Granulocyte transfusions: Current science and perspectives

Seminars in Hematology 56 (2019) 241–247

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Granulocyte transfusions: Current science and perspectives Kamille A. West∗, Cathy Conry-Cantilena Department of Transfusion Medicine, National Institutes of Health Clinical Center, Bethesda, MD

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Keywords: Granulocyte transfusion Neutropenia Neutrophil dysfunction Bacteremia Invasive fungal infection Fungemia

a b s t r a c t Severe neutropenia renders patients susceptible to life-threatening bacterial and fungal infections. Despite improvements in supportive care and antimicrobial therapy, morbidity and mortality remains significant. Since the 1960s, granulocyte transfusions have been used to either treat or prevent serious infections in patients with neutropenia or neutrophil dysfunction. Despite significant optimizations in product collection, the practice of granulocyte transfusion therapy remains controversial. The use of granulocytes varies widely across institutions and countries in terms of indications, procurement, dose, infusion frequency, and duration of therapy. There are limited and conflicting data concerning its clinical effectiveness; current evidence from clinical trials does not support or refute efficacy. In this narrative review, we summarize the current evidence, discuss persistent concerns and consider future possibilities of the role of granulocyte transfusions. © 2019 Published by Elsevier Inc.

Basis for granulocyte transfusion Neutrophils or polymorphonuclear leukocytes (PMNs) are an essential component of the innate immune response and the first line of defense against a wide array of invading pathogens. Neutrophils are recruited to sites o inflammation, where their primary function is phagocytic ingestion and intracellular killing of invading bacteria and certain fungal elements, using antimicrobial peptides, enzymes, and reactive oxygen species [1]. Neutrophils can also kill microbes extracellularly by the release of granule proteins and chromatin to form neutrophil extracellular traps (NETs) [2,3]. Significant neutropenia or qualitative neutrophil dysfunction are therefore associated with a marked susceptibility to infection. In normal physiologic conditions, neutrophils represent the most abundant population (50%-70%) of circulating leukocytes, about 4.5 × 109 /L on average, with environmental and genetic factors accounting for variation between individuals and ethnic groups [4]. Neutrophil homeostasis is complex, with granulopoeisis, circulation in peripheral blood, intravascular margination and clearance operating in concert. Based on classic experiments performed with re-infusion of ex vivo-labeled neutrophils, it is commonly believed that neutrophils have a brief half-life of 6-8 hours in circulation, although one group suggests somewhat controversially that this period may be as long as 5 days [5]. To maintain

∗ Corresponding author. Kamille A. West, MD, Department of Transfusion Medicine, National Institutes of Health Clinical Center, 10 Center Drive, Room 1C711, Bethesda, MD 20892. E-mail addresses: [email protected] (K.A. West), [email protected] (C. Conry-Cantilena).

https://doi.org/10.1053/j.seminhematol.2019.11.002 0037-1963/© 2019 Published by Elsevier Inc.

circulating levels they must be produced by the marrow at a rate of 5-10 × 1010 cells/day [6]. The rationale of granulocyte transfusion (GTX) is to provide functional circulating granulocytes of a sufficient number that persist long enough to traffic to sites of infection and eliminate pathogens. Studies in the 1960s and 70s demonstrated that granulocyte transfusions from a variety of sources cleared bacteremia and reduced fever in severely neutropenic leukemia patients [7]. Despite these early studies and the hypothetical validity of the approach, the transfusion medicine community remains less convinced of the utility of GTX compared to other types of cellular transfusion therapy. For example, therapeutic red blood cell (RBC) transfusion is widely utilized in the management of anemia, although benefit has never been directly proven [8], and specific practice guidelines vary [9,10]. In contrast, granulocyte transfusions are infrequently used in practice; commonly cited reasons include the paucity of supporting evidence of efficacy, concern regarding potential adverse effects, and limited availability of high-quality products. Granulocyte procurement Granulocytes are collected by blood banks from healthy donors. Because granulocytes are only viable for 24 hours after collection, embarking on a course of granulocyte transfusion therapy requires planning and co-ordination. In the early history of granulocyte procurement, the difficulty in obtaining large doses of neutrophils from a single healthy donor was the limiting factor. Whole blood-derived buffy coats contain only about 5 × 108 to 1 × 109 neutrophils, but early data suggested that at least 1 × 1010 neutrophils would be required to achieve a therapeutic effect [11]. The

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Table 1 Content of granulocyte components according to procurement method.

