Antifungal prophylaxis of patients undergoing allogenetic hematopoietic stem cell transplantation in China: a multicenter prospective observational study

Background Antifungal prophylaxis is currently regarded as the gold standard in situations with allo-genetic hematopoietic stem cell transplantation (allo-HSCT). However, the epidemiological information regarding prophylaxis of invasive fungal diseases (IFDs) is not clear in China. Methods We report the first large-scale (1053 patients) observational study of the prophylaxis and management of IFDs among patients with allo-HSCT in China. Results The incidence rates of IFD after primary antifungal prophylaxis (PAP), secondary antifungal prophylaxis (SAP), and non-prophylaxis were 22.7 vs. 38.6 vs. 68.6 %, respectively (P = 0.0000). The median time from transplantation to IFD was 45 days in PAP patients, 18 days in SAP patients, and 12 days in non-prophylaxis patients. Aspergillus spp. represents the most common type of fungal infection. Independent risk factors for IFD in allo-HSCT patients with PAP were age, having human leukocyte antigen (HLA)-haploidentical or matched unrelated donor, decreased albumin levels, and the use of itraconazole as the prophylactic antifungal agent. Among SAP transplant recipients, there was no significant risk factor for IFDs. The incidence rates of overall survival (OS) in the PAP, SAP, and no prophylaxis groups were 85.07, 78.80, and 74.82, respectively (P = 0.01). Conclusions This observational study indicates that prophylaxis of IFD is helpful to reduce the incidence of IFDs and improve the OS of patients after allo-HSCT.

This first large-scale observational study of invasive fungal disease (IFD) in China indicated that prophylaxis of IFD is helpful to improve the OS of patients after allogenetic hematopoietic stem cell transplantation (allo-HSCT). For patients with a history of IFD, effective prevention of IFD is particularly important because they have higher incidence of IFD.

Background
Although the control of bacterial infections in patients with hematological malignancies has been significantly improved with broad-spectrum antibiotics in the past decades, treating invasive fungal diseases (IFDs) is still a major problem in these patients, especially in patients undergoing allogenetic hematopoietic stem cell transplantation (allo-HSCT) [1][2][3]. Epidemiological data from the USA, China, and parts of Europe have shown that the incidence of IFDs in patients after allo-HSCT has increased dramatically in the past several years, substantially increasing the morbidity and mortality rates [4][5][6][7][8][9].
The diagnosis and treatment guidelines for IFDs, including some that are specific to HSCT patients, have been developed by academic societies in different world regions [10][11][12]. Notwithstanding the guidance and new forms of antifungal agents, treatment is often delayed because of nonspecific disease presentation and a lack of reliable diagnostic techniques, leading to poor clinical outcomes. Prophylaxis of IFDs is currently regarded as the gold standard in situations with allo-HSCT. Over the years, different scientific societies have established a series of recommendations for antifungal prophylaxis based on prospective studies performed with different drugs [13][14][15][16][17].
However, until recently, data on the prophylaxis of IFDs and real-world management of IFDs in patients with allo-HSCT have been gleaned primarily from single-center and retrospective studies in China. Here, we report the first large-scale observational study of the prophylaxis and management of IFDs among adults and children who have undergone allo-HSCT in China. Data came from the China Assessment of Antifungal Therapy in Hematological Disease (CAESAR) study.

Study design
The CAESAR study was a multicenter, prospective, observational study performed in 35 hematology centers across China, including two children's hospitals. Subjects were consecutive patients of any age with a hematological malignancy who were hospitalized during the study period either after allogeneic or autologous HSCT or to receive intravenous chemotherapy. The overall study methods have been described previously [7,18]. This observational study was conducted as a part of the CAESAR study and focused on the prophylaxis of IFDs in 1053 patients who underwent allo-HSCT in 31 HSCT centers.
All patients in each study center who were hospitalized and had undergone allo-HSCT between January 1, 2011 and October 30, 2011 were enrolled in this study. Data were collected by chart reviews and included the following parameters: demographic characteristics, antecedent hematologic disease, type of HSCT, conditioning regimen, IFD risk factors, clinical features suggestive of IFD, laboratory findings such as imaging, histopathology, and fungal cultures, treatment outcome of IFD, and mortality. Each IFD was categorized as proven, probable, or possible, according to the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) 2008 criteria [19]. Patients were diagnosed as having suspected IFD if they had IFD risk factors; were observed to have symptoms, radiological abnormalities, or indirect microbiological evidence of fungal infection; and were treated empirically with antifungal agents but could not be diagnosed with proven, probable, or possible IFD according to the EORTC/MSG 2008 criteria [20]. Patients were followed for 6 months after the date of transplantation; the follow-up was completed on April 30, 2012. In all, 18 patients were lost to follow-up.

