of mitigating this could be to establish donor pools where blood is collected in advance of high-risk missions and then reinfused back to the donor if not needed to treat casualties. METHODS: Four hundred fifty milliliters plus 56 mL of blood was collected, rested for 2 hours in room temperature, and stored at 4°C. The blood was reinfused 22 to 24 hours after donation and the donor observed for adverse reactions. Samples were collected before and 20 minutes after each donation for hematology, immunoglobulin G, ferritin, C-reactive protein, total protein, lactate dehydrogenase, bilirubin, haptoglobin, and activated partial thromboplastin time. RESULTS: Nine participants went through a total of 36 donation and reinfusion procedures. Four donors participated in five rounds, two in four rounds, two in three rounds, and one in two rounds. A significant drop was seen in hemoglobin (14.6 ± 0.9 to 13.9 ± 0.9) and ferritin (179 ± 70 to 149 ± 78) from before the first donation to after the last reinfusion (p < 0.05). Other parameters were unaffected. CONCLUSION: This small pilot study suggests that repeated donations and reinfusions may be both feasible and safe. Blood collected in this way should be labeled with the donor's full name and social security number (or similar) and the identity visually verified by the donor immediately before both donation and reinfusion. To further reduce risk, this form of donation should be restricted to scenarios where there is no other option for making blood available. LEVEL OF EVIDENCE: Therapeutic/Care management study, level V.

Efficacy of Humanized Mesenchymal Stem Cell Cultures for Bone Tissue Engineering: A Systematic Review with a Focus on Platelet Derivatives.

Fetal bovine serum (FBS) is the most commonly used supplement for ex vivo expansion of human mesenchymal stem cells (hMSCs) for bone tissue engineering applications. However, from a clinical standpoint, it is important to substitute animal-derived products according to current good manufacturing practice (cGMP) guidelines. Humanized alternatives to FBS include three categories of products: human serum (HS), human platelet derivatives (HPDs)-including platelet lysate (PL) or platelet releasate (PR), produced by freeze/thawing or chemical activation of platelet concentrates, respectively, and chemically defined media (serum-free) (CDM). In this systematic literature review, the in vitro and in vivo osteogenic potential of hMSCs expanded in humanized (HS-, HPD-, or CDM-supplemented) media versus hMSCs expanded in FBS-supplemented media, was compared. In addition, PL and PR were compared in terms of their growth factor (GF)/cytokine-content and cell-culture efficacy. When using either 10-20% autologous or pooled HS, 3-10% pooled HPDs or CDM supplemented with GFs, in comparison with 10-20% FBS, a majority of studies reported similar or superior in vitro proliferation and osteogenic differentiation, and in vivo bone formation in ectopic or orthotopic rodent models. Moreover, a trend for higher GF content was observed in PL versus PR, although evidence for cell culture efficacy is limited. In summary, humanized supplements seem at least equally effective as FBS for hMSC expansion and osteogenic differentiation. Although pooled HPDs appear to be the most favorable supplement for large-scale hMSC expansion, further efforts are needed to standardize the preparation and composition of these products in compliance with cGMP standards.

Immunomodulation Induced by Stem Cell Mobilization and Harvesting in Healthy Donors: Increased Systemic Osteopontin Levels after Treatment with Granulocyte Colony-Stimulating Factor.

Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and harvested by leukapheresis are commonly used for allogeneic stem cell transplantation. The frequency of severe graft versus host disease is similar for patients receiving peripheral blood and bone marrow allografts, even though the blood grafts contain more T cells, indicating mobilization-related immunoregulatory effects. The regulatory phosphoprotein osteopontin was quantified in plasma samples from healthy donors before G-CSF treatment, after four days of treatment immediately before and after leukapheresis, and 18-24 h after apheresis. Myeloma patients received chemotherapy, combined with G-CSF, for stem cell mobilization and plasma samples were prepared immediately before, immediately after, and 18-24 h after leukapheresis. G-CSF treatment of healthy stem cell donors increased plasma osteopontin levels, and a further increase was seen immediately after leukapheresis. The pre-apheresis levels were also increased in myeloma patients compared to healthy individuals. Finally, in vivo G-CSF exposure did not alter T cell expression of osteopontin ligand CD44, and in vitro osteopontin exposure induced only small increases in anti-CD3- and anti-CD28-stimulated T cell proliferation. G-CSF treatment, followed by leukapheresis, can increase systemic osteopontin levels, and this effect may contribute to the immunomodulatory effects of G-CSF treatment.

