Cancer type predicts alloimmunization following RhD-incompatible RBC transfusions.

BACKGROUND: Immunosuppressed, RhD-negative oncology patients tend to have lower rates of sensitization to the D antigen when they receive transfusion with RhD-positive blood components. Clinical factors associated with alloimmunization to the D antigen in RhD-negative oncology patients when they receive transfusion with RhD-positive red blood cells (RBCs) have not been well defined. STUDY DESIGN AND METHODS: This was a 4-year, retrospective analysis identifying RhD-negative oncology patients who received RhD-positive RBCs and were not previously alloimmunized to the D antigen. Age, sex, race, ABO group, primary oncology diagnosis, and numbers of RhD-incompatible RBC transfusions were recorded. The association between antibody formation and clinical factors was studied. The incidence of alloanti-D was calculated from a subsequent antibody-detection test performed at least 28 days after receipt of the first transfusion of RhD-positive RBCs. RESULTS: In total, 545 RhD-negative oncology patients received 4295 RhD-positive RBC transfusions. Of these, 76 (14%) became alloimmunized to the D antigen. Diagnosis type was the only factor significantly associated with responder status. The logistic regression model indicated that patients who had myelodysplastic syndrome or solid malignancies were more likely to be responders than those who had acute leukemia. CONCLUSION: We measured a 14% sensitization rate to the D antigen in our RhD-negative oncology population. The rate of alloimmunization was higher in patients who had solid cancers (22.6%) or myelodysplastic syndrome (23%) compared with those who had other hematologic malignancies (7%). Knowledge of diagnoses that predispose to RhD alloimmunization enables better utilization of RhD-negative RBCs during times of shortage.

Pure Red Cell Aplasia in Major ABO-Mismatched Allogeneic Hematopoietic Stem Cell Transplantation Is Associated with Severe Pancytopenia.

In major ABO-mismatched allogeneic hematopoietic stem cell transplantation (HSCT) persistence of antidonor isohemagglutinins leads to pure red cell aplasia (PRCA). To investigate severe pancytopenia noted in a previous study of PRCA, we analyzed all major ABO-mismatched HSCT between January 2003 and December 2012. Of 83 PRCA patients, 13 (16%) had severe pancytopenia. Severe pancytopenia was defined as an absolute neutrophil count (ANC) < 1.5 K/µL or requiring granulocyte colony-stimulating factor, platelets < 50 K/µL or transfusion dependent, and PRCA with RBC transfusion dependence at post-transplant day 90. In 6 patients (46%) severe pancytopenia resolved after PRCA resolution. Two patients (15%) received a second transplant because of persistent pancytopenia/secondary graft failure, 1 (8%) died from secondary graft failure despite a stem cell boost, 1 (8%) did not recover his platelet counts despite RBC/ANC recovery, and 3 patients (23%) died from disease relapse. We found that severe pancytopenia is frequently associated with PRCA in 16% of major ABO-incompatible HSCT with a higher incidence in males and pancytopenia resolved with resolution of PRCA in 46% of patients.

Complement-Binding Donor-Specific Anti-HLA Antibodies and Risk of Primary Graft Failure in Hematopoietic Stem Cell Transplantation.

Detection of donor-specific anti-HLA antibodies (DSA) has been associated with graft rejection in all forms of transplantation. The mechanism by which DSA increase the risk of graft failure remains unclear. We hypothesized that complement-binding DSA are associated with engraftment failure in hematopoietic stem cell transplantation (HSCT) and analyzed 122 haploidentical transplant recipients tested prospectively for DSA. Retrospective analysis to detect C1q binding DSA (C1q+DSA) was performed on 22 allosensitized recipients. Twenty-two of 122 patients (18%) had DSA, 19 of which were women (86%). Seven patients with DSA (32%) rejected the graft. Median DSA level at transplant for patients who failed to engraft was 10,055 mean fluorescence intensity (MFI) versus 2065 MFI for those who engrafted (P = .007). Nine patients with DSA were C1q positive in the initial samples with median DSA levels of 15,279 MFI (range, 1554 to 28,615), compared with 7 C1q-negative patients with median DSA levels of 2471 MFI (range, 665 to 12,254) (P = .016). Of 9 patients who were C1q positive in the initial samples, 5 patients remained C1q positive at time of transplant (all with high DSA levels [median, 15,279; range, 6487 to 22,944]) and experienced engraftment failure, whereas 4 patients became C1q negative pretransplant and all engrafted the donor cells (P = .008). In conclusion, patients with high DSA levels (>5000 MFI) and complement-binding DSA antibodies (C1q positive) appear to be at much higher risk of primary graft failure. The presence of C1q+DSA should be assessed in allosensitized patients before HSCT. Reduction of C1q+DSA levels might prevent engraftment failure in HSCT.

