Itraconazole Research Articles

Repurposing Itraconazole As an Anti-Cancer Agent Against Diffuse Large B-Cell Lymphoma

Introduction: Results from the Retrospective Non-Hodgkin Lymphoma Research (SCHOLAR-1) study demonstrated that relapsed/refractory Diffuse Large B cell Lymphoma (r/r DLBCL) have a low response rate to third-line therapy (20-30%) and an overall survival of 6 months. It stresses the need to develop novel therapeutic strategies for patients with r/r lymphoma. Itraconazole (ITZ) is a common widely clinical used orally bioavailable azole antifungal drug, with a more than thirty years well-documented safety record. Recently, scientific evidence has highlighted ITZ anti-tumor effects at clinically achievable doses in patients with variety of solid tumor malignancies. However, little is known about ITZ activity in B-cell lymphoma, especially in chemotherapy resistant lymphoma. Here we are the first group to report that potent anti-tumor activity of ITZ as a single agent or combined with other chemotherapeutic drugs in both sensitive and resistant lymphoma pre-clinical models. We also further explored the mechanism of action of ITZ to overcome drug resistance.Methods: A panel of rituximab-chemotherapeutic therapy-sensitive (TSCL), -resistant (TRCL) cell lines, developed in our laboratory, and primary tumor cells isolated from B-cell lymphoma patients were exposed to escalating doses of ITZ (0-20uM). IC50 values were calculated using the GraphPad Prism6 software. Changes in cell viability, cell number and cellular ATP levels were evaluated using the Presto Blue, Trypan Blue and Titer Glo assay, respectively. Mitochondrial respiration rate indicated as oxygen consumption rate (OCR) were measured by an XFe96 flux analyzer. Mitochondrial membrane potential (MMP) were quantified by DiOC6 staining and flow cytometry. Subsequently, TSCL and TRCL cells were exposed to ITZ alone or in combination with various chemotherapy agents or small inhibitors and differences in cell viability was determined. Coefficient of synergy was calculated using the CalcuSyn software. HKII-VDAC complex disruption by ITZ was identified by co-immunoprecipitation and western blot. HKII knockdown was generated by using ON-TARGET plus SMART pool siRNA containing a mixture of 4 siRNAs designed to specifically target HKII. Standard 51Cr release assays were conducted to study the effects of ITZ on the anti-tumor activity of monoclonal antiCD20 antibodies against TSCL and TRCL cell lines. For in vivo studies, severe combined immunodeficiency (SCID) mice were inoculated via tail vein injection (iv) with Raji cells (day 0) and assigned to observation, rituximab (at 10mg/kg/dose), obinotuzumab (at 10mg/kg/dose) with or without ITZ. Anti-tumor activity was evaluated by functional imaging and length of survival.Results: We found that ITZ induced a dose-dependent cell death in TSCL, TRCL and primary patient lymphoma cells. Single agent of ITZ decreased cell growth, reduced ATP production, lower the mitochondrial respiration rate and MMP in both TSCL and TRCL. ITZ exhibited additive/synergistic effects when combined with chemotherapeutic drugs (dexamethasone and cisplatin), proteasome inhibitors (bortezomib, carfilzomib, or ixazomib), targeted inhibitors (ABT199 and A-1210477 [an MCL-1 inhibitor]). At the molecular level, we found that, ITZ disrupted HKII/VDAC complex formation. Silencing HKII in TRCL attenuated the killing effect of ITZ. More importantly, ITZ in combination with rituximab or obinutuzumab led to reduced tumor intensity by functional imaging than rituximab or obinotuzumab single agent in vivo.Conclusion: Our findings support the potential benefits for using ITZ as an anti-cancer drug are: 1) reduce costs, 2) clear defined toxicity profile, 3) availability, and 4) pre-clinical activity in rituximab-chemotherapy resistant lymphomas. ITZ can be an inexpensive and effective agent to enhance standard chemotherapy in B-cell lymphoma. Repurposing of ITZ can result in novel therapeutic options for lymphoma patients. (Research supported by Roswell Park Cancer Institute Alliance Grant, National Natural Science Foundation of China (NSFC) 81573063) DisclosuresNo relevant conflicts of interest to declare.