A study protocol for a randomised controlled trial evaluating clinical effects of platelet transfusion products: the Pathogen Reduction Evaluation and Predictive Analytical Rating Score (PREPAReS) trial.

INTRODUCTION: Patients with chemotherapy-induced thrombocytopaenia frequently experience minor and sometimes severe bleeding complications. Unrestrictive availability of safe and effective blood products is presumed by treating physicians as well as patients. Pathogen reduction technology potentially offers the opportunity to enhance safety by reducing bacterial and viral contamination of platelet products along with a potential reduction of alloimmunisation in patients receiving multiple platelet transfusions. METHODS AND ANALYSIS: To test efficacy, a randomised, single-blinded, multicentre controlled trial was designed to evaluate clinical non-inferiority of pathogen-reduced platelet concentrates treated by the Mirasol system, compared with standard plasma-stored platelet concentrates using the percentage of patients with WHO grade ≥ 2 bleeding complications as the primary endpoint. The upper limit of the 95% CI of the non-inferiority margin was chosen to be a ≤ 12.5% increase in this percentage. Bleeding symptoms are actively monitored on a daily basis. The adjudication of the bleeding grade is performed by 3 adjudicators, blinded to the platelet product randomisation as well as by an automated computer algorithm. Interim analyses evaluating bleeding complications as well as serious adverse events are performed after each batch of 60 patients. The study started in 2010 and patients will be enrolled up to a maximum of 618 patients, depending on the results of consecutive interim analyses. A flexible stopping rule was designed allowing stopping for non-inferiority or futility. Besides analysing effects of pathogen reduction on clinical efficacy, the Pathogen Reduction Evaluation and Predictive Analytical Rating Score (PREPAReS) is designed to answer several other pending questions and translational issues related to bleeding and alloimmunisation, formulated as secondary and tertiary endpoints. ETHICS AND DISSEMINATION: Ethics approval was obtained in all 3 participating countries. Results of the main trial and each of the secondary endpoints will be submitted for publication in a peer-reviewed journal. TRIAL REGISTRATION NUMBER: NTR2106; Pre-results.

Survival after ultramassive transfusion: a review of 1360 cases.

BACKGROUND: Information about patient survival after transfusion of multiple blood volumes is limited, and most reports have focused on trauma patients. STUDY DESIGN AND METHODS: Retrospective study of blood use and survival at 11 hospitals in six nations between 2009 and 2013. Ultramassive transfusion (UMT) was defined as transfusion of 20 or more red blood cell (RBC) units over the course of any 2 consecutive calendar days. RESULTS: A total of 1975 patients received UMT and a representative sample of 1360 patients was studied in detail. Patients were grouped into seven diagnostic categories: solid organ transplantation (n = 411), cardiac or major vascular surgery (n = 317), general surgery (n = 228), trauma (n = 221), general medicine (n = 124), obstetrics (n = 23), and other (n = 36). During the 7 days after initiation of UMT, these patients used more than 120,000 blood components. The median (interquartile range) blood use was 35 (26-50) RBC units, 30 (20-47) plasma units, and 7 (4-13) platelet doses. Five- and 30-day survival significantly declined with increasing RBC use. Overall survivals of patients receiving UMT were 71% (5 day) and 60% (30 day), and in the subset of 165 patients receiving 60 or more RBC units over 2 consecutive days, 5-day survival was 54% ranging from 17% (trauma) to 75% (solid organ transplant). The decline in survival with increasing RBC transfusions was minimal for patients undergoing solid organ transplantation and was most pronounced for trauma and nonsurgical bleeding patients. CONCLUSION: Trauma was not the leading cause of UMT. Increasing RBC requirements were significantly associated with decreasing survival. However, survival was more strongly associated with diagnostic category than total RBCs transfused, with highest survival rates in solid organ transplant surgery.