Synergistic effect of hydralazine associated with triazoles on Candida spp. in planktonic cells
Abstract
Objective: To evaluate the antifungal activity of hydralazine hydrochloride alone and in synergy with azoles against Candida spp. and the action mechanism. Methods: We used broth microdilution assays to determine the MIC, checkerboard assays to investigate synergism, and flow cytometry and molecular docking tests to ascertain action mechanism. Results: Hydralazine alone had antifungal activity in the range of 16–128 μg/ml and synergistic effect with itraconazole versus 100% of the fungal isolates, while there was synergy with fluconazole against 11.11% of the isolates. There was molecular interaction with the receptors exo-B(1,3)-glucanase and CYP51, causing reduced cell viability and DNA damage. Conclusion: Hydralazine is synergistic with itraconazole and triggers cell death of Candida spp. at low concentrations, demonstrating antifungal potential.
Tweetable abstract
We evaluated the effect of hydralazine on Candida spp. and observed its fungicidal effect as a consequence of the damage to the fungal DNA, as well as the enhanced effects of association with itraconazole.
Papers of special note have been highlighted as: • of interest; •• of considerable interest
References
- 1. . Revisiting species distribution and antifungal susceptibility of Candida bloodstream isolates from Latin American medical centers. J. Fungi 3(2), 11–17 (2017). • Shows epidemiology of candidemia in Latin America.
- 2. Clinical predictors and outcome impact of community-onset polymicrobial bloodstream infection. Int. J. Antimicrob. Agents 54(6), 716–722 (2019).
- 3. Evaluation of Candida bloodstream infection and antifungal utilization in a tertiary care hospital. BMC Infect. Dis. 18(1), 187 (2018).
- 4. . Invasive fungal disease in humans: are we aware of the real impact? Mem. Inst. Oswaldo Cruz 115(9), e200430 (2020).
- 5. Changes in prevalence of health care-associated infections in US hospitals. N. Engl. J. Med. 379(18), 1732–1744 (2018).
- 6. . Changes in the epidemiological landscape of invasive candidiasis. J. Antimicrob. Chemother. 73(January), i4–i13 (2018).
- 7. . Secular trends of candidemia at a Brazilian tertiary care teaching hospital. Brazilian J. Infect. Dis. 22(4), 273–277 (2018).
- 8. . Pathogenesis of Candida albicans biofilm. Pathog. Dis. 74(4), ftw018 (2016).
- 9. . Increased antimicrobial resistance during the COVID-19 pandemic. Int. J. Antimicrob. Agents. 57(4), 106324 (2020).
- 10. . Drug repurposing: a promising tool to accelerate the drug discovery process. Drug Discov. Today 24(10), 2076–2085 (2019).
- 11. . Drug repurposing for antimicrobial discovery. Nat. Microbiol. 4(4), 565–577 (2019).
- 12. . The antihypertensive drug hydralazine activates the intrinsic pathway of apoptosis and causes DNA damage in leukemic T cells. Oncotarget 7(16), 21875–21886 (2016). • Shows that hydralazine causes cell death via mitochondrial apoptotic pathway and causes DNA damage.
- 13. . Covalent adduct formation between the antihypertensive drug hydralazine and abasic sites in double- and single-stranded DNA. Chem. Res. Toxicol. 27(12), (2014).
- 14. M27-A3. Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts; Approved Standard (3rd Edition). Clinical and Laboratory Standards Institute, PA, USA (2008).
- 15. M27-S4 Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts. Clinical and Laboratory Standards Institute. PA, USA (2012).
- 16. . Comparação de métodos para avaliação da atividade antimicrobiana e determinação da concentração inibitória mínima (cim) de extratos vegetais aquosos e etanólicos. Arq. Inst. Biol. (Sao Paulo) 81(3), (2014).
- 17. . Synergy, antagonism, and what the chequerboard puts between them. J. Antimicrob. Chemother. 52(1), 1 (2003).
- 18. Antifungal activity of naphthoquinoidal compounds in vitro against fluconazole-resistant strains of different Candida species: a special emphasis on mechanisms of action on Candida tropicalis. PLoS ONE 9(5), (2014).
- 19. Kauren-19-oic acid induces DNA damage followed by apoptosis in human leukemia cells. J. Appl. Toxicol. 29(7), 560–568 (2009).
- 20. Berberine antifungal activity in fluconazole-resistant pathogenic yeasts: action mechanism evaluated by flow cytometry and biofilm growth inhibition in Candida spp. Antimicrob. Agents Chemother. 60(6), 3551–3557 (2016).
- 21. . MarvinSketch and MarvinView: molecule applets for the world wide web. Proceedings of The 3rd International Electronic Conference on Synthetic Organic Chemistry. p1775 (2019).
- 22. . Capillary surfaces with free boundary in a wedge. Adv. Math. (NY) 262, 476–483 (2014).
- 23. . Merck molecular force fieldI. Basis, form, scope, parameterization, andperformance of MMFF94. J. Comput. Chem. 17(5), 490–519 (1996).
- 24. . Structure of the catalytic core of S. cerevisiae DNA polymerase η: implications for translesion DNA synthesis. Mol. Cell 8(2), 417–426 (2001).
- 25. . Structural features and thermodynamics of the J4/5 loop from the Candida albicans and Candida dubliniensis group I introns. Biochemistry 43(50), 15822–15837 (2004).