Design, development, and validation of a high-throughput drug-screening assay for targeting of human leukemia.

BACKGROUND: The authors developed an ex vivo methodology to perform drug library screening against human leukemia. METHODS: The strategy for this screening relied on human blood or bone marrow cultures under hypoxia; under these conditions, leukemia cells deplete oxygen faster than normal cells, causing a hemoglobin oxygenation shift. Several advantages were observed: 1) partial recapitulation of the leukemia microenvironment, 2) use of native hemoglobin oxygenation as a real-time sensor/reporter, 3) cost-effectiveness, 4) species specificity, and 5) a format that enables high-throughput screening. RESULTS: For a proof of concept, a chemical library (size, approximately 20,000 compounds) was screened against human leukemia cells. Seventy compounds were identified ("hit" rate, 0.35%; Z-factor = 0.71) that had activity, and 20 compounds were examined to identify 18 true-positive compounds (90%). Finally, the results demonstrated that carbonohydraxonic diamide group-containing compounds are potent antileukemia agents that induce cell death in leukemia cells and in patient-derived samples. CONCLUSIONS: The current results indicated that this unique functional assay can identify novel drug candidates and can help with the development of future applications in personalized drug selection for patients with leukemia.

Septic transfusion reactions during blood transfusion via indwelling central venous catheters.

BACKGROUND: Transfusion of blood products requires a vascular port. Use of an indwelling central venous catheter (CVC) provides this port readily and safely in general; however, potential risks require assessment. STUDY DESIGN AND METHODS: The objective was to examine septic reactions to blood transfusions performed via CVCs owing to subclinical microbial catheter colonization. All transfusion reactions that occurred from 2007 to 2011 at The University of Texas MD Anderson Cancer Center were analyzed and correlated with microbiology culture results. Data on the reactions, including vascular access via a catheter or peripheral venipuncture, were collected prospectively. RESULTS: A total of 999 reactions were reported, with an incidence of two per 1000 transfusion events. A total of 738 reactions occurred in 642 patients during transfusion through a CVC. Among them, 606 reactions occurred in patients that had cultures of blood samples drawn within 7 days before or after reaction. Sixty of these (9.9%) had at least one significant microorganism isolated from their catheters and/or peripheral blood. The blood culture results and timing suggested that these patients likely had catheter-related bloodstream infections caused by transfusion through a CVC with subclinical microbial colonization. Fever and chills occurred in 35 of these patients (58%), which resembled febrile nonhemolytic transfusion reactions. Culture results of the transfused blood products, although not performed in all cases, were mostly negative in these CVC-related reactions. CONCLUSION: Blood transfusion through an indwelling CVC may lead to septic reaction owing to subclinical microbial colonization. This risk should be considered before transfusion and during investigation of transfusion reactions.

Incidence and natural history of pure red cell aplasia in major ABO-mismatched haematopoietic cell transplantation.

Major ABO mismatching is not considered a contraindication to allogeneic haematopoietic stem cell transplantation (HSCT). Modern reduced-intensity conditioning and reduced-toxicity regimens cause much less myeloablation than conventional myeloablative regimens, such as cyclophosphamide with busulfan or total body irradiation, which may affect the incidence of pure red cell aplasia (PRCA). We estimated the incidence and described the natural history of PRCA in patients with major ABO-mismatched donor stem cells. Between 2007 and 2008, 161 (27% of all patients undergoing HSCT) underwent allogeneic HSCT with major ABO-mismatched stem cells and 12 (7·5%) of these patients developed PRCA. Thirty and ninety day T-cell and myeloid cell chimerism and neutrophil and platelet engraftment did not differ between patients who developed PRCA and those who did not. The only risk factor associated with PRCA was the use of a fludarabine/busulfan conditioning regimen. All patients with PRCA needed red cell transfusion for several months after HSCT resulting in significant iron overload. Pure red cell aplasia resolved spontaneously in the majority (seven patients) but only resolved after stopping tacrolimus in three patients. Hence, after major ABO-mismatched HSCT, the incidence of PRCA was 7·5% and it resolved spontaneously or after withdrawal of immunosuppression in the majority of patients.

