1,3-β-D-glucan synthase enzyme complex
Rezafungin targets the fungal β-1,3-glucan synthase enzyme complex, which is essential for the synthesis of β-1,3-glucan, a critical component of the fungal cell wall. By binding to and inhibiting this enzyme, rezafungin disrupts the integrity of the cell wall, leading to osmotic instability, leakage of cellular contents, and cell death. This mechanism is fungicidal against Candida species and fungistatic against Aspergillus species. Its target is not present in mammalian cells, providing selective toxicity against fungi.

Rezafungin is an echinocandin antifungal drug. It inhibits the 1,3-β-D-glucan synthase enzyme complex present in the cell walls of fungi, responsible for the formation of 1,3-β-D-glucan. Since 1,3-β-D-glucan is an essential component of fungal cell walls, rezafungin disrupts the cell wall of fungal species, including _Candida_ spp., and acts as a concentration-dependent _in vitro_ fungicidal. Mammalian cells do not express β-1-3-glucan synthase; therefore, the action of rezafungin is specific to fungi. Rezafungin is a member of the family of echinocandins that inhibits 1,3-beta-D-glucan synthase.
In vitro, rezafungin exhibits potent activity against a wide range of Candida species, including C. albicans, C. glabrata, C. parapsilosis, and C. tropicalis, with MIC₉₀ values typically ≤0.5 µg/mL. It also shows activity against Aspergillus fumigatus and other molds. Time-kill studies demonstrate rapid fungicidal activity against Candida, with >99.9% killing within 2-4 hours. The compound has a low propensity for resistance, and cross-resistance with other echinocandins is uncommon. Its activity is not affected by serum protein binding, making it effective in vivo.

In a mouse infection model, rezafungin (Biafungin) has demonstrated strong in vivo prophylaxis against Pneumocystis jiroveci [1].
In vivo, rezafungin has demonstrated efficacy in animal models of invasive candidiasis, including murine models of disseminated and intra-abdominal candidiasis. Intraperitoneal or intravenous administration at doses of 1-10 mg/kg reduced fungal burden in kidneys, liver, and spleen. The compound's prolonged half-life (>30 hours in humans) allows for once-weekly dosing, which has been confirmed in clinical trials. In a Phase 2/3 trial (STRIVE and ReSTORE), rezafungin was non-inferior to caspofungin for the treatment of candidemia and invasive candidiasis.

In vitro susceptibility testing for rezafungin follows CLSI or EUCAST guidelines using broth microdilution. MICs are determined by visual or spectrophotometric reading after 24 hours of incubation in RPMI medium. Quality control strains (e.g., C. albicans ATCC 90028) are used to validate the assay. The compound's activity against biofilms can be assessed using a 96-well plate model and measuring metabolic activity via XTT reduction. The minimal fungicidal concentration (MFC) is determined by subculturing the contents of clear wells on solid medium.

In vitro cellular experiments for rezafungin involve co-incubation with human cell lines (e.g., HepG2, THP-1) to assess cytotoxicity. The compound shows minimal toxicity to mammalian cells at concentrations 100-fold higher than its MICs. The effect on cytokine release can be measured in human peripheral blood mononuclear cells (PBMCs) to evaluate immunomodulatory potential. Fungal killing is assessed using a time-kill assay with Candida species in RPMI broth, with samples plated on Sabouraud agar at intervals to enumerate surviving colonies.

Animal/Disease Models: C3H/HeN mice [1]
Doses: 1 mg/kg
Route of Administration: intraperitoneal (ip) injection; one time/day for 6 days.
Experimental Results: The burden of cell nuclei and ascus was Dramatically diminished.
In vivo animal studies for rezafungin are typically performed in immunosuppressed mouse models (e.g., cyclophosphamide or corticosteroid-treated). Animals are infected intravenously or intraperitoneally with a lethal dose of Candida or Aspergillus. Rezafungin is administered via IV once or twice weekly. Efficacy is measured by survival, fungal burden in target organs (CFU/g), and histopathology. Pharmacokinetic/pharmacodynamic (PK/PD) indices (e.g., AUC/MIC) are derived to optimize dosing regimens.