Buffy Coat (single) [20] Pooled Buffy Coat [20] Apheresis (steroid only)a Apheresis (G-CSF ± steroid)a a

Volume (mL)

Hct (%)

Neutrophils (×1010 )

Lymphocytes (×109 )

Platelets (×109 )

55-65 232-272 215-330 300-375

35-59 18-24 10-15 10-15

0.04-0.2 0.7-1.17 1.2-2.7 4.0-8.4

0.6-2.8 5.2-8.2 1.5-2.0 1.5-5.0

50-105 226-585 50-180 50-250

Internal data, NIH.

Fig. 1. Apheresis granulocyte concentrate undergoing gravity sedimentation. (A) Unmanipulated apheresis granulocyte concentrate. (B) Granulocyte concentrate postsedimentation, with red blood cells sharply demarcated at the bottom of the product. (C) Red cell-depleted, granulocyte-rich aliquot separated postsedimentation. Images B and C courtesy of Barbara Jean Bryant, MD, FCAP, FASCP, University of Texas Medical Branch, and Susan F. Leitman, MD, National Institutes of Health Intramural Research Program.

advent of automated cell separators, starch sedimentation, and the use of steroids with or without growth factors to mobilize neutrophils into the circulation greatly improved the feasibility of collecting larger doses of granulocytes via apheresis. In the United States, apheresis granulocyte concentrates are most commonly utilized, collected from donors stimulated with steroids and containing approximately 1 to 2.5 × 1010 neutrophils [12]. Co-administration of granulocyte colony-stimulating factor (G-CSF) and a single dose of dexamethasone increases the yield to 4 to 8 × 1010 neutrophils or higher [13], however this strategy is used only in a few centers. This is due in part to fear of long-term adverse effects of repeated stimulation, particularly the theoretical risk of leukemia, although clinical data to date have failed to identify any increased incidence of AML in G-CSF mobilized donors [14,15] Identifying a suitable donor and administering medications prior to granulocyte apheresis requires at least 1 day of lead-time. Donor availability depends on the size of donor pool and the requirements of the patient (such as CMV status or HLA match). Many centers limit G-CSF-mobilized granulocyte collections to related donors or friends of the patient; maintaining a committed pool of unrelated community granulocyte donors is feasible, but difficult. The collection process entails minimal risk to carefullyscreened allogeneic donors with informed consent. The short-term side effects of G-CSF such as bone pain, headache, and myalgia are generally mild and treatable [16]; thus far, long-term follow-up of granulocyte donors stimulated with G-CSF and dexamethasone suggests that granulocyte donation is safe. [17,18] Alternatively, multiple buffy coats may be administered to achieve a sufficient granulocyte dose, as is done in the United Kingdom and elsewhere [19]. One advantage of using buffy coat collections is the ready availability of granulocyte products compared to apheresis collections. A dose of 10 buffy coats for adults

and 10-20 mL/kg for children less than 50 kg is recommended. Pooled granulocyte components prepared from whole blood containing approximately 1 × 1010 neutrophils are available for transfusion [20,21]; a low-volume pooled product has been shown to be functional in vitro and may represent a promising alternative [22]. Granulocyte concentrates contain variable amounts of red blood cells, platelets, and other white cells such as lymphocytes; the volume and cellular content of the product depends on the method of procurement (Table 1). After collection, granulocytes are irradiated to prevent transfusion-associated graft versus host disease [23], and stored at room temperature for a maximum of 24 hours to preserve adequate function [24]. Because of this short shelf life, the results of donor infectious disease testing may not be completed prior to transfusion. Granulocyte concentrates contain up to 50 mL of red blood cells (RBC), therefore the product should be cross match compatible with the recipient to prevent hemolytic transfusion reactions. If a major ABO compatible donor is not available, additional RBC sedimentation may be performed on apheresis granulocyte concentrates to deplete incompatible RBCs (Fig. 1) [25]. Clinical indications GTX is most commonly considered in the context of severe neutropenia due to disease or therapy such as in hematologic malignancies, and aplastic anemia; less commonly, GTX may be used in patients with neutrophil dysfunction, such as in chronic granulomatous disease (CGD). Recommendations generally suggest initiating GTX only where endogenous neutrophil recovery is expected, to avoid embarking on futile, unsustainable courses of therapy.