Targeting neuropilin-1 in human leukemia and lymphoma.

Targeted drug delivery offers an opportunity for the development of safer and more effective therapies for the treatment of cancer. In this study, we sought to identify short, cell-internalizing peptide ligands that could serve as directive agents for specific drug delivery in hematologic malignancies. By screening of human leukemia cells with a combinatorial phage display peptide library, we isolated a peptide motif, sequence Phe-Phe/Tyr-Any-Leu-Arg-Ser (F(F)/(Y)XLRS), which bound to different leukemia cell lines and to patient-derived bone marrow samples. The motif was internalized through a receptor-mediated pathway, and we next identified the corresponding receptor as the transmembrane glycoprotein neuropilin-1 (NRP-1). Moreover, we observed a potent anti-leukemia cell effect when the targeting motif was synthesized in tandem to the pro-apoptotic sequence (D)(KLAKLAK)2. Finally, our results confirmed increased expression of NRP-1 in representative human leukemia and lymphoma cell lines and in a panel of bone marrow specimens obtained from patients with acute lymphoblastic leukemia or acute myelogenous leukemia compared with normal bone marrow. These results indicate that NRP-1 could potentially be used as a target for ligand-directed therapy in human leukemias and lymphomas and that the prototype CGFYWLRSC-GG-(D)(KLAKLAK)2 is a promising drug candidate in this setting.

Technical Note: Use of a Novel Bacterial Culture Sampling Device for Pooled Whole Blood Platelets.

OBJECTIVE: Transfusion associated sepsis is a serious risk after platelet transfusion. Although platelet culture can be performed to avoid such risk, culture results are often available after transfusion due to the 4-hour shelf-life after pooling. To decrease such risk, we implemented a needleless closed system device to culture for contamination before pooling and release for transfusion. METHODS: We customized a needleless device to permit sterile sampling of whole blood platelets without retrograde or cross-contamination. Then aliquots of platelets were injected into culture media for detection of aerobic organisms and cultured for 24 hours but released for transfusion after 12 hours of negative culture. RESULTS: In a period of two years, we used this device in 5,741 whole blood derived pooled platelets and only 24 units tested positive (0.4%) but none of initial positive was later confirmed. There were 11 Staphylococcus and 9 Bacillus species identified. All but one of the positive units were discarded; there was no clinical impact in the patient who received the positive unit. CONCLUSION: This device allows for sampling of whole blood derived platelets before pooling, warranting a transfusion of a culture negative unit after 12 hours of negative culture, consequently reducing transfusion of bacterially contaminated whole blood derived pooled platelets.

Impact of safety concerns and regulatory changes on the usage of erythropoiesis-stimulating agents and RBC transfusions.

PURPOSE: Safety concerns raised in the recent oncology trials with erythropoiesis-stimulating agents (ESAs) have led to regulatory restrictions on their use. We wished to determine the impact of these changes on the use of ESAs and RBC transfusions. METHODS: In a retrospective observational study of patients treated at a comprehensive cancer center in 2006-2008, data on all ESA doses dispensed, RBCs transfused, and hemoglobin levels on the days of transfusions and ESA initiations were analyzed. RESULTS: Compared with 2006, the total patients treated was 14% higher (28,339 versus 24,806) in 2007 and 22% higher (30,254) in 2008. Patients receiving ESAs decreased by 26% and 61%, and ESA units dispensed decreased by 29% (from 30,206 units to 21,409 units) and 80% (6,102 units) in 2007 and 2008, respectively. However, RBC transfusions increased by only 2% (from 38,218 units to 38,948 units) in 2007 and by 8% (41,438) in 2008. The mean hemoglobin on the day of transfusion was the same for each year (8.4 g/dl); however, an increasing proportion of patients initiated ESAs at lower hemoglobin (< 10 g/dl) levels. After adjusting for demographics and diagnostic variables for 3 years (n = 83,399), a multivariate logistic regression showed a significant decline in ESA use (p < .0001) without an increase in RBC transfusions. CONCLUSIONS: Recent ESA safety concerns and regulatory restrictions have significantly decreased ESA use. The lack of a significant impact on transfusions may be related to a lower hemoglobin threshold used to initiate ESAs or treatment of patients less likely to respond.

