The Antifungal and Synergistic Effect of Bisphosphonates in Cryptococcus

Abstract

New treatment strategies are required for cryptococcosis, a leading mycosis in HIV-AIDS patients. Following the identification of Cryptococcus proteins differentially expressed in response to fluconazole, we targetted farnesyl pryrophosphate synthetase (FPPS), an enzyme in the squalene biosynthesis pathway, using nitrogenous bisphosphonates. We hypothesised that these would disrupt squalene synthesis and thereby produce synergy with fluconazole, which acts on a downstream pathway that requires squalene. The susceptibilities of 39 clinical isolates from 6 different species of Cryptococcus were assessed for bisphosphonates and fluconazole, used both independently and in combination. Effective fluconazole-bisphosphonate combinations were then assessed for fungicidal activity, efficacy against biofilms and ability to resolve cryptococcosis in an invertebrate model. The nitrogenous bisphosphonates risedronate, alendronate and zoledronate were antifungal against all strains tested. Zoledronate was the most effective (GM MIC = 113.03 mg/L; risedronate = 378.49 mg/L; alendronate = 158.4 for) and was broadly synergistic when combined with fluconazole, with an FICI ≤ 0.5 in 92% of isolates. Fluconazole and zoledronate in combination were fungicidal in a time-kill assay, inhibited Cryptococcus biofilms, prevented the development of fluconazole resistance and resolved infection in a nematode model. Supplementation with squalene eliminated bisphosphonate-mediated synergy, demonstrating that synergy was due to the inhibition of squalene biosynthesis. This study demonstrates the utility of targeting squalene synthesis for improving the efficacy of azole-based antifungal drugs, and suggests bisphosphonates are promising lead compounds for further antifungal development.

Augmenting Azoles with Drug Synergy to Expand The Antifungal Toolbox

“…Like statins, bisphosphonates operate on the mevalonate pathway where they target farnesyl pyrophosphate synthase (or Erg11; Figure 2) [287]. Of the bisphosphonates tested, zoledronate resulted in strong synergy across numerous strains of Cryptococcus tested in vitro and in an in vivo model and significantly limited the development of antifungal resistance [167]. Due to their propensity to bind to bone mineral and their implication in osteonecrosis, market bisphosphonates have limited applicability for the treatment of invasive mycoses [287,288].…”

Advances in Antifungal Development: Discovery of New Drugs and Drug Repurposing

“…When it comes to drug delivery through stratum corneum, microemulsions are very resourceful considering the ability of the oils and surfactants included in their composition to act as skin penetration enhancers [170,172]. Furthermore, they are also good candidates for oral delivery of poorly water-soluble drugs as they can improve their solubilization.…”

“…Cryptococcus sp. [172] Immunomodulators promethazine [173,174] terfenadine Effective against biofilms [175] ebastine Effective against biofilms [175] dexamethasone Effective against biofilms [176] budesonide Effective in vivo [177] methotrexate [178] Psychoactives bromperidol [179] fluoxetine Effective in vivo, C. albicans only [180] haloperidol [173] sertraline Incl.…”

One‐Pot Microwave‐Assisted Synthesis, in Vitro Anti‐inflammatory Evaluation and Computer‐Aided Molecular Design of Novel Sulfamide‐Containing Bisphosphonates Derivatives

“…[10] Nitrogen bisphosphonates or aminobisphosphonates (N-BPs), the most potent class, strongly inhibit bone resorption, and have a higher binding affinity than non-aminobisphosphonates, due to electrostatic interactions such as hydrogen bonds on the hydroxyapatite surface, the main mineral constituent of bone. [11] Furthermore, these compounds have gained considerable attention in the field of medicinal chemistry due to their role in many biological activities, including anti-inflammatory, [12] antioxidant, [13] antifungal, [14,15] antibacterial and antiviral, [16] anticancer, [17] antiparasitic, [18] herbicidal, [19] as well as chelating agents. [20,21] However, an additional bioactive functionalized moiety in the bisphosphonate molecule such as 1H-indazole, 1Hpyrazolo [3,4-b]pyridine and 1H-pyrazolo [3,4-b]quinolone, [22] 2amino-benzothiazole, [23] nitrogen and sulfur, [24] sulfonylamino, [25] fluoro-substituted amine, [26] pyrazole, [27] acrylamide-derived monomer, [28] amino acid, [29] hyaluronic acid, [30] and ethylidene-pyridine bisphosphonate polyoxovanadates complexes, [31] give new derivatives for this class of molecules and offer promising and remarkable biological and industrial applications.…”

“…Furthermore, these compounds have gained considerable attention in the field of medicinal chemistry due to their role in many biological activities, including anti‐inflammatory, [12] antioxidant, [13] antifungal, [14,15] antibacterial and antiviral, [16] anticancer, [17] antiparasitic, [18] herbicidal, [19] as well as chelating agents [20,21] …”