Leishmania;Plasmodium

The infusion-related toxicity of amphotericin B, which includes fever and chills, limits its administration. This effect is thought to be caused by innate immune cells producing proinflammatory cytokines. TLR2 and CD14-expressing cells release inflammatory cytokines and undergo signal transduction when exposed to amphotericin B[1]. Amphotericin B's relative toxicity limits its usefulness as it interacts with cholesterol, the primary sterol found in mammalian membranes. In the subphase, amphotericin B is distributed either as a highly aggregated state or as a pre-micellar state[2].Amphotericin B only kills Leishmania promastigotes (LPs) that are unicellular when they form aqueous pores that are permeable to small cations and anions. A polarization potential is induced by amphotericin B (0.1 mM) in liposomes loaded with KCl and suspended in an iso-osmotic sucrose solution, signifying K+ leakage. The negative membrane potential nearly completely collapses when amphotericin B (0.05 mM) is added, indicating Na+ entry into the cells[3].

In the hamster scrapie model, amphotericin B causes the incubation period to be extended and the accumulation of PrPSc to be reduced. In mice suffering from transmissible subacute spongiform encephalopathies (TSSE), amphotericin B significantly lowers PrPSc levels[4]. In mouse malaria, amphotericin B directly affects Plasmodium falciparum and has an impact on parasitemia, host survival, and eryptosis of infected erythrocytes. In mice infected with Plasmodium berghei, amphotericin B tends to postpone the development of parasitemia and considerably postpones host death[5].

Polyfect reagent and DEAE-dextran are used to transiently transfect THP-1 and HEK293 cells, respectively. Genes encoding the NF-κB-dependent pELAM-luc luciferase reporter, TLR2, TLR4, CD14, and MD2 are present in transfected plasmids. In 12-well plates, cells (5×105 THP-1 or 1×105 HEK293) are added, and after 18 hours, they are washed and stimulated for 5 hours. Following the instructions, cells are lysed in reporter lysis buffer, and the lysates are subjected to luminescence analysis using a Monolight 3010 luminometer and Promega luciferase substrate.

AmB-induced cell death kinetics against Leishmania promastigotes are monitored via fluorometry employing ethidium bromide (EB), a compound that binds DNA. A SPEX Fluorolog II spectrophotometer is used to measure fluorescence at excitation-emission wavelengths of 365–580 nm. Promastigotes are added to a fluorescence cuvette containing 2 mL of various buffered solutions, always containing 10 mM glucose and 50 mM EB, and incubated for 5 minutes with gentle stirring at a final concentration of 25×106 cells/mL.Following the attainment of signal stabilization, AmB is introduced and dissolved in dimethylsulfoxide. Digitonin (50 mg/mL) is always added to achieve maximum EB incorporation. A buffer of 75 mM TRIS (pH 4 7.6) is applied to all solutions, which also contain 150 mM KCl (BK+), 150 mM NaCl (BNa+), 150 mM choline chloride, 100 mM sucrose, and 100 mM NaCl. A sophisticated instrument called the SW2 osmometer is always used to adjust the osmolarity of all solutions to 390±5 mOsm.

Efficacy of PEO-b-p(HASA)/AmB. Efficacy was assessed by organism killing in the kidneys of a neutropenic murine model of disseminated fungal infection as described previously by Andes et al. A clinical isolate of Candida albicans (K-1) was grown and quantified on SDA. For 24 h prior to infection, the organism was subcultured at 35 °C on SDA slants. A 106 CFU/mL inoculum (CFU, colony forming units) was prepared by placing six fungal colonies into 5 mL of sterile, depyrogenated normal (0.9%) saline warmed to 35 °C. Six-week-old ICR/Swiss specific-pathogen-free female mice were obtained from Harlan Sprague Dawley . All animal studies were approved by the Animal Research Committee of the William S. Middleton Memorial VA Hospital (Madison, WI). The mice were weighed (23−27 g) and given intraperitoneal injections of cyclophosphamide to render neutropenia. (For the purposes of this study, neutropenia was defined as <100 polymorphonuclear leukocytes/mm3.) Each mouse was dosed with 150 mg/kg of cyclophosphamide 4 days prior to infection and 100 mg/kg 1 day before infection. Disseminated candidiasis was induced via tail vein injection of 100 μL of inoculum. [5]
The AmB/polymeric micelle formulations or micelle blanks were reconstituted with 1.0 mL of 5% dextrose. The treatment group was given single 200 μL intravenous (iv) injections of reconstituted AmB/PEO-b-p(HASA), 91% 2 h postinfection. Doses were calculated in terms of mg of AmB/kg of body weight. Control animals were given a placebo of “blank” polymeric micelles. Over time, two animals per experimental condition were sacrificed by CO2 asphyxiation. The kidneys from each animal were removed and homogenized. The homogenate was diluted 10-fold with 9% saline and plated on SDA. The plates were then incubated for 24 h at 35 °C and inspected for CFU determination. The lower limit of detection for this technique is 100 CFU/mL. To compare the antifungal activity of the AmB/ micelle formulations with that of Fungizone, animals were dosed with equivalent doses of AmB as Fungizone as described above. The control animals for the Fungizone group received 200 μL iv injections of 5% dextrose. All results are expressed as the mean CFU per kidney for two animals (four kidneys total). The change in the area under the time−kill curves was calculated by ΔAUCTK = AUCcontrol − AUCtreatment. Outcomes were compared using ANOVA on ranks.[5]