A portable system for processing donated whole blood into high quality components without centrifugation.

BACKGROUND: The use of centrifugation-based approaches for processing donated blood into components is routine in the industrialized world, as disparate storage conditions require the rapid separation of 'whole blood' into distinct red blood cell (RBC), platelet, and plasma products. However, the logistical complications and potential cellular damage associated with centrifugation/apheresis manufacturing of blood products are well documented. The objective of this study was to evaluate a proof-of-concept system for whole blood processing, which does not employ electromechanical parts, is easily portable, and can be operated immediately after donation with minimal human labor. METHODS AND FINDINGS: In a split-unit study (n = 6), full (~500mL) units of freshly-donated whole blood were divided, with one half processed by conventional centrifugation techniques and the other with the new blood separation system. Each of these processes took 2-3 hours to complete and were performed in parallel. Blood products generated by the two approaches were compared using an extensive panel of cellular and plasma quality metrics. Comparison of nearly all RBC parameters showed no significant differences between the two approaches, although the portable system generated RBC units with a slight but statistically significant improvement in 2,3-diphosphoglyceric acid concentration (p < 0.05). More notably, several markers of platelet damage were significantly and meaningfully higher in products generated with conventional centrifugation: the increase in platelet activation (assessed via P-selectin expression in platelets before and after blood processing) was nearly 4-fold higher for platelet units produced via centrifugation, and the release of pro-inflammatory mediators (soluble CD40-ligand, thromboxane B2) was significantly higher for centrifuged platelets as well (p < 0.01). CONCLUSION: This study demonstrated that a simple, passive system for separating donated blood into components may be a viable alternative to centrifugation-particularly for applications in remote or resource-limited settings, or for patients requiring highly functional platelet product.

Quantitative real-time polymerase chain reaction for detection of circulating cells with t(14;18) in volunteer blood donors and patients with follicular lymphoma.

The chromosomal rearrangement t(14;18)(q32;21) involves the major (MBR) or minor (mcr) breakpoint cluster regions and the immunoglobulin heavy chain joining regions (JH) in most follicular lymphomas. As a first step towards determining the clinical significance of circulating cells with t(14;18), we detected and quantitated circulating cells in samples obtained from volunteer blood donors and follicular lymphoma patients. The t(14;18) was co-amplified with beta-actin with real-time quantitative PCR (QRT-PCR) in reactions containing 1 microg of DNA from peripheral blood or bone marrow aspirates. The cell number was quantitated using linear regression and an external standard of serially diluted DNA from cell lines with MBR/JH or mcr/JH rearrangements. At dilutions of 10(5) and 106, sensitivity was 100 and 55% for MBR/JH, and 100 and 10% for mcr/JH rearrangements. Among 102 volunteer blood donors MBR/JH vs. mcr/JH amplicons were detected in 22 vs. 4% with duplicate 1 microg DNA reactions, and in 41 vs. 6% with a total 10 microg DNA analyzed in multiple reactions. Among volunteer blood donors the mean number of circulating cells with MBR/JH vs. mcr/JH rearrangements were 0.8 vs. 0.1/microg DNA, and exceeded the upper normal limit (defined as the mean of all volunteer samples plus two standard deviations) in 3% vs. 2%, respectively. Analysis for MBR/JH rearrangements revealed that follicular lymphoma patients vs. volunteer blood donors were positive in 76% vs. 22% (p = 0.008 by Fisher's exact test); that the mean number of MBR/JH cells per microg of DNA was 91 vs. 0.8 (p = 0.0002 by Mann-Whitney test); and the number of the MBR/JH cells exceeded the upper normal limit in 32% vs. 3% of subjects (p = 0.0001 by Fisher's exact test). Circulating cells with mcr/JH were not detected among any of these 25 lymphoma patients. We conclude that patients with follicular lymphoma are more frequently positive, have higher numbers of circulating cells with t(14;18), which exceed upper normal limit more frequently than in volunteer blood donors.