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Although newer antimicrobial agents have improved the outcome of patients with chemotherapy-induced neutropenia, and prophylactic myelopoietic growth factors reduce the risk of febrile neutropenia and infections in adult patients receiving chemotherapy [26], infections in neutropenic patients are still associated with hospital admission, organ damage, and mortality. GTX may represent a potential therapeutic adjunct, but many practitioners do not routinely consider this option. In a recent report of 471 patients receiving myeloablative chemotherapy, only 9 of 34 (26%) patients deemed eligible to receive GTX were treated with this therapy [27]. A survey conducted in England and North Wales reported little consensus among hematology units on the use of GTX, even in clinical scenarios where patients clearly met national guideline criteria. The most commonly cited reason for reluctance to use GTX was lack of evidence of effect [28]. Practice guidelines for granulocyte collection and administration vary amongst different centers, and indeed across countries. In the USA, the Food and Drug Administration (FDA) does not recognize granulocyte concentrates as a licensed blood component. However, the Circular of Information (COI) for the use of Human Blood and Blood Components outlines indications for which granulocytes are typically used; in patients with documented infections unresponsive to antimicrobial therapy in the setting of neutropenia [absolute neutrophil count (ANC) <0.5 × 109 /L (500/μL)] and expected eventual marrow recovery [29]. According to a recent international survey, national guidance on the indications for therapeutic use of granulocytes is available in France, Germany, and the UK [30]. In Canada, Australia and Brazil, there are standards or guidelines for collection and preparation of granulocyte concentrates, but no official national recommendations on the clinical use of GTX. In Belgium, the Superior Health Council considers granulocyte transfusions an experimental treatment in neutropenic patients with refractory severe bacterial or fungal infections. Professional evidence-based guidelines from Infectious Disease Societies with bacterial or fungal agents do not provide explicit recommendations regarding the use of GTX. However, some clinicians consider granulocytes as an adjunct in the treatment of microbiologically confirmed infections, as well as in the event of life-threatening suspected or “possible” invasive fungal infections, where patients with prolonged neutropenia demonstrate typical clinical and radiologic findings, but the culpable organism is not demonstrable. [31,74] GTX is often administered daily or on alternate days, although clinically measurable WBC increments have been reported using high-dose granulocyte concentrates 2 or 3 times per week [30]. The course of GTX is discontinued in the event of neutrophil recovery or clinical resolution of infection; courses may last several weeks. Granulocytes should be infused through a standard blood administration filter over 1 to 2 hours; for obvious reasons, leukoreduction filters must never be used with granulocytes. Outcomes: evidence of clinical efficacy Previous reviews have summarized the supporting evidence for the use of GTX for the prevention or treatment of infections in general [32-35]. The existing literature is predominantly comprised of case reports and uncontrolled case series, with limitations including publication bias. Interpretation of the literature is challenging because authors report heterogeneous patient populations, use varying doses of granulocytes, and have inconsistent criteria for determining successful outcomes. GTX has been associated with observable increments in absolute neutrophil counts, particularly in patients receiving >1 × 1010 transfused granulocytes; the degree of increment has been associated with survival in some series [36]. Defervescence and resolu-