Statistical analysis
In accordance with common practice and to maximize diagnostic accuracy, the incidence of IFD was mainly calculated based on proven and probable cases combined. The cumulative incidence was calculated as the incidence of proven plus probable IFD for the first 187 days after transplantation, divided by the number of cases at risk. Data were grouped according to primary antifungal prophylaxis (PAP) or secondary antifungal prophylaxis (SAP). PAP refers to patients without a history of fungal infection and need to be protected to avoid fungal infection after HSCT [21]. SAP is a rational strategy for patients with proven or probable IFD within 6 months before transplantation [22]. In particular, all allo-HSCT patients (n = 1045) in the CAESAR study were included in the analysis of overall survival (OS) for comparison of the OS differences in the PAP, SAP, and no prophylaxis groups. The characteristics of all of the allo-HSCT patients were described in the CAESAR study [7].
Statistics were primarily descriptive and were compared using analysis of variance, the Wilcoxon rank-sum test, or the chi-squared test, as appropriate. Risk factors for IFD were analyzed using univariate analysis followed by multivariate logistic regression. Kaplan-Meier analysis and the log-rank test were used to compare OS between different groups of patients. A two-sided P value of less than 0.05 was denoted as statistically significant. All statistical analyses were performed with SAS software version 9.1 (SAS, Cary, NC, USA).

Patient characteristics and antifungal prophylaxis regimen
A total of 1053 patients who underwent allo-HSCT were enrolled from 31 HSCT centers across China. Their baseline demographic and clinical characteristics at the time of admission for transplantation are shown in Table 1. Among them, 906 patients received prophylactic antifungal treatment before or after transplantation. Previous IFDs were noted in 88 patients (8.4 %, 88/1053), including 7 with proven IFDs and 81 with probable IFDs.
The average duration of antifungal prophylaxis before HSCT was 10.9 ± 6.56 days (10.8 ± 6.57 days in the PAP group and 12.0 ± 6.43 days in the SAP group). Single antifungal agents were the most common prophylaxis regimen (667/818 in the PAP group and 69/88 in the SAP group). Fluconazole (429/818, 64.9 %) and voriconazole (32/88, 46.4 %) were the most widely used agents for PAP and SAP, respectively ( Table 1). All antifungal agents were given in accordance with the recommended doses and schedule.

Efficacy of PAP and SAP on the prophylaxis of IFD occurrence
In    Table 3. Of the 89 etiological pathogens identified in patients with proven or probable IFDs, 62 (69.7 %) were Aspergillus, including 19 that were exclusively diagnosed with two positive galactomannan tests. Unspecified Aspergillus was the most common mold (31 cases), followed by Aspergillus flavus (5 cases), and Aspergillus fumigatus (2 case) isolates.
The infection sites of all categories of IFD were also analyzed. Of all the 54 infection sites identified in patients with IFDs, the most common location of infection was the lower respiratory tract (64.8 %, 35/54), followed by blood stream infections only (13.0 %, 7/ 54), central nervous system (5.6 %, 3/54), spleen (1.9 %, 1/54), and three other sites ( Table 4). Among the SAP transplant recipients, there was no significant risk factor for proven or probable IFD.