Impact of Oral Voriconazole As Prophylaxis during Intensive Chemotherapy for Adult Patients with Acute Myeloid Leukemia: A Retrospective Cohort Study Using Japanese Administrative Database

BackgroundPrevention of invasive fungal infections (IFIs) during chemotherapy is an important issue for patients with hematologic malignancies. Although preventive effects of antifungal drugs on hematological stem cell transplantation (HSCT) has been reported, the appropriate approach for prophylaxis of IFIs during intensive chemotherapy for patients with acute myeloid leukemia (AML) has been still unclear. To solve this issue, we analyzed the Diagnosis Procedure Combination/Per-Diem Payment System (DPC/PDPS) database instituted by the Ministry of Health, Labour and Welfare of Japan as a payment system for medical services, which contains a large nationwide inpatient administrative data. The objective of this study is to investigate the effectiveness of oral voriconazole (VRCZ) as prophylaxis of IFIs in adult AML patients undergoing intensive chemotherapy.MethodsThis is a retrospective cohort study analyzing data of which were extracted from the DPC database collected from July 2010 to June 2015. The prophylactic usage of oral antifungal drugs is defined as administration of these drugs for more than 3 consecutive days prescribed within 3 days after the onset of chemotherapy. The selection criteria were as follows: patients diagnosed as AML, aged 18 years or older, conducted intensive chemotherapy under aseptic management, and administrated either of oral fluconazole (FLCZ), itraconazole (ITCZ) or VRCZ for the prophylactic usage defined as above. The data for HSCT were excluded. The main analyses were conducted in these 2 groups: the group of patients administrated oral VRCZ, and the other oral azoles (FLCZ or ITCZ). Primary outcome was the intravenous antifungal drug usage within 30 days after the onset of chemotherapy. Subgroup analyses were performed in the following categories: 1) VRCZ group vs FLCZ group/VRCZ group vs ITCZ group; 2) Age < 65/ Age ≥ 65. Secondary outcomes were 1) length of hospital stay, and 2) in-hospital mortality. The log-transformed values were used for analyses of length of hospital stay because its distribution was right-skewed. Panel instrumental variable (IV) method was carried out to address unobserved time-invariant characteristics of individuals, time-variant factors, and the endogeneity due to the unmeasured bias.ResultsA total of 63,197 inpatient data of 923 hospitals for AML patients aged 18 years or older were screened. Finally, 10,083 data of 413 hospitals, including 655 in VRCZ group, 4,445 in FLCZ group, and 4,983 in ITCZ group, were met to the selection criteria. The result of panel IV approach showed that prophylactic administration of oral VRCZ was significantly associated with the reduction of intravenous antifungal drug usage within 30 days after the onset of chemotherapy compared to the other azoles (RD −0.13, 95%CI −0.22 to −0.03, p = 0.012). It was also associated with the shortening the log-transformed value of length of hospital stay compared to the other oral azoles (RD −0.11, 95%CI −0.23 to −0.00, p = 0.043), but not with in-hospital mortality (RD 0.02, 95%CI −0.04 to 0.08, p = 0.513). In the subgroup analyses, prophylactic administration of oral VRCZ was significantly associated with the reduction of intravenous antifungal drug usage compared with FLCZ (RD −0.18, 95%CI −0.28 to −0.08, p < 0.001), but not with ITCZ (RD −0.07, 95%CI −0.16 to 0.34, p = 0.150). When the data was analyzed by age groups, it was significantly associated with the reduction of intravenous antifungal drug usage in patients under the age of 65 (RD −0.18, 95%CI −0.35 to −0.02, p = 0.033), but not in patients aged 65 years or older (RD −0.09, 95%CI −0.22 to 0.04, p = 0.196) compared to the other azoles.ConclusionsBy analyzing the large nationwide database, the present results revealed that the prophylactic administration of oral VRCZ would be effective for reducing the intravenous antifungal drug usage and shortening of hospital stay, which might contribute to reduction of IFIs and improvement of quality of life especially for young AML patients receiving the intensive chemotherapy. DisclosuresYoshida:Bristol-Myers Squibb: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; ONO PHARMACEUTICAL CO., LTD.: Honoraria; Nippon Kayaku Co.,Ltd.: Honoraria.

Cotreatment with Aspirin and Azole Drugs Increases Sensitivity of Candida albicans in vitro.

PurposeThis study aimed to investigate the effects of aspirin (acetyl salicylic acid [ASA]) combined with fluconazole (FCA), itraconazole (ITR), or voriconazole (VRC) on Candida albicans under planktonic and biofilm conditions.MethodsA total of 39 clinical C. albicans strains were used to perform the in vitro drug sensitivity assay under different conditions using the M27-A4 broth microdilution method. The minimal inhibitory concentrations (MICs) and fractional inhibitory concentration index (FICI) values were calculated. C. albicans ZY23 was chosen for the further analyses.ResultsUnder planktonic conditions, the half maximal MIC (MIC50) values of FCA, ITR, and VRC were 64–0.5 μg/mL, 32–0.0625 μg/mL, and 16–0.125 μg/mL, respectively, when applied, whereas in combination with ASA, the values decreased to 32–0.25 μg/mL, 8–0.0313 μg/mL, and 8–0.0313 μg/mL, respectively. Under biofilm conditions, FCA, ITR, or VRC alone showed MIC50 values of 128–8 μg/mL, 32–4 μg/mL, and 32–0.5 μg/mL, whereas in combination with ASA the values were decreased to 32–0.5 μg/mL, 16–0.5 μg/mL, and 8–0.0625 μg/mL, respectively. Analysis of the FICI showed that the sensitization rate of ASA to FCA, ITR, and FCA under planktonic conditions was 43.59%, whereas the sensitization rates of ASP to FCA, ITR, and FCA under biofilm conditions were 46.15%, 46.15%, and 48.72%, respectively. Additionally, the time-growth and time-kill curves of C. albicans ZY23 further verified the synergistic effects of ASA on azole drugs.ConclusionASA may act as an enhancer of the inhibitory effects of azole drugs on the growth of clinical C. albicans under planktonic and biofilm conditions.

Comparison of the Sensititre YeastOne® and CLSI M38-A2 microdilution methods in determining the activity of nine antifungal agents against dermatophytes.

Dermatophytes are the most common fungal pathogens causing superficial infections in humans with a high prevalence worldwide. The treatment of these infections is based on the use of topical and systemic antifungal agents. A convenient method with a high predictive value for testing the susceptibilities of dermatophytes is necessary. To evaluate the ability of the Sensititre YeastOne® in testing the activity of nine antifungal agents against dermatophytes. We compared Sensititre® with reference procedure for anidulafungin (ANID), micafungin sodium (MCF), caspofungin acetate (CAS), 5-fluorocytosine (5FC), posaconazole (PCZ), voriconazole (VCZ), itraconazole (ITZ), fluconazole (FLZ) and amphotericin B (AMB) against 79 dermatophyte isolates, the essential agreement (EA) and categorical agreement (CA) between the two methods were obtained. The MICs or MECs obtained by the Sensititre® were usually lower than those obtained by the M38-A2. The overall EA between the two methods of nine antifungals was best for 5FC (100%), followed by MCF (94.9%), PCZ (84.8%), AMB (67.1%), FLZ (65.8%), VCZ (63.3%), ANID (29.1%), ITZ (20.3%) and CAS (2.5%). The overall CA between the two methods for all drugs was 100% except for ANID (97.4%), MCF (95%) and PCZ (92.5%). Substantial discrepancies were observed with all drugs except for VCZ and 5FC. The results of M38-A2 in terms of GMIC (or GMEC) and MIC90 (or MEC90) were, in increasing order, as follows: MCF, PCZ, VCZ, ANID, ITZ, CAS, AMB, FLZ and 5FC. The Sensititre YeastOne® shows poor EA with the reference method for dermatophytes; therefore, M38-A2 should remain the reference procedure for antifungal susceptibility testing against dermatophytes.

Activity of cinnamaldehyde, carvacrol and thymol combined with antifungal agents against Fusarium spp

ABSTRACT Fusarium spp. are able to cause a wide range of opportunistic infections in both immunocompromised and immunocompetent individuals. Thus, new therapeutic options should be considered. We evaluated the antifungal activity of three phytocompounds (cinnamaldehyde, CIN; carvacrol, CRV; and thymol, THY), as well as their activities when combined with amphotericin B (AMB), caspofungin (CAS), itraconazole (ITC) and voriconazole (VRC) against 20 clinical isolates of Fusarium. Broth microdilution (M38-A2) and checkerboard methodologies were used to obtain MICs (Minimum inhibitory concentration) and FICIs (Fractional Inhibitory Concentration Index), respectively. The MICs observed for the phytocompounds were: 20–80 for CIN, 40-> 320 for CRV and 40–320 µg/mL for THY. The most effective combinations (synergism rates) were: THY + CAS (96%), CRV + CAS (88%), THY + AMB (76%), CRV + AMB (68%), THY + VRC (68%) and CRV + VRC (60%). Our findings suggest that CRV and THY have potent activity when combined with CAS, AMB or VRC.

Antifungal Activity of Minocycline and Azoles Against Fluconazole-Resistant Candida Species.

Candida species are the most common fungal pathogens to infect humans, and can cause life-threatening illnesses in individuals with compromised immune systems. Fluconazole (FLU) is the most frequently administered antifungal drug, but its therapeutic efficacy has been limited by the emergence of drug-resistant strains. When co-administered with minocycline (MIN), FLU can synergistically treat clinical Candida albicans isolates in vitro and in vivo. However, there have been few reports regarding the synergistic efficacy of MIN and azoles when used to treat FLU-resistant Candida species, including Candida auris. Herein, we conducted a microdilution assay wherein we found that MIN and posaconazole (POS) showed the best in vitro synergy effect, functioning against 94% (29/31) of tested strains, whereas combinations of MIN+itraconazole (ITC), MIN+voriconazole (VOR), and MIN+VOR exhibited synergistic activity against 84 (26/31), 65 (20/31), and 45% (14/31) of tested strains, respectively. No antagonistic activity was observed for any of these combinations. In vivo experiments were conducted in Galleria mellonella, revealing that combination treatment with MIN and azoles improved survival rates of larvae infected with FLU-resistant Candida. Together, these results highlight MIN as a promising synergistic compound that can be used to improve the efficacy of azoles in the treatment of FLU-resistant Candida infections.

The effects of secreted aspartyl proteinase inhibitor ritonavir on azoles‐resistant strains of Candida albicans as well as regulatory role of SAP2 and ERG11

Background Candida albicans, the main human fungal pathogen, can cause fungal infection and seriously affect people's health and life. This study aimed to investigate the effects of ritonavir (RIT) on C. albicans and the correlation between SAP2 as well as ERG11 and drug resistance.ResultsSecreted aspartyl proteinases (Saps) activities and pathogenicity of C. albicans with different drug resistance were measured. M27‐A4 broth microdilution method was used to analyze the drug sensitivity of RIT combined with fluconazole (FCA) on C. albicans. After that, SAP2 and ERG11 mutations were examined by polymerase chain reaction (PCR) and sequencing, and quantitative real‐time PCR was utilized to determine the expression of the two genes. By analyzing pz values, the Saps activity of cross‐resistant strains was the highest, followed by voriconazole (VRC)‐resistant strains, FCA‐resistant strains, itraconazole (ITR)‐resistant strains, and sensitive strains. The pathogenicity of C. albicans in descending order was as follows: cross‐resistant strains, VRC‐resistant strains, ITR‐resistant strains, FCA‐resistant strains, and sensitive strains. With the increase of RIT concentrations, the Saps activity was gradually inhibited. Drug sensitivity results showed that there was no synergistic effect between RIT and FCA. Additionally, no gene mutation sites were found in SAP2 sequencing, and 17 synonymous mutations and 6 missense mutations occurred in ERG11 sequencing. Finally, the expression of SAP2 and ERG11 was significantly higher in the resistant strains compared with the sensitive strains, and there was a positive liner correlation between SAP2 and ERG11 messenger RNA expression (r = .6655, p < .001).ConclusionThese findings may help to improve our understanding of azole‐resistant mechanisms of C. albicans and provide a novel direction for clinical therapeutics of C. albicans infection.

Effect of itraconazole on the pharmacokinetics of faldaprevir in healthy subjects.

Faldaprevir (FDV), a substrate of CYP3A/P-glycoprotein (P-gp), is a selective inhibitor of the hepatitis C virus (HCV) NS3/4 protease. FDV is currently under clinical development for application in interferon-free treatment regimens for patients with chronic HCV infection. Understanding the drug-drug interaction potential of FDV is critical, as certain drug combinations may facilitate the more rapid achievement of steady-state-that is, the ideal drug concentration and balanced metabolic cycle of absorption and elimination that optimize drug efficacy. We thus conducted this study to investigate the effect of itraconazole (ICZ), a strong inhibitor of CYP3A and a moderate inhibitor of P-gp, on the pharmacokinetics (PK) of FDV. Eighteen healthy male and female volunteers participated in this open-label, fixed-sequence study. FDV 120 mg twice daily (BID) was administered on Day 1, followed by 120 mg once daily (QD) from Day 2 until the end of the 10-day study; after 6 days of FDV alone, ICZ 200 mg was added to FDV for an additional 4 days (BID on Day 7 and QD from Day 8 to Day 10). Intensive PK sampling was performed after 6 days of FDV treatment and again after 4 days of combined FDV/ICZ treatment. The adjusted geometric mean (gMean) ratios (%) of area under the concentration curve over dosing interval at steady-state (AUCτ, ss) and maximal concentration at steady-state (Cmax, ss) for combined FDV/ICZ treatment vs. FDV treatment alone were 198.6% and 180.6%, respectively, with 90% confidence intervals (CIs) of 182.4-216.1 and 165.7-196.9. Administration of FDV alone or in combination with ICZ was observed to be safe and well-tolerated. Co-administration with ICZ, however, resulted in an approximately two-fold increase in FDV steady-state exposure. Furthermore, FDV required no dosage adjustment when co-administered with ICZ.

Effects of calcineurin inhibitors, cyclosporine A and tacrolimus (FK506), on the activity of antifungal drugs against Candida spp.

Introduction. The simultaneous use of antifungals with immunosuppressive agents has become a necessity for patients taking immunosuppressive therapy. However, antifungal drugs are problematic because of their limited target.Hypothesis. Scientists have been searching for new antifungals and some compounds with at least additive effects on antifungals. Calcineurin inhibitors used as immunosuppressive agents also attract attention due to their antifungal property.Aim. To evaluate the activity of two calcineurin inhibitors alone and in combination with amphotericin B (AMB), caspofungin (CAS), itraconazole (ITR), voriconazole (VOR) and fluconazole (FLU).Methodology. MICs of AMB, CAS, ITR, VOR, FLU and cyclosporine A (CsA) and tacrolimus (TAC) as calcineurin inhibitors were evaluated by the broth microdilution method against Candida albicans (n=13), C. krusei (n=7) and C. glabrata (n=10). Checkerboard and time-kill methods were performed to investigate the activity of combining calcineurin inhibitors with antifungal drugs.Results. The lowest MIC values were detected with VOR for all Candida isolates tested. Although we did not detect any inhibition for CsA or TAC alone at concentrations tested in this study, the combinations of CAS with CsA showed the highest synergistic activity (36.7%) by the checkerboard method, and CAS with CsA and ITR with TAC combinations exhibited apparent synergistic interaction by the time-kill method. However, the combinations of both CsA and TAC with AMB resulted in antagonistic interactions, especially against C. krusei isolate in time-kill testing.Conclusion. Synergistic interactions in the combinations of TAC or CsA with antifungal drugs, except for AMB, in many concentrations was found to be promising in terms of the treatment of patients with fungal infections.

Diphenyl diselenide alone and in combination with itraconazole against Sporothrix schenckii s.str. and Sporothrix globosa.

We evaluated the in vitro susceptibility of Sporothrix schenckii s.str. and Sporothrix globosa to diphenyl diselenide (PhSe)2 alone and in association with itraconazole (ITZ). Eight clinical isolates were tested in microdilution and checkerboard assays. (PhSe)2 alone inhibited all isolates in concentration ≤ 8µg/mL and was effective in killing one S. schenckii isolate. Inhibitory and fungicidal beneficial effects in its interaction with ITZ were shown against 87.5% (7/8) and 50% (4/8) of the isolates tested, respectively. Our study demonstrates the in vitro antifungal activity of (PhSe)2 against two pathogenic Sporothrix species, suggesting studies of in vivo applications are warranted.

Preparation of Topical Itraconazole with Enhanced Skin/Nail Permeability and In Vivo Antifungal Efficacy against Superficial Mycosis.

In this study, a stable and highly skin-permeable topical delivery system for itraconazole (ITZ) was designed to provide effective treatment against superficial mycosis. Herein, ITZ was incorporated into a solution composed of ethanol, benzyl alcohol, hydrochloric acid, Transcutol P, and cyclomethicone as a delivery vehicle, solubilizer, protonating agent, permeation enhancer, and spreading agent, respectively. At 72 h, the optimal topical ITZ formulation (ITZ–TF#11) exhibited 135% enhanced skin permeability, which led to increases in drug deposition in the stratum corneum, epidermis, and dermis of 479%, 739%, and 2024%, respectively, compared with the deposition of 1% ITZ in ethanol (control). Moreover, on day 7, ITZ–TF#11 demonstrated 2.09- and 2.30-fold enhanced nail flux and drug deposition, compared with the control. At a dose of 40 mg/kg/day, ITZ–TF#11 showed 323% greater lesion recovery, a 165% lower mean erythema severity score, and a 37% lower mean logarithm of viable fungal cells in skin in the treated area, compared with mice that received oral ITZ at the same dose. Overall, the findings imply that ITZ–TF#11 is a superior alternative to oral ITZ for treatment of superficial mycosis.

Itraconazole resistance of Trichophyton rubrum mediated by the ABC transporter TruMDR2.

Dermatophytes showing reduced sensitivity to antifungal agents have emerged in several countries. One terbinafine resistant strain of Trichophyton rubrum, TIMM20092, also showed reduced sensitivity to itraconazole (ITC) and voriconazole (VRC). The expression of two genes (TruMDR2 and TruMDR3) encoding multidrug transporters of the ABC family was found to be highly up-regulated in this strain. Deletion of TruMDR3 in TIMM20092 abolished its resistance to VRC but only slightly reduced its resistance to ITC. We examined the potential of T rubrum to develop resistance to ITC by analysing the mechanism of ITC resistance in TIMM20092. The deletion of TruMDR2 by gene replacement was performed in TIMM20092 and one TruMDR3-lacking mutant (∆TruMDR3) previously generated from TIMM20092. TruMDR2 single and TruMDR2/TruMDR3 double mutants (∆TruMDR2 and ∆TruMDR2/3) were successfully obtained, respectively. The suppression of TruMDR2 was shown to abolish resistance to ITC in TIMM20092 and the TruMDR3-lacking mutant, strongly suggesting that TruMDR2 is a major contributor to ITC resistance in TIMM20092. Our study highlights the possible role of the ABC transporter TruMDR2 in ITC resistance of T.rubrum.

BIOACTIVITY CHARACTERIZATION OF PURIFIED RECOMBINANT HYPOTHETICAL PROTEIN CODED BY OPEN READING FRAME-112 OF STREPTOMYCES.

This study was aimed to investigate the Open Reading Frame-112 (ORF-112) gene, which encoded for a hypothetical 218 amino acids protein in Streptomyces bacteria. A complete ORF-112 gene was synthesized, with addition of a 6xHis-Tag at the N-terminal location. The synthesized DNA nucleotides were sub-cloned into bacterial expression plasmid pBAT4. The pBAT4-ORF-112 plasmid transformed in bacterial cells BL21(De3)pLysS, intended for protein over expression, induced by isopropyl β- d-1-thiogalactopyranoside (IPTG). The IMAC affinity chromatography technique was deployed for protein purifications. Highly-purified fractions of ORF-112 were achieved by using affinity Ni++-columns. The purified ORF-112 protein was tested for possible biological activity. The SDS-PAGE analysis exhibited a soluble 30 kDa size purified ORF-112 protein which showed a slight gel shifting from the predicted size. The virulent activity test on purified fractions of ORF-112 was measured using the Disk Diffusion Test and it disclosed a clear zone formed in response to fungi Candida albicans growth. The data implies that the ORF-112 protein has an acceptable protective effect against the fungus C. albicans as compared to positive control ketoconazole (KCZ) (P &lt; 0.05) while the protein has a significantly lower protective effect against the fungus than Itraconazole (ICZ) (P &gt; 0.05). The results clarify the hypothetical ORF-112 protein is a novel protein with protective response effect against fungi cells C. albicans on disk-diffusion.test.

Bioadhesive-Thermosensitive In Situ Vaginal Gel of the Gel Flake-Solid Dispersion of Itraconazole for Enhanced Antifungal Activity in the Treatment of Vaginal Candidiasis.

The poor solubility of itraconazole (ITZ) has limited its efficacy in the treatment of vaginal candidiasis. Accordingly, the improvement of ITZ solubility using a solid dispersion technique was important to enhance its antifungal activity. Besides, as the purpose of this research was to develop local-targeting formulations, bioadhesive-thermosensitive in situ vaginal gel combined with the gel-flake system was found to be the most suitable choice. To obtain optimum solubility, entrapment efficiency, and drug-loading capacity, optimization of solid dispersion (SD) and gel-flake formulations of ITZ was performed using a composite central design. The results showed that the optimized formulation of SD-ITZ was able to significantly enhance its solubility in both water and simulated vaginal fluid to reach the values of 4.211 ± 0.23 and 4.291 ± 0.21 mg/mL, respectively. Additionally, the optimized formulation of SD-ITZ gel flakes possessed desirable entrapment efficiency and drug-loading capacity. The in situ vaginal gel containing SD-ITZ gel flakes was prepared using PF-127 and PF-68, as the gelling agents, with the addition of hydroxypropyl methylcellulose (HPMC) as the mucoadhesive polymer. It was found that the obtained in situ vaginal gel provided desirable physicochemical properties and was able to retain an amount of more than 4 mg of ITZ in the vaginal tissue after 8 h. Importantly, according to the in vivo antifungal activity using infection animal models, the incorporation of the solid dispersion technique and gel-flake system in the formulation of the bioadhesive-thermosensitive in situ vaginal gel led to the most significant decrease of the growth of Candida albicans reaching <1 log colony-forming units (CFU)/mL or equivalent to <10% of the total colony after 14 days, indicating the improvement of ITZ antifungal activity compared to other treated groups. Therefore, these studies confirmed a great potential to enhance the efficacy of ITZ in treating vaginal candidiasis. Following these findings, several further experiments need to be performed to ensure acceptability and usability before the research reaches the clinical stage.

Species identification and antifungal susceptibility testing of Aspergillus strains isolated from patients with otomycosis in northern China

There are limited studies on species distribution and susceptibility profiles of Aspergillus strains isolated from patients with otomycosis in China. A total of 69 confirmed Aspergillus species isolates were obtained from ear swabs of patients diagnosed with otomycosis from 2017 to 2018 in northern China. Identification of these Aspergillus isolates at the species level was performed using conventional morphological methods and MALDI-TOF MS in combination with molecular sequencing, and invitro susceptibility to nine antifungal agents was evaluated using the Sensititre YeastOne system. The Aspergillus section Nigri had the greatest distribution of Aspergillus isolates. A. welwitschiae (n=25) was the most predominant isolate in section Nigri, followed by A. tubingensis (n=12) and A. niger (n=11). Other Aspergillus species were also isolated, including A. terreus (n=11), A. flavus/A. oryzae (n=8), and A. fumigatus (n=2). Amphotericin B, posaconazole, and echinocandins were highly invitro active against all the isolates tested. 2.9% (2/69) of the isolates were resistant to azoles in our study, including one A. niger isolate with a high MIC value for itraconazole (ITR) (16mg/L) and one A. tubingensis isolate cross-resistant to both voriconazole (VOR) (MIC >8mg/L) and ITR (MIC >16mg/L). One A. welwitschiae and one A. niger isolate both had increased MIC values of 4mg/L against VOR. A. welwitschiae was the most prevalent Aspergillus species isolated from patients with otomycosis. Our findings also indicated that the azole-resistant Aspergillus section Nigri should be utilized to guide clinical medication for Otomycosis.

In vitro and in vivo assessment of hydroxypropyl cellulose as functional additive for enabling formulations containing itraconazole

Using polymers as additives to formulate ternary amorphous solid dispersions (ASDs) has successfully been established to increase the bioavailability of poorly soluble drugs, when one polymer is not able to provide both, stabilizing the drug in the matrix and the supersaturated solution. Therefore, we investigated the influence of low-viscosity hydroxypropyl cellulose (HPC) polymers as an additive in HPMC based ternary ASD formulations made by hot-melt extrusion (HME) on the bioavailability of itraconazole (ITZ). The partitioning potential of the different HPC grades was screened in biphasic supersaturation assays. Solid-state analytics were performed using differential scanning calorimetry (DSC), X-ray powder diffraction (XRPD). The addition of HPCs, especially HPC-UL, resulted in a superior partitioned amount of ITZ in biphasic supersaturation assays. Moreover, the approach in using HPCs as an additive in HPMC based ASDs led to an increase in partitioned ITZ compared to Sporanox® in biorelevant biphasic dissolution studies. The results from the biphasic dissolution experiments correlated well with the in vivo studies, which revealed the highest oral bioavailability for the ternary ASD comprising HPC-UL and HPMC.

Prevalence and Antifungal Susceptibility of Candida parapsilosis Species Complex in Eastern China: A 15-Year Retrospective Study by ECIFIG.

Candida parapsilosis complex is one of the most common non-albicans Candida species that cause candidemia, especially invasive candidiasis. The purpose of this study was to evaluate the antifungal susceptibilities of both colonized and invasive clinical C. parapsilosis complex isolates to 10 drugs: amphotericin (AMB), anidulafungin (AFG), caspofungin (CAS), micafungin (MFG), fluconazole (FLZ), voriconazole (VRZ), itraconazole (ITZ), posaconazole (POZ), 5-flucytosine (FCY), and isaconazole (ISA). In total, 884 C. parapsilosis species complex isolates were gathered between January 2005 and December 2020. C. parapsilosis, Candida metapsilosis, and Candida orthopsilosis accounted for 86.3, 8.1, and 5.5% of the cryptic species, respectively. The resistance/non-wild-type rate of bloodstream C. parapsilosis to the drugs was 3.5%, of C. metapsilosis to AFG and CAS was 7.7%, and of C. orthopsilosis to FLZ and VRZ was 15% and to CAS, MFG, and POZ was 5%. The geometric mean (GM) minimum inhibitory concentrations (MICs) of non-bloodstream C. parapsilosis for CAS (0.555 mg/L), MFG (0.853 mg/L), FLZ (0.816 mg/L), VRZ (0.017 mg/L), ITZ (0.076 mg/L), and POZ (0.042 mg/L) were significantly higher than those of bloodstream C. parapsilosis, for which the GM MICs were 0.464, 0.745, 0.704, 0.015, 0.061, and 0.033 mg/L, respectively (P < 0.05). The MIC distribution of the bloodstream C. parapsilosis strains collected from 2019 to 2020 for VRZ, POZ, and ITZ were 0.018, 0.040, and 0.073 mg/L, significantly higher than those from 2005 to 2018, which were 0.013, 0.028, and 0.052 mg/L (P < 0.05). Additionally, MIC distributions of C. parapsilosis with FLZ and the distributions of C. orthopsilosis with ITZ and POZ might be higher than those in Clinical and Laboratory Standards Institute studies. Furthermore, a total of 143 C. parapsilosis complex isolates showed great susceptibility to ISA. Overall, antifungal treatment of the non-bloodstream C. parapsilosis complex isolates should be managed and improved. The clinicians are suggested to pay more attention on azoles usage for the C. parapsilosis complex isolates. In addition, establishing the epidemiological cutoff values (ECVs) for azoles used in Eastern China may offer better guidance for clinical treatments. Although ISA acts on the same target as other azoles, it may be used as an alternative therapy for cases caused by FLZ- or VRZ-resistant C. parapsilosis complex strains.

Novel cocrystals of itraconazole: Insights from phase diagrams, formation thermodynamics and solubility

In this work, the cocrystallization approach was applied to itraconazole (ITR), a very slightly soluble triazole antifungal drug, which led to the formation of two new solid forms of ITR with 4-aminobenzoic acid (4AmBA) and 4-hydroxybenzamide (4OHBZA). A thermodynamic analysis of the solid-liquid binary phase diagrams for the (ITR + 4AmBA) and (ITR + 4OHBZA) systems provided conclusive evidence of the cocrystal stoichiometry: 1:1 for the cocrystal with 4-aminobenzoic acid, and 1:2 for the cocrystal with 4-hydroxybenzamide. Powder X-Ray diffraction analysis confirmed the formation of two different polymorphic forms of the [ITR + 4OHBZA] (1:2) cocrystal obtained either through solution or melt crystallization. Cocrystal formation and polymorphic transition processes were investigated in detail by the DSC and HSM methods. The thermodynamic functions of cocrystal formation were estimated from the solubility of the cocrystals and the corresponding solubility of the pure compounds at different temperatures. The combination of ITR and 4OHBZA was found to be more favorable than the reaction between ITR and 4AmBA in terms of both Gibbs energy and enthalpy. The pH-solubility behavior of the cocrystals was investigated at different pH values using eutectic concentrations of the components and the cocrystal solubility advantage was estimated. It was found that the cocrystallization of itraconazole with 4OHBZA and 4AmBA can potentially increase the drug solubility at pH1.2 and 37 °C by 225 and 64 times, respectively. The cocrystal dissolution behavior in biorelevant media was analyzed in terms of Cmax, σmax parameters (the maximum ITR concentration and supersaturation), and AUC (the concentration area under the curve during the dissolution – supersaturation – precipitation process). The cocrystals had similar σmax values during the dissolution and sustained supersaturation for up to 6 h, which gave them an advantage in the AUC values (13–37 times higher) over the drug. The differences in the dissolution profiles of the cocrystals were rationalized in terms of their dissolution rate values.

Drug Repurposing of Itraconazole and Estradiol Benzoate against COVID-19 by Blocking SARS-CoV-2 Spike Protein-Mediated Membrane Fusion.

SARS‐CoV‐2 caused the emerging epidemic of coronavirus disease in 2019 (COVID‐19). To date, there are more than 82.9 million confirmed cases worldwide, there is no clinically effective drug against SARS‐CoV‐2 infection. The conserved properties of the membrane fusion domain of the spike (S) protein across SARS‐CoV‐2 make it a promising target to develop pan‐CoV therapeutics. Herein, two clinically approved drugs, Itraconazole (ITZ) and Estradiol benzoate (EB), are found to inhibit viral entry by targeting the six‐helix (6‐HB) fusion core of SARS‐CoV‐2 S protein. Further studies shed light on the mechanism that ITZ and EB can interact with the heptad repeat 1 (HR1) region of the spike protein, to present anti‐SARS‐CoV‐2 infections in vitro, indicating they are novel potential therapeutic remedies for COVID‐19 treatment. Furthermore, ITZ shows broad‐spectrum activity targeting 6‐HB in the S2 subunit of SARS‐CoV and MERS‐CoV S protein, inspiring that ITZ have the potential for development as a pan‐coronavirus fusion inhibitor.

Preparation and characterization of itraconazole loaded nanomicelles based on dextran–behenic acid for cutaneous leishmaniasis treatment

Cutaneous leishmaniasis is known as the most prevalent clinical form of leishmaniasis. It needs the development of new therapies due to the serious side-effects promoted by taking the current drugs. In the present study, dextran–behenic acid (DEX–BA) based nanomicelles were developed to direct the delivery of itraconazole (ITZ) to the macrophages and enhance its toxic effects against Leishmania parasites. DEX–BA was synthesized through the esterification of dextran with behenic acid. The critical micelle concentration of the newly developed conjugate was evaluated using pyrene as the fluorescent probe. The nanomicelles were generated by the dialysis method; then they were optimized by applying a Box–Behnken design. The effects of the dialysis temperature, polymer content, and sonication time on the characteristics of micelles were subsequently studied. Furthermore, in vitro efficacy against Leishmania major promastigotes and parasite-infected macrophages was evaluated. The optimized formulation showed the particle size of 195.16 ± 3.06 nm, the polydispersity index of 0.39 ± 0.01, the zeta potential of −16.29 ± 0.89 mV, the encapsulation efficiency % of 56.11 ± 4.9, and the release efficiency % of 51.29 ± 1.97. According to scanning electron microscopy, the nanomicelles were found to be nearly spherical in shape. ITZ-loaded nanomicelles showed the strongest anti-leishmanial activities when compared with the free ITZ and drug-free nanomicelles. It could be, therefore, concluded that ITZ-loaded nanomicelles might be useful as an alternative therapy for the treatment of cutaneous leishmania.