Prevalence of hepatitis C in a drug using and non-using welfare population.

CONTEXT: Drug use is a primary route for the transmission of the Hepatitis C virus (HCV). A substantial proportion of welfare recipients have been shown to be substance abusers. In addition, federal legislation has imposed limits on the number of months individuals may receive benefits and has mandated most recipients to participate in a 'work activity' in exchange for benefits. HCV symptoms may inhibit welfare recipients' ability to seek and maintain employment. OBJECTIVE: To assess the prevalence of HCV in a sample of Temporary Assistance to Needy Families (TANF) recipients and the effects of HCV antibody seropositivity on employability. DESIGN, SETTING, AND PARTICIPANTS: The sample for this study consisted of 380 individuals participating in a longitudinal study of employment patterns among TANF recipients in Houston, TX, funded by the National Institute on Drug Abuse (NIDA). Private interviews regarding welfare receipt, employment, and drug use were conducted at intake into the study and at 4-month intervals for one year. Participants agreed to a one-time blood test. Blood samples were tested for the presence of HCV antibodies by enzyme linked immunoassay. MAIN OUTCOME MEASURE: Employment status over time by HCV antibody status. RESULTS: Overall, 12% of all participants tested positive for the presence of HCV antibodies. A significantly greater proportion of chronic drug users (31%) than non-drug users (4%) tested positive for the presence of Hepatitis C antibodies. Those who tested positive for hepatitis C had significantly lower rates of employment. CONCLUSIONS: Potential infection with Hepatitis C may constitute an employment barrier for many welfare recipients.

Transfusion of RhD-incompatible blood components in RhD-negative blood marrow transplant recipients.

BACKGROUND: Transfusion-dependent bone marrow transplant recipients are routinely transfused with ABO group and RhD-compatible blood components. However, because of the scarcity of RhD-negative blood components, particularly platelets, a policy was developed to transfuse RhD-positive blood components to RhD-negative patients during periods of shortage. METHODS: We reviewed the records of 78 RhD-negative patients with hematologic malignancies who received RhD-negative bone marrow and/or peripheral blood stem cells, from June 1995 to August 2000. The patients transfused with RhD-incompatible blood components were screened periodically for evidence of the development of red blood cell (RBC) alloimmunization. RESULTS: Three of 78 patients (4%) developed anti-D antibodies after receiving RhD-incompatible platelet transfusions. One of the patients developed evidence of anti-RhD antibodies after receiving 42 units of RhD-positive random donor platelets; the second patient developed such evidence after receiving 6 apheresis platelets and 2 infusions of intravenous immunoglobulin G (positive for anti-RhD). The third patient received 206 RhD-positive random donor platelets and 5 apheresis units. All patients were discharged from the hospital. The overall immunization rate was 4%. Six patients received Rh-incompatible packed RBCs and showed no evidence of neither anti-RhD nor any other anti-RBC antibodies. All 78 patients had received RhD-incompatible platelets throughout their engraftment period. CONCLUSION: Transfusion of RhD-positive blood components to Rh-negative patients with hematologic cancers, who have received RhD-negative bone marrow and/or peripheral blood stem cells, are at low risk of developing RhD antibodies. These findings allow for a flexible strategy of blood component therapy support for this special patient population during periods of shortage.

Underreporting of minor transfusion reactions in cancer patients.

