Fluconazole plus flucytosine is a good alternative therapy for non‐HIV and non‐transplant‐associated cryptococcal meningitis: A retrospective cohort study

Summary Cryptococcal meningitis (CM) carries a high risk of mortality with increasing incidences in immune competent hosts. Current treatments are not well tolerated, and evaluation of other treatments is needed. Fluconazole and 5‐flucytosine in treating immune competent hosts have not been characterised. To evaluate the efficacy of fluconazole and 5‐flucytosine in treating non‐HIV‐ and non‐transplant‐associated CM. We performed a retrospective cohort study of the outcomes in immune competent patients with CM treated with fluconazole and 5‐flucytosine or deoxycholate‐amphotericin B and 5‐flucytosine. The primary outcome was treatment response evaluated at the 12th week after initiation of antifungal therapy. A total of 43 and 47 patients received amphotericin B deoxycholate and 5‐flucytosine or fluconazole and 5‐flucytosine, respectively. A total of 38 (88.4%) patients cannot tolerate recommended doses of amphotericin B deoxycholate and 5‐flucytosine (patients needed dose reduction during the treatment). Patients given fluconazole and 5‐flucytosine had higher baseline cryptococcal burdens (median 3632 versus 900 cryptococci/mL, P = 0.008). No significant differences were seen in cryptococcus clearance (74.4% vs 70.2%, P = 0.814), treatment time (39 days, 20‐69 days vs 21 days, 7‐63 days, P = 0.107) and successful response (including complete and partial responses) rates (69.7% vs 72.3%, P = 0.820). Fluconazole and 5‐flucytosine treatment had lower total adverse events (19.1% vs 90.7%, P < 0.001). Fluconazole and 5‐flucytosine had relatively high efficacy with few adverse events in treating CM. Fluconazole and 5‐flucytosine therapy is promising in patients that do not tolerate or are not suited for amphotericin B deoxycholate treatment.


| INTRODUC TI ON
Cryptococcus neoformans (C. neoformans s.l.) and Cryptococcus gattii (C. gattii s.l.) species complex are opportunistic pathogens, and cryptococcal meningitis (CM) is the most common cause of fungal meningitis by a considerable high morbidity and mortality. It was estimated the global incidence of CM to be substantial at 223 100 cases annually, resulting in 181 100 annual deaths in 2014. 1 This infection is common in immunosuppressed patients including those with HIV or who have undergone solid organ transplantation. [2][3][4] There has been an increase in the incidence of this disease among immunocompetent hosts that are not HIV-infected or not transplant recipients in recent years. [5][6][7] Reports have noted that about half of all reported cases of cryptococcosis are in non-HIV-infected and non-transplant patients (between 44% and 55%). [8][9][10][11] Additionally, CM is associated with excessively high disability and mortality rates. 12,13 Induction therapy is a critical treatment for CM in non-HIV-infected and non-transplant CM patients.
Induction therapy of amphotericin B (AmB) (0.7-1.0 mg/kg per day) combined with 5-flucytosine (5-FC) (100 mg/kg per day) for at least four weeks is the globally preferred regimen for non-HIV-infected and non-transplant patients with CM worldwide. 14 However, high doses of AmB and 5-FC can result in severe toxic side effects such as phlebitis, liver impairment, renal impairment, haematological impairment, myocardial damage and a majority of CM patients do not tolerate the recommended dosages. [15][16][17][18][19][20] The triazole antifungal drug fluconazole (FCZ) has been widely used in treating deep mycosis and has low rates of adverse events. FCZ combined with 5-FC was recommended to treat HIV-associated CM. 14 However, FCZ combined with 5-FC in treating non-HIV-and non-transplant-associated CM has not been well characterised and thus the data are limited. Thus, in this study we compared the efficacy of combination therapies using AmB and 5-FC versus FCZ and 5-FC. Our goals were to analyse this drug combination therapy in order to propose alternative therapeutic regimens for CM in patients without HIV or patients that are not transplant recipients.

This study was approved by the Ethics Committees of The Third
Affiliated Hospital of Sun Yat-Sen University. The study was in compliance with the Declaration of Helsinki and its later amendments. All study participants provided informed consent. Identifiable data involving the individuals in this study were encrypted.

| Laboratory examination
Enrolled patients underwent lumbar punctures (LP) at least once a week in accordance with treatment guidelines. 14,22 Then CSF open pressure, CSF white blood cell count and classification, glucose, protein, India ink stain and cultures were recorded. In addition, other conventional blood tests and imaging studies including brain computerised tomography (CT) and/or magnetic resonance imaging (MRI) were also performed during treatment. CSF burden of cryptococcal organisms was evaluated by CSF cryptococcal organisms count through India ink stain. We use canavanine-glycine-bromothymol (CGB) medium to distinguish C. gattii s.l. from C. neoformans s.l.
CGB medium showed clear colour change for C. gattii s.l. and can be used to differentiate it from C. neoformans s.l. 23

| Therapeutic methods
In group I, all patients were treated with amphotericin B deoxycho-

| Outcome assessments
The primary outcome was treatment response evaluated at the 12th

| Assessment of adverse events
Data of adverse events were collected and assessed by National Cancer Institute Common Terminology Criteria for Adverse Events version 5.0 (NCI CTCAE v5.0).

| Statistical analysis
Baseline demographic and clinical characteristics are presented as percentages, mean with standard deviations (SD) or medians with range; comparisons were performed using the chi-square or Fisher's exact tests for categorical data, and with Student's t or Mann-Whitney U tests for continuous data. Efficacy of treatment responses was estimated using the Wilcoxon rank sum test based on the five grades over ten weeks between the two study groups. Chi-square tests were used to compare CSF sterilisation within 2, 4 and 12 weeks. Chi-square and Fisher's exact tests were used to estimate the incidence of adverse events between groups. Statistical analyses were performed using SPSS statistics version 19 (IBM). All analyses were two-sided and Pvalues of < 0.05 were considered statistically significant.  Table 1. Patients in the FCZ and 5-FC group had higher median CSF burden of cryptococcal organisms before treatment compared to patients in the AmB and 5-FC group (median 3632 vs 900 Cryptococci/mL, P = 0.008). No significant differences were observed between groups with respect to age, gender, CSF opening pressure, CSF WBC, CSF protein and CSF glucose.

| CSF sterility
CSF sterility results within 12 weeks are detailed in Table 2, and no significant differences were observed in the incidence of CSF cryptococcus clearance (

| Treatment response
The primary treatment response outcome was evaluated in the 12th week following initial therapy. No significant difference was observed in the treatment response rate as shown in Table 3 AmB + 5-FC (n = 43)

| D ISCUSS I ON
We found that the majority of patients with non-HIV-and non-transplant-associated CM do not tolerate the recommended induction therapy doses of AmB or 5-FC. Our findings were in accordance with those of previous studies which depict medication intolerance as a commonly documented problem in China. 15,17,18 Although dosages of AmB and 5-FC were adjusted, the incidence of adverse events in this study was still high (90.7%). AmB activity is concentration-dependent, so it is vital to ensure that drug concentrations reach the level of therapeutic effect. 25  Our study was limited due in part to its retrospective nature and the fact that the total number of enrolled patients was limited.
Moreover, we did not further apply molecular methods to distinguish C. neoformans s.l. from C. gattii s.l. after CGB culture. As a result, a definite consensus regarding the efficacy of FCZ and 5-FC therapy for non-HIV-and non-transplant-associated CM was needed. Future studies, particularly additional prospective large-scaled randomised controlled trials including molecular methods to distinguish cryptococcal species, are warranted in order to definitively determine the best standard of care in these patient populations.
In conclusion, our study found that the majority of patients with non-HIV-and non-transplant-associated CM do not tolerate the recommended doses of AmB during induction therapy due to side effect severity. Adjusting the dosages and treatment regimen of AmB still resulted in a high incidence of adverse events in this group.
We showed that FCZ and 5-FC combination therapy had relatively high efficacy with few adverse events in treating non-HIV-and nontransplant-associated CM. FCZ and 5-FC combination treatment proved to be a good alternative therapy for patients with non-HIVand non-transplant-associated CM. This treatment regimen is particularly efficacious in patient groups that are unable to tolerate the recommended dosages of AmB or in those that are not suitable candidates for AmB treatment.

CO N FLI C T O F I NTE R E S T
There are no conflicts of interest to declare.