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tion of signs of infection have been attributed to GTX therapy in multiple reports [37]. Transfused granulocytes have been shown to migrate into bacterially infected solid tissues [38,39] and to joint cavities inflamed by fungal infection [40]. The question of whether or not GTX improves survival in patients with infection is the unresolved issue; thus far, there is little evidence from controlled trials that granulocyte transfusions improve the outcome of current standard treatment of infections during neutropenia. Case series and controlled trials of GTX in adult patients Early controlled trials of GTX were conducted 30-40 years ago in prophylactic [41-49] and therapeutic settings [50-57], primarily in patients with acute myeloid leukemia undergoing induction chemotherapy or bone marrow transplantation. Granulocytes for transfusion were derived from unstimulated donors and hence of low dose. Results of these studies ranged from clear to marginal to no benefit, therefore GTX fell out of favor. However, in the early 1990s, the availability of recombinant G-CSF facilitated marrow stimulation of healthy donors, and led to renewed interest in GTX. Retrospective case series and prospective studies have since been conducted in adults with overall inconclusive results. The context in which GTX is used in these studies is not uniform. Generally, patients received therapeutic GTX for the treatment of established refractory infections; some series also included patients with febrile neutropenia without microbiologically documented infection. Prophylactic or pre-emptive, GTX has been used in patients considered at risk of infection (primary prophylaxis), or to prevent recurrent or worsening infection during a period of transient neutropenia, often just prior to or during HSCT (secondary prophylaxis). In some series, GTX is used in more than one of the approaches described above, which further confounds the clarity of the results. Regarding the response of different types of infectious agents to GTX, many authors reported that patients with bacterial infections had better outcomes than those with fungal infections (Table 2); interestingly, the reverse has also been reported. In a phase I/II trial using G-CSF and dexamethasone-mobilized GTX, the authors report resolution of infection in 8/19 patients (42%); none of the patients with invasive aspergillosis cleared the infection [58]. Conversely, in another prospective study, favorable responses were observed in 40% of patients, especially in those with fungal or Gramnegative infections [39]. A single-center report of 11 patients with invasive fusariosis treated with high-dose GTX reported 91% 30-day survival [59], compared to clinical response in only 30% in prior published cases of Fusarium infection treated with GTX. Some series claimed successful prophylactic GTX in patients with recent fungal infections undergoing allogeneic HSCT [60,61]; other authors reported no benefit of prophylactic GTX [62]. GTX has also been reported to stabilize or improve fungal infection [63], and to mitigate the high risk of mortality in adults [64-66] and children [67] with IFI undergoing HSCT. Retrospective analyses have reported mixed results regarding survival benefit afforded by GTX, ranging from clinical benefit compared with historical controls [68], no difference in mortality [69] or even increased likelihood of death [70] in patients with fungal infections who received GTX compared to patients who did not. In a retrospective review of patients with aplastic anemia treated with GTX, survival was strongly correlated with hematopoietic recovery; 93% of the patients who recovered hematopoiesis survived; of the 18 patients who did not, only 22% survived to hospital discharge [66]. A prospective study evaluating the effect of GTX therapy on survival and microbial response compared to concurrent or historic controls showed that the number of fatal or progressive fungal infections and survival was comparable in both groups [71].

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Table 2 Case series of adult patients treated with GTX in the G-CSF era. Study

N

Study design

Response (Fungal)

Response (Bacterial)

Grigg (1996) [60] Price (20 0 0) [58] Lee (2001) [39] Illerhaus (2002) [62] Hubel (2002) [71] Rutella (2003) [68] Mousset (2005) [61] Safdar (2006) [104] Ofran (2007) [105] Quillen (2009) [66] Al-Tanbal (2010) [106] Ang (2011) [107] Kim (2011) [91] Safdar (2014) [108] Wang (2014) [109] Marciano (2017) [88]

11 19 25 18 74 22 44 20 47 32 22 15 128 74 56 40

Retrospective Prospective Prospective Retrospective Retrospective Retrospective Prospective Retrospective Retrospective Retrospective Retrospective Retrospective Retrospective Retrospective Retrospective Retrospective

0% resolved 0% with IA cleared infection 73% response 55% IPA responded 18% mould, 55% yeast stable 57% (0% response in IFI) 78% response at 30 days 45% CR or PR, 15% stable 64% infection-related survival 44% survival 75% survival 31% cleared 47% control of IFI 45% patients had IFI 87% 30-day survival 82% success

100% resolved 100% resolution 45% response 78% septicaemia responded NS 54% response 92% response at 30 days NS 53% infection-related survival 58% overall survival to discharge 68% clinical improvement 63% cleared 53% overall control of infection 46% overall response 92% 30-day survival 94% success

CR = complete response; IFI = invasive fungal infection; IA = invasive aspergillosis; IPA = invasive pulmonary aspergillosis; PR = partial response; NS = not specified.