Discussion
Invasive infections remain major infectious threats to patients undergoing allo-HSCT and are associated with high fatality rates [23][24][25][26][27][28]. The diagnosis of IFDs continues to be difficult to establish because they do not manifest with specific clinical or radiographic signs or symptoms [29]. In the recent years, there has been a concerted effort to identify alternative procedures for the future diagnosis of fungus. They include the targeting of fungal antigens by enzyme-linked immunosorbent assay (ELISA) or lateral flow devices (LFDs) [30], detection of siderophores [31], and amplification of fungal nucleic acids from tissue and body fluids [32] as well as application of matrix-assisted laser desorption ionization TOF mass spectrometry (MALDI-TOF MS) [33]. Each of the above methods has advantages and disadvantages, and not all of which have entered clinical practice [34]. The difficulty in obtaining a timely diagnosis as well as the high morbidity and mortality rates associated with IFDs provide a rationale for antifungal prophylaxis in patients undergoing allo-HSCT. Nonetheless, antifungal prophylaxis remains a topic of some controversy, with no clear consensus among different centers [35][36][37].
The CAESAR study is the first population-based and the largest prospective observational study of the incidences of IFDs in patients receiving HSCT [7]. This Fig. 1 Overall survival of patients with proven/probable IFD, possible IFD, or suspected IFD and who received antifungal therapy; those with no suspected IFD and who did not receive antifungal therapy; and those without any IFD (proven, probable, or possible) observational study was conducted as a part of the CAE-SAR study and focused on the prophylaxis of IFDs in patients who underwent allo-HSCT. In the present analysis of adults and children at risk for IFD due to allo-HSCT, non-prophylaxis transplant recipients had a higher incidence of proven/probable/possible IFD than PAP and SAP transplant recipients at 6 months (68.6 vs. 22.7 and 38.6 %, P = 0.0000). This result was in accord with the classic randomized clinical trials on PAP in HSCT recipients and patients with hematological malignancies [38][39][40]. Furthermore, the OS was significantly different among PAP, SAP, and no prophylaxis patients. It was revealed that antifungal prophylaxis was helpful to reduce the incidence of IFD and improve the survival of patients after transplantation.
The most recently published European guidelines pointed out that SAP should be administered to patients with a previous IFD to prevent recurrence of the previous IFD or onset of a new IFD during a new at-risk phase, mainly referring to a prolonged neutropenic period induced by chemotherapy or a phase of severe immunosuppression after allo-HSCT [16]. Several studies have reported success rates for SAP, which was proven to be effective in preventing IFD recurrence [41,42]. In the study, the cumulative incidence of IFD increased particularly rapidly during the first month after transplantation in the SAP group and non-prophylaxis group, suggesting that this is a high-risk period during which health care providers should pay particularly close attention to signs of emerging IFD in SAP and nonprophylaxis patients. On the contrary, the median time of IFD occurrence in the PAP group was 45 days after HSCT. This finding indicated that patients who had a previous IFD and had no antifungal prophylaxis were more likely to experience a breakthrough fungal infection in the early stage after transplantation.
As 22.7 % of patients treated with PAP and 38.6 % of patients treated with SAP in the study went on to develop proven, probable, or possible IFD, there remains an unmet need for education about the appropriate timing and choice of antifungal agent for prophylaxis in China. Among PAP transplant recipients, the most commonly prescribed prophylactic agents were fluconazole and itraconazole, which may be less effective than posaconazole [43]. The use of itraconazole as the antifungal prophylactic agent also proved the independent risk factor for IFD occurrence in our study. An update to the cost-effectiveness of posaconazole vs. fluconazole or itraconazole in the prevention of IFD among neutropenic patients in the USA has shown that posaconazole is a cost-effective alternative to fluconazole or itraconazole in the prevention of IFD among neutropenic patients [44]. Among SAP transplant recipients, there was no significant risk factor for proven or probable IFD. The characteristics of the observational study and the small sample size of the patients treated with SAP may be the main reasons. Prospective and randomized studies assessing the risk factors for SAP are needed in the future.
The epidemiological characteristics of IFD continue to evolve in transplant patients. A major contributor is the widespread use of azole prophylaxis since the early 1990s, which results in less candidiasis but more frequent mold infections in hematologic malignancies [25,45]. In most [46][47][48] but not all [49] cases, Aspergillus spp. represent the most frequent cause of fungal-related morbidity in patients with HSCT. In our study, Aspergillus spp. also predominated in culture-proven or histologically proven pathogens and was more than two times as common as Candida spp. The mortality among patients who developed IFD (24.1 %) was almost double than that in the overall population of HSCT patients (15.0 %), and regression analysis confirmed that the development of IFD is a significant independent risk factor for death, highlighting the grave prognosis of those with IFD and the need for a timely diagnosis and prompt treatment. The incidence rates of invasive aspergillosis, however, need to be interpreted with caution as establishing this diagnosis often requires invasive procedures that are difficult to perform in severely ill patients. The low rate of proven/probable IFD in the present study suggests the need to improve diagnostic techniques to treat IFD as early and accurately as possible. Under the existing conditions, in order to reduce the IFDrelated mortality of allo-HSCT patients, preemptive antifungal therapy should be given to patients with possible or suspected IFD.
The current study has several limitations. The main limitation was its observational nature. Confounding factors cannot be controlled effectively in observational studies, frequently due to biased selection of patients or Fig. 2 Overall survival of allo-HSCT patients receiving PAP, SAP, or no prophylaxis treatment protocols. In the analysis of mortality by antifungal therapy, the uncontrolled study design means that no firm conclusions can be drawn regarding the relative effect of different treatments. Another limitation was the relatively short duration. The fact that the overall treatment duration was not recorded in patients who were still taking antifungal treatment after discharge from the hospital further limited data analysis. A longer follow-up would have allowed a more comprehensive analysis of patients who developed IFD after discharge. Procedures for diagnosis, prophylaxis, and treatment of IFD were not prespecified in the protocol but were performed according to usual practice and local clinical guidelines [19]. Furthermore, in some centers, diagnostic procedures were conducted only among patients with suspected clinical signs or symptoms of IFD. The observational study also meant that the diagnosis of specific infections, although guided by EORTC/MSG 2008 criteria, was limited to the data collected according to local hospitals' usual practice; there was no mandatory requirement for diagnostic microbiological testing or the use of a centralized laboratory to validate the results.

Conclusions
In conclusion, the results of the present observational study indicate that prophylaxis of IFD among patients receiving allo-HSCT for hematological malignancy in China is broadly in line with the recommended practice and is helpful to reduce the incidence of IFD and improve the OS of patients after transplantation. For patients with a history of IFD, active and effective prevention of fungal infections is particularly important because these patients have a higher incidence of IFD occurrence and a shorter incubation period. Different from the traditional view, for the allo-HSCT patients with PAP, itraconazole was a less effective treatment than other antifungal drugs. Aspergillus spp. represents the most common type of fungal infection in patients with allo-HSCT. Due to the limited diagnostic techniques, it is necessary to give antifungal therapy to patients who do not meet the EORTC/MSG 2008 criteria but show clinical evidence of fungal infection.