Absorption
Rezafungin dosage ranges from 50 mg (equivalent to 0.125 times the maximum recommended loading dose) to 400 mg, with both Cmax and AUC increasing proportionally with the dose, regardless of whether it is a single or multiple dose. In patients with candidemia and invasive candidiasis, an initial loading dose of 400 mg rezafungin, followed by weekly 200 mg doses, resulted in Cmax of 19.2 mcg/mL, AUC0-168 (AUC0-168) of 0-168 hours post-dose, and Cmin of 2.4 mcg/mL on day 1. In the same cohort, Cmax, AUC0-168, and Cmin of day 15 were 11.8 mcg/mL, 667 mcg/mL, and 2.2 mcg/mL, respectively. Compared with healthy subjects, patients with candidemia had reduced AUC0-168 and Cmax by 30% and 19%, respectively. Age, sex, race, weight, and hepatic impairment had no clinically significant effect on the pharmacokinetics of rezafenib.
Elimination Route
Rizafenib is primarily excreted in feces. In healthy subjects, 74.3% of the drug was primarily excreted in feces as rezafenib, while 25.7% was primarily excreted in urine as inactive metabolites of rezafenib.
Volume of Distribution
The volume of distribution of rezafenib is 67 L.
Clearance
The clearance of rezafenib is 0.35 L/hr.
Protein Binding
Rizafenib has a high protein binding rate. Protein binding in patients increased from 87.5% to 93.6%, and in healthy adults from 95.6% to >98.6%.
Metabolism/Metabolites
Rizafenone is metabolized via hydroxylation of the terphenylpentyl ether side chain, producing three hydroxylated metabolite isomers: 2'-, 3'-, or 4'-hydroxypentylrezafenone. Rizafenone can also lose the pentyl group via O-dealkylation to produce depentylrezafenone. Subsequent binding (sulfation) reactions of the hydroxyl metabolites are minimal. Rizafenone is not metabolized in the liver and is not expected to become a clinically relevant substrate for CYP450 enzymes.
Biological Half-Life
The terminal half-life of rizafenone is 152 hours.
The pharmacokinetic properties of rezafungin are characterized by a very long half-life (approximately 130-150 hours in humans) and linear dose-proportional exposure. It has high protein binding (>99%), but this does not affect its antifungal activity. The compound is not metabolized significantly by CYP450 enzymes; it is slowly degraded via peptide hydrolysis, and excretion occurs unchanged in feces and urine. The long half-life enables once-weekly intravenous administration, which is a major clinical advantage over daily echinocandins.

Effects During Pregnancy and Lactation
◉ Overview of Use During Lactation
There is currently no information regarding the use of rezafenamic acid during lactation. Because rezafenamic acid binds to plasma proteins at a rate of approximately 90% and has low oral bioavailability, it is unlikely to enter breast milk and be absorbed by the infant. If the mother needs to use rezafenamic acid, breastfeeding should not be discontinued.
◉ Effects on Breastfed Infants
As of the revision date, no relevant published information was found.
◉ Effects on Lactation and Breast Milk
As of the revision date, no relevant published information was found.
The toxicity profile of rezafungin is favorable. In clinical trials, the most common adverse events were mild-to-moderate, including gastrointestinal disturbances (nausea, vomiting), headache, and infusion-related reactions. These were comparable to placebo or other echinocandins. Hepatotoxicity (elevated transaminases) was observed but was generally transient. No significant nephrotoxicity or cardiotoxicity has been reported. The compound is well-tolerated, with a safety profile similar to other echinocandins, and no dose-limiting toxicity at therapeutic doses.

[1]. Melanie Cushion, et al. Rezafungin Prophylactic Efficacy in a Mouse Model of Pneumocystis Pneumonia. VOLUME 25, ISSUE 3, SUPPLEMENT, S366, MARCH 01, 2019.
[2]. Sofjan AK, et al. Rezafungin (CD101), a next-generation echinocandin: A systematic literature review and assessment of possible place in therapy. J Glob Antimicrob Resist. 2018 Sep;14:58-64.