In a recent Cochrane database systematic review [72], the authors concluded that in patients who are neutropenic due to chemotherapy or HSCT, there is low-grade evidence that prophylactic granulocyte transfusions decrease the risk of bacteremia or fungemia. This effect of prophylactic granulocyte transfusions may be dose-dependent, with a dose of at least 1 × 1010 per day being more effective. There was insufficient evidence to determine any difference in mortality rates due to infection. In light of the risk of serious adverse events, primary prophylactic GTX is no longer recommended. Recent controlled trials have thus far failed to confirm or refute the benefit of therapeutic GTX. Seidel et al [73] conducted a phase III RCT of GTX in 74 patients with hematologic or malignant diseases. The authors found no difference in probability of 28-day survival, and no effect of GTX on survival until day 100 in patients with fungal or bacterial infection. Price et al reported the results of the RING (Resolving Infection in Neutropenia with Granulocytes) study [74], the largest randomized controlled trial of granulocyte transfusions, comprised 114 patients with neutrophil count <0.5 × 109 /L and proven or probable bacterial or fungal infection. The primary endpoint of this study was a composite of survival plus microbial response at 42 days. Differences in primary endpoint success rates for granulocyte and control arms were not statistically significantly different for any infection type. The granulocyte dose was also lower than anticipated; the target of ≥4.0 × 1010 neutrophils per transfusion was only achieved in 70% of subjects. In a post-hoc analysis, subjects who received an average dose per transfusion of ≥ 0.6 × 109 granulocytes/kg tended to have better outcomes than those receiving a lower dose. Both of these trials were underpowered due to low enrollment. A systematic Cochrane review of therapeutic granulocyte transfusion incorporated the results of the RING trial, as well as other older studies [75]. There were no clear differences in other outcomes and there was insufficient data to report on risks of harm, such as serious pulmonary adverse events. Overall, it was not possible to establish whether granulocyte transfusions affect all-cause mortality. Case series of GTX in pediatric patients Infections in neutropenic children are associated with a severe prognosis, despite treatment with appropriate antimicrobial therapy, with up to 85% mortality in bone marrow transplant recipients with invasive aspergillosis. A number of series of pediatric patients with high-risk febrile neutropenia, proven or suspected serious infections have been published (Table 3). GTX was determined to be

beneficial in retrospective analyses [76-79] and prospective uncontrolled studies [80]. In an open, single-center, prospective phase II clinical trial of early-onset G-CSF-mobilized GTX in neutropenic children with severe refractory infections [81], GTX was well tolerated, with 93% clearance of initial infection and no pulmonary transfusion reactions. Because of their small size, pediatric patients may obtain a superior increment in ANC post-GTX compared to adults; however one prospective study showed that neither body weight nor granulocyte dose impacted infection outcome and survival in pediatric patients [82]. More modest responses were reported in other series, with promising short-term survival but the majority of patients ultimately dying of their infection [83]. Some studies suggest a role for granulocyte transfusions in preventing infections or progression of infections in children with anticipated prolonged neutropenia after HSCT or chemotherapy [84,85]. Case reports and small series of children and young adults with CGD report benefit from the addition of GTX to the therapeutic arsenal [86,87]. The largest cohort of CGD patients with refractory infections treated with of GTX as an adjunctive therapy reported that overall >80% infections improved with this approach [88]. To date, no randomized controlled trials of GTX have been conducted in children. A Cochrane review concluded that there is inconclusive evidence from randomized controlled trials (RCTs) to support or refute the routine use of granulocyte transfusions in neutropenic, septic neonates [89]. Outcomes: adverse effects of granulocyte transfusion Febrile and allergic reactions are commonly associated with GTX, occurring at a rate of 10%-15%. Febrile transfusion reactions are attributed to recipient antibody reactions against HLA antigens on donor leukocytes in the transfused product, or infusion of inflammatory cytokines produced during room temperature storage. These reactions may be associated with chills and rigors, minor fluctuations in blood pressure and/or pulse rate; symptomatic management with antipyretics will usually suffice. GTX is associated with increased risk of transmission of intracellular pathogens such as CMV [90]. Since granulocyte recipients often require CMV-safe blood products, CMV seronegative donors are generally recommended for seronegative recipients. Pulmonary complications, including hypoxemia, hemoptysis, pulmonary infiltrates, acute lung injury and fluid overload have been reported in 10%-18% of patients [91,92]. In an early report, 14 of 22 (64%) patients receiving amphotericin B during GTX developed respiratory deterioration, compared to 2 of 35 (6%) who did not receive amphotericin [93]; however this association has not

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Table 3 Case series including pediatric patients treated with GTX in the G-CSF era. Study

N

Study design

Response (Fungal)

Response (Bacterial)

Grigull (2006) [84] Kikuta (2006) [80] Sachs (2006) [81] Drewniak (2008) [76] Seidel (2009) [82] Graham (2009) [83] Atay (2011) [78] Ozturkmen (2013) [77] Diaz (2014) [79] Nikolajeva (2015) [110]

32 13 27 18 69 13 35 10 18 28

Retrospective Prospective Prospective Prospective Prospective Retrospective Retrospective Retrospective Retrospective Retrospective

67% survival 50% response 100% response 73% response 28-day survival probability 0.51 ± 0.12 50% survived to discharge 55% clinical response 50% response 80% response 79% 100-day survival