Absorption, Distribution and Excretion
The bioavailability for intravenous infusion is 100%.
39 ± 22 mL/hr/kg [Patients with febrile neutropenia, cancer, and bone marrow transplant, receiving an infusion of 1 mg/kg/day on day 1]
17 ± 6 mL/hr/kg [Patients with febrile neutropenia, cancer, and bone marrow transplant, receiving an infusion of 1 mg/kg/day after 3-20 days]
51 ± 44 mL/hr/kg [Patients with febrile neutropenia, cancer, and bone marrow transplant, receiving an infusion of 2.5 mg/kg/day on day 1]
22 ± 15 mL/hr/kg [Patients with febrile neutropenia, cancer, and bone marrow transplant, receiving an infusion of 2.5 mg/kg/day after 3-20 days]
21 ± 14 mL/hr/kg [Patients with febrile neutropenia, cancer, and bone marrow transplant, receiving an infusion of 5 mg/kg/day on day 1]
11 +/- 6 mL/hr/kg [For patients with febrile neutropenia, cancer, and bone marrow transplantation, an infusion of 5 mg/kg/day is administered 3–20 days later.]
The pharmacokinetics of amphotericin B vary depending on the route of administration, such as conventional amphotericin B (prepared with sodium deoxycholate), amphotericin B cholesterol sulfate complex, amphotericin B lipid complex, or amphotericin B liposomes. Therefore, the pharmacokinetic parameters of one amphotericin B formulation should not be used to predict the pharmacokinetics of any other amphotericin B formulation.
Amphotericin B is poorly absorbed from the gastrointestinal tract and must be administered via parenteral route to treat systemic fungal infections. In one study, after an intravenous infusion of 30 mg amphotericin B over several hours, the mean peak serum concentration was approximately 1 μg/ml; at a dose of 50 mg, the mean peak serum concentration was approximately 2 μg/ml. The serum concentration of amphotericin B after infusion did not exceed 10% of the administered dose. It has been reported that the mean minimum serum concentration (recorded before the next infusion) is approximately 0.4 μg/ml when administered at a daily dose of 30 mg or every other day at a dose of 60 mg. Information on the distribution of amphotericin B is limited, but it is clearly multicompartmental. The reported volume of distribution after routine administration of amphotericin B is 4 L/kg. The reported steady-state volume of distribution after administration of amphotericin B cholesterol sulfate is 3.8–4.1 L/kg. Following intravenous administration of routine amphotericin B, the concentrations of amphotericin B in the inflamed pleura, peritoneum, synovium, and aqueous humor are approximately 60% of the corresponding plasma concentrations; the drug can also be distributed in vitreous fluid, pleural fluid, pericardial fluid, peritoneal fluid, and synovial fluid. Amphotericin B has been reported to cross the placenta and reach lower concentrations in amniotic fluid. Following intravenous administration of routine amphotericin B, the concentration of the drug in cerebrospinal fluid is approximately 3% of the corresponding serum concentration. Intrathecal administration is usually required to achieve the cerebrospinal fluid concentrations needed for inhibition. In patients with meningitis, intrathecal injection of 0.2–0.3 mg of conventional amphotericin B via subcutaneous reservoir sheath resulted in peak cerebrospinal fluid (CSF) concentrations of 0.5–0.8 ug/mL; 24 hours after administration, CSF concentrations were 0.11–0.29 ug/mL. Amphotericin B is cleared from CSF via arachnoid villi and appears to be stored in the extracellular spaces of the brain, which may serve as a reservoir for the drug. For more complete data on the absorption, distribution, and excretion of amphotericin B (14 in total), please visit the HSDB record page. Metabolites/Metabolites Excreted only by the kidneys. Biological Half-Life The initial plasma half-life is approximately 24 hours, and the elimination half-life is approximately 15 days. The distribution half-life of the amphotericin B cholesterol sulfate complex is 3.5 minutes, and the elimination half-life is 27.5–28.2 hours. Amphotericin B Cholesterol Sulfate Complex
In patients with normal renal function before treatment, the initial plasma half-life after intravenous administration of conventional amphotericin B is approximately 24 hours. After 24 hours, the clearance of amphotericin B decreases, and its elimination half-life has been reported to be approximately 15 days.
Elimination half-life: Neonates: Large individual variability (range: 18 to 62.5 hours). Children: Large individual variability (range: 5.5 to 40.3 hours). Adults: Approximately 24 hours. Terminal half-life: Approximately 15 days. Note: There is significant individual variability in the elimination of amphotericin B in neonates. Amphotericin B may persist in neonates for up to 17 days after discontinuation. Based on a series of sacrifice experiments in rats following a single administration of 3.2 mg/kg aerosol amphotericin B, the elimination half-life of amphotericin B from the lungs of rats was 4.8 days.