BACKGROUND: Minor adverse reactions following transfusion of blood components to cancer patients are not uncommon. Reporting these minor reactions to the transfusion service needs a careful evaluation. The objectives of this study were to closely monitor the transfusion reactions that occurred and had not been reported to the transfusion service and to evaluate the process by which the medical and nursing staff recognized and managed these reactions. METHODS: We prepared a questionnaire with the nursing staff of a selected inpatient unit that addressed important questions, such as signs and symptoms during the transfusion, premedications given, process for physician notification, recommended action, and blood component implicated. Charts of the patients were reviewed, and the process was monitored for a 6-month period. RESULTS: A total of 58 cases were completed and analyzed. Platelet concentrates were transfused in 43 cases (74.1%), packed red blood cells in 9 cases (15.6%), and fresh frozen plasma in 6 cases (10.3%). Minor adverse reactions that were documented included chills in 11 cases (19.0%), low-grade fever in 11 cases (19.0%), hives and itching in 24 cases (41.4%), nausea and vomiting in 1 case (1.7%), and headaches and nonspecific mild pains in 11 cases (19.0%). Transfusions had been resumed in 27 cases (46.6%) and stopped completely in 13 cases (22.4%). Twenty-seven of 58 (46.6%) were first-time events. CONCLUSION: We conclude that underreporting of minor transfusion reactions, such as a febrile nonhemolytic transfusion reaction and allergic reactions, exists. To ensure safety to our cancer patients who are transfusion-dependent, we suggest that careful evaluation of any suspected transfusion reaction event should be referred to the transfusion medicine physicians who will evaluate each case and discuss it with the attending physician. This process will prevent detrimental, acute transfusion reactions.

Granulocyte concentrates from a single high-yield apheresis can be split to support multiple patients.

Allogeneic granulocyte transfusion has evolved into a viable therapeutic option for immunocompromised severely neutropenic leukemic patients and those with hematopoietic stem cell transplant with life-threatening bacterial and fungal infections. The collection of larger cell doses of granulocyte concentrates (GCs) has been facilitated by the stimulation of donors with granulocyte colony stimulating factor (G-CSF) and dexamethasone. The synergistic effect of G-CSF and dexamethasone has allowed the collection of larger cell doses of GCs and its use has increased steadily. This has allowed us to split the high-yield GC products and facilitated distribution of the split GC products to a second or third patient who needs GCs but lacks donors. The main objective of this article was to present our rationale for splitting GC products and how the split GC units were transfused to multiple patients. We believe that split GCs are as equally effective as unsplit GCs and that multiple patients benefit from splitting GCs.

Results of HLA antibody testing using ELISA vs the fluorescent bead method and retrospective review of data for recipients of packed RBCs and platelets from male HLA-immunized donors.

We reviewed HLA antibody testing results using an enzyme-linked immunosorbent assay (ELISA) for all male blood donors at our institution during a 3.5-month period to look for HLA immunization. Confirmatory testing of 33 blood samples positive for HLA class I and/or II antibodies was performed using the fluorescent bead method. A retrospective review of recipients of packed RBCs and platelets processed from these 33 HLA-immunized male donors were conducted to identify transfusion-related acute lung injury and cognate antigens. The agreement rates between the methods for HLA class I and II antibodies were 21% (7/33) and 6% (2/33), respectively. We noted HLA antibodies in the male donors corresponding to cognate antigens in 2 recipients of packed RBCs and in 3 recipients of platelets. Of 8 donors positive for HLA antibodies, 5 did not have a history of blood transfusion. We conclude that ELISA was too sensitive and had a high false-positive rate for the detection of HLA class II antibodies.

Impact on patient outcome following transfusion of bacterially contaminated platelets: the M.D. Anderson Cancer Center experience.

This study examined the clinical outcome of every patient who received a bacterially contaminated unit of platelets at The University of Texas M.D. Anderson Cancer Center, Houston, during 2007. Samples of platelets were aerobically cultured and read for 1 day at 35 degrees C. Positive bottles were subcultured in the appropriate media. The effect of independent variables in the clinical outcome of patients infused with bacterially contaminated platelet units was analyzed. A total of 23,199 platelet units were transfused, 71 of which were bacterially contaminated units; 8 were apheresis platelets and 63 were whole blood platelets. Of the 71 units, 70 were contaminated with gram-positive bacteria and 1 with gram-negative bacteria. Only 1 patient developed fever, and coagulase-negative staphylococci were isolated from the transfused unit. Transfusion of fresh units and antibiotic therapy possibly explain the lack of clinical consequences in our patients.