Rezafungin is an echinocandin antibiotic that inhibits 1,3-β-D-glucan synthase. Developed by Cidara Therapeutics, it is approved for the treatment of candidemia and invasive candidiasis in patients aged 18 years and older with limited alternative treatment options. Rezafungin is an echinocandin antifungal drug, belonging to the quaternary ammonium ion class of antibiotics, and is also a nitrogen-containing macrocyclic compound, homocyclic peptide, and aromatic ether compound. Rezafungin is a new generation of semi-synthetic cyclic lipopeptide and echinocandin derivative with potential antifungal activity. After administration, rezafungin inhibits the fungal-specific enzyme 1,3-β-D-glucan synthase (which is crucial for fungal cell wall synthesis), resulting in reduced β(1,3)-D-glucan synthesis. This weakens the fungal cell wall, leading to osmotic lysis, fungal cell wall rupture, and fungal cell death. See also: Rizafenib (note moved to).
Rezafungin is a next-generation echinocandin antifungal agent approved for the treatment of invasive candidiasis and candidemia. Its once-weekly dosing schedule represents a significant advancement over daily echinocandins, improving patient convenience and potentially reducing healthcare costs. It has been evaluated in Phase 2 (STRIVE) and Phase 3 (ReSTORE) trials, which demonstrated non-inferiority to caspofungin. Rezafungin is also being investigated for the prevention of invasive fungal infections in hematopoietic stem cell transplant recipients. It offers a valuable option for managing serious fungal infections, especially in hospitalized patients.

C63H85N8O17

1226.3924176693

1225.603

1396640-59-7

Rezafungin acetate;1631754-41-0; 1396640-59-7 (cation) ; 1396640-60-0 (TFA)

78318119

Typically exists as solid at room temperature

2.9

13

17

18

88

2300

15

CCCCCOC1=CC=C(C=C1)C2=CC=C(C=C2)C3=CC=C(C=C3)C(=O)N[C@H]4C[C@H]([C@H](NC(=O)[C@@H]5[C@H]([C@H](CN5C(=O)[C@@H](NC(=O)[C@@H](NC(=O)[C@@H]6C[C@H](CN6C(=O)[C@@H](NC4=O)[C@@H](C)O)O)[C@@H]([C@H](C7=CC=C(C=C7)O)O)O)[C@@H](C)O)C)O)OCC[N+](C)(C)C)O

LNFCWEXGZIEGJW-TXSVMFMRSA-O

InChI=1S/C63H84N8O17/c1-8-9-10-28-87-45-25-21-40(22-26-45)38-13-11-37(12-14-38)39-15-17-42(18-16-39)56(80)64-46-31-48(76)61(88-29-27-71(5,6)7)68-60(84)52-53(77)34(2)32-70(52)63(86)50(36(4)73)66-59(83)51(55(79)54(78)41-19-23-43(74)24-20-41)67-58(82)47-30-44(75)33-69(47)62(85)49(35(3)72)65-57(46)81/h11-26,34-36,44,46-55,61,72-73,75-79H,8-10,27-33H2,1-7H3,(H5-,64,65,66,67,68,74,80,81,82,83,84)/p+1/t34-,35+,36+,44+,46-,47-,48+,49-,50-,51-,52-,53-,54-,55-,61+/m0/s1

2-[[(3S,6S,9S,11R,15S,18S,20R,21R,24S,25S,26S)-6-[(1S,2S)-1,2-dihydroxy-2-(4-hydroxyphenyl)ethyl]-11,20,25-trihydroxy-3,15-bis[(1R)-1-hydroxyethyl]-26-methyl-2,5,8,14,17,23-hexaoxo-18-[[4-[4-(4-pentoxyphenyl)phenyl]benzoyl]amino]-1,4,7,13,16,22-hexazatricyclo[22.3.0.09,13]heptacosan-21-yl]oxy]ethyl-trimethylazanium

Rezafungin; rezafungin acetate; 1631754-41-0; Rezafungin (acetate); acetate de rezafungine; acetato de rezafungina; Biafungin; Rezzayo

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)