81% survival 73% response NS 100% response 28-day survival probability 0.89 ± 0.06 100% survived to discharge 65% clinical response 80% response 100% response 50% 100-day survival

Practice considerations. When considering a course of GTX for a patient, determine: Is GTX indicated? • Does my patient meet the generally agreed upon criteria referred to below, for patients most likely to clinically benefit? o Bacterial or fungal infections unresponsive to antimicrobial therapy o Profound neutropenia [absolute neutrophil count < 0.5 × 109 /L (500/μL)] o Expected eventual marrow recovery Is GTX feasible? • Can my hospital blood bank or transfusion service acquire granulocyte concentrates? • Can enough doses be provided to support daily or other chosen frequency of infusions? Is GTX safe? • Do the potential benefits outweigh the risk of harm (e.g. severe pulmonary reactions)?

since been proven or reproduced. In the RING trial, severe reactions (including hypoxia requiring temporary ventilatory support) were observed in <5% of granulocyte transfusions [74]. GTX can cause alloimmunization to HLA and human neutrophil antigens (HNA), resulting in subsequent platelet and leukocyte transfusion refractoriness, or rejection of a subsequent allogeneic HSCT [94]. Red cell alloimmunization has also been reported [95]. Patients with pre-existing granulocyte-reactive (HLA or HNA) antibodies have been associated with more frequent febrile and pulmonary reactions; diminishing increment in the ANC [96], and decreased localization of transfused granulocytes to sites of inflammation, leading to impaired efficacy of GTX [97]. Interestingly, the RING study group reported no statistically significant difference in WBC alloimmunization in the GTX arm, and no demonstrable effect of the presence of alloimmunization on survival and microbial response at 42 days, the occurrence of transfusion reactions, or neutrophil increments [98]. Nevertheless, many remain convinced that in light of the clinical and laboratory evidence to date, the presence of leukocyte-reactive antibodies should be taken seriously [99].

Persistent concerns and future considerations After years of published reports and extensive discussion, controversy persists. Many clinicians remain disinclined to use GTX because high-dose, G-CSF-mobilized granulocyte concentrates are not available at their institutions; low-dose granulocyte concentrates may pose risks to the recipient without likely clinical benefit. Obstacles to conducting future randomized controlled trials include cost, logistical challenges, and low enrollment. A prospective international registry of granulocyte transfusions has been initiated by the BEST Collaborative Group, which may capture helpful data on granulocyte transfusion practice as well as clinical efficacy [100]. Current limitations of donor-derived granulocyte concentrates include limited supply, short shelf-life, and cumbersome collection processes, hindering future clinical trials and the use of GTX in common practice. Functionally mature neutrophils have been generated at a small scale from induced pluripotent stem cells (iPS) [101,102] or CD34+ hematopoietic progenitor cells; in the future, granulocyte concentrates may be produced in advance, cryopreserved and made available as an off-the-shelf product [103].

Summary and conclusions GTX has remained in clinical use for over 40 years, despite persistent controversy regarding its efficacy. Severely neutropenic patients with refractory bacterial or fungal infections tend to be very ill with high risk of mortality, therefore clinical response to GTX might be difficult to determine in this setting. Overall, the quality of the data in the published literature is low, and predominantly limited to individual cases and uncontrolled case series. GTX may have a role in preventing progression of refractory fungal infections during HSCT-induced neutropenia. Modern controlled studies, including the RING trial, have been unable to demonstrate clinical benefit of GTX; however, this study was underpowered due to under-enrollment, and many patients received suboptimal doses. Transfused granulocytes may cause adverse reactions, including severe pulmonary reactions, HLA alloimmunization, and CMV infection. The risks and benefits must be weighed on a case-by-case basis. GTX may be helpful in certain patients if rapidly available at high neutrophil doses, particularly if neutrophil recovery is anticipated. Future studies should ideally evaluate the effects of highdose GTX using clearly defined parameters of response to therapy. Author contributions KW and CC wrote and critically reviewed the manuscript. Declaration of competing interest The authors have no relevant conflicts of interest to disclose. The views expressed are the authors’ own and do not represent the National Institutes of Health, the Department of Health and Human Services, or the U.S. Federal government. Acknowledgment The authors thank Dr Harvey Klein for reviewing the draft manuscript. This work was supported by the Intramural Research Program of the NIH Clinical Center (project Z99 CL999999). References [1] Mayadas TN, Cullere X, Lowell CA. The multifaceted functions of neutrophils. Annu Rev Pathol 2014;9:181–218.

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