Effects During Pregnancy and Lactation
◉ Overview of use during lactation: While there is no information on the excretion of amphotericin B in breast milk, it has a high protein binding rate, a large molecular weight, is almost not absorbed orally, and has been administered directly to infants; therefore, most reviewers believe it is safe for use by lactating women. ◉ Effects on breastfed infants: No relevant published information was found as of the revision date. ◉ Effects on lactation and breast milk: No relevant published information was found as of the revision date. Protein binding Highly bound to plasma proteins (>90%). Interactions Because nephrotoxicity may be additive, the simultaneous or sequential use of amphotericin B and other drugs with similar toxic potential (e.g., aminoglycosides, capreomycin, colistin B, cisplatin, cyclosporine, methoxyflurane, pentamicillin, polymyxin B, vancomycin) should be avoided whenever possible.
It has been reported that corticosteroids may exacerbate potassium loss caused by amphotericin B and therefore should not be used. Amphotericin B should not be treated concurrently with antineoplastic drugs (e.g., nitrogen mustard) unless to control adverse reactions. Antineoplastic drugs may increase the risk of nephrotoxicity, bronchospasm, and hypotension in patients receiving amphotericin B treatment; therefore, such combination therapy should be used with caution. In a randomized, double-blind study, researchers evaluated the use of routine intravenous amphotericin B and amphotericin B cholesterol sulfate complex in patients with febrile neutropenia and normal baseline serum creatinine levels. Results showed that the incidence of nephrotoxicity in adult and pediatric patients receiving amphotericin B cholesterol sulfate complex in combination with cyclosporine or tacrolimus was 31% (defined as a doubling or increase of 1 mg/dL or more in serum creatinine from baseline, or a decrease of 50% or more in calculated creatinine clearance from baseline), compared to 68% in patients not receiving amphotericin B cholesterol sulfate complex in combination with cyclosporine or tacrolimus. In patients receiving standard amphotericin B treatment, the incidence of nephrotoxicity was 8% in adults and children not receiving cyclosporine or tacrolimus who received amphotericin B cholesterol sulfate complex treatment, compared to 35% in those receiving standard amphotericin B treatment. For more complete data on interactions of amphotericin B (15 in total), please visit the HSDB record page. Non-human toxicity values: Intravenous LD50 in mice: 4 mg/kg; Intraperitoneal LD50 in mice: 88 mg/kg

[1]. The antifungal drug amphotericin B promotes inflammatory cytokine release by a Toll-like receptor- and CD14-dependent mechanism. J Biol Chem. 2003 Sep 26;278(39):37561-8. Epub 2003 Jul 14.

[2]. The effect of aggregation state of amphotericin-B on its interactions with cholesterol- or ergosterol-containing phosphatidylcholine monolayers. Chem Phys Lipids. 1997 Feb 28;85(2):145-55.

[3]. Amphotericin B kills unicellular leishmanias by forming aqueous pores permeable to small cations and anions. J Membr Biol. 1996 Jul;152(1):65-75.

[4]. Pharmacological studies of a new derivative of amphotericin B, MS-8209, in mouse and hamster scrapie. J Gen Virol. 1994 Sep;75 (Pt 9):2499-503.

[5]. Amphotericin B encapsulated in micelles based on poly(ethylene oxide)-block-poly(L-amino acid) derivatives exerts reduced in vitro hemolysis but maintains potent in vivo antifungal activity. Biomacromolecules. 2003 May-Jun;4(3):750-7.

Therapeutic Uses
Amphotericin B; Antibiotic, Antifungal; Antibiotic, Macrolide; Antibiotic for Animals
Drug: Antifungal; (Veterinary): Antifungal
Drug (Veterinary): ... Blastomycosis, Histoplasmosis.
Amphotericin B for injection may be used as an adjunct therapy for the treatment of paracoccidioidomycosis caused by Paracocytosporum brasiliensis. /Not included in the US product label/
For more complete data on the therapeutic uses of amphotericin B (19 in total), please visit the HSDB record page.

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Drug Warnings Patients receiving amphotericin B have reported rashes (including maculopapular or vesicular rashes), purpura, pruritus, urticaria, sweating, exfoliative dermatitis, erythema multiforme, alopecia, dry skin, and skin discoloration or ulceration.
Intravenous administration of conventional amphotericin B, amphotericin B cholesterol sulfate complex, amphotericin B lipid complex, or amphotericin B liposomes may cause erythema, pain, or inflammation at the injection site. There have been reports of phlebitis or thrombophlebitis occurring with conventional intravenous amphotericin B. Manufacturers of conventional intravenous amphotericin B and some clinicians recommend adding 500-1000 units of heparin to the amphotericin B infusion solution, using a pediatric scalp vein needle, or administering the drug every other day to potentially reduce the incidence of thrombophlebitis. Extravasation can cause local irritation.
The incidence of adverse reactions to conventional intravenous amphotericin