- 26. . Analyses of Candida Cdc13 orthologues revealed a novel OB fold dimer arrangement, dimerization-assisted DNA binding, and substantial structural differences between Cdc13 and RPA70. Mol. Cell. Biol. 32(1), 186–198 (2012).
- 27. Candida albicans SOD5 represents the prototype of an unprecedented class of Cu-only superoxide dismutases required for pathogen defense. Proc. Natl Acad. Sci. USA 111(16), 5866–5871 (2014).
- 28. . Minor structural consequences of alternative CUG codon usage (Ser for Leu) in Candida albicans exoglucanase. Protein Eng. 13(10), 735–738 (2000).
- 29. . Structural basis for Mep2 ammonium transceptor activation by phosphorylation. Nat. Commun. 7, 1–11 (2016).
- 30. Structural analyses of Candida albicans sterol 14α-demethylase complexed with azole drugs address the molecular basis of azole-mediated inhibition of fungal sterol biosynthesis. J. Biol. Chem. 292(16), 6728–6743 (2017).
- 31. Tricyclic 1,5-naphthyridinone oxabicyclooctane-linked novel bacterial topoisomerase inhibitors as broad-spectrum antibacterial agents-SAR of left-hand-side moiety (part-2). Bioorganic Med. Chem. Lett. 25(9), 1831–1835 (2015).
- 32. . Specific roles of protein–phospholipid interactions in the yeast cytochrome bc1 complex structure. EMBO J. 20(23), 6591–6600 (2001).
- 33. . Using AutoDock 4 and AutoDock Vina with AutoDockTools: a tutorial. Scripps Res. Inst. Mol. (December), 32 (2012).
- 34. . AutoDock Vina: improving the speed and accuracy of docking with a new scoring function, efficient optimization, and multithreading. J. Comput. Chem. 31(2), 455–461 (2009).
- 35. UCSF Chimera – a visualization system for exploratory research and analysis. J. Comput. Chem. 25(13), 1605–1612 (2004).
- 36. . Virtual screening based on molecular docking of possible inhibitors ofCovid-19 main protease. Microbial Pathogenesis 148, 104365 (2020).
- 37. . An alternative method for the evaluation of docking performance: RSR vs RMSD. J. Chem. Inf. Model. 48(7), 1411–1422 (2008).
- 38. . Lipophilicity, pharmacokinetic properties, and molecular docking study on sars-cov-2 target for betulin triazole derivatives with attached 1,4-quinone. Pharmaceutics 13(6), 781 (2021).
- 39. . Combination antifungal therapy: a review of current data. J. Clin. Med. Res. 9(6), 451–456 (2017).
- 40. . Candida biofilms on implanted biomaterials: a clinically significant problem. FEMS Yeast Res. 6(7), 979–986 (2006).
- 41. Synergistic anticandidal activity of etomidate and azoles against clinical fluconazole-resistant Candida isolates. Future Microbiol. 14(17), 1477–1488 (2019).
- 42. . Anti-candidal activity of selected analgesic drugs used alone and in combination with fluconazole, itraconazole, voriconazole, posaconazole and isavuconazole. J. Mycol. Med. 28(2), 327–331 (2018).
- 43. A three-dimensional model of lanosterol 14α-demethylase of Candida albicans and its interaction with azole antifungals. J. Med. Chem. 43(13), 2493–2505 (2000).
- 44. . Apoptosis-inducing factor: structure, function, and redox regulation. Antioxidants Redox Signal. 14(12), 2545–2579 (2011).
- 45. . A novel mechanism of fluconazole: fungicidal activity through dose-dependent apoptotic responses in Candida albicans. Microbiology 164(2), 194–204 (2018).
- 46. . Antimicrobial activity and acetylcholinesterase inhibition by extracts from chromatin modulated fungi. Brazilian J. Microbiol. 49(1), 169–176 (2018).
- 47. . Fungal persister cells: the basis for recalcitrant infections? PLoS Pathog. 14(10), 1–14 (2018).
- 48. . Characterisation of Candida parapsilosis CYP51 as a drug target using Saccharomyces cerevisiae as host. J. Fungi 8(1), 69 (2022). • Suggests that CYP51 as a target of antifungals can overcome the resistance of pathogenic fungi to existing azole drugs.
- 49. Nature of β-1,3-glucan-exposing features on Candida albicans cell wall and their modulation. MBio 13(6), 1–19 (2022). • Shows that b-1,3-glucan exposed in Candida is the target of phagocytic attack and that lactate-induced masking reduces exposure and phagocytosis.
- 50. A phase II study of epigenetic therapy with hydralazine and magnesium valproate to overcome chemotherapy resistance in refractory solid tumors. Ann. Oncol. 18(9), 1529–1538 (2007).
- 51. . Encouraging results with the compassionate use of hydralazine/valproate (TRANSKRIP™) as epigenetic treatment for myelodysplastic syndrome (MDS). Ann. Hematol. 96(11), 1825–1832 (2017). •• Study on the repurpose of hydralazine in anticancer therapy.
- 52. The role of Cryptococcus neoformans histone deacetylase genes in the response to antifungal drugs, epigenetic modulators and to photodynamic therapy mediated by an aluminium phthalocyanine chloride nanoemulsion in vitro. J. Photochem. Photobiol. B Biol. 216, 112131 (2021).
- 53. The Rpd3/Hda1 family of histone deacetylases regulates azole resistance in Candida albicans. J. Antimicrob. Chemother. 70(7), 1993–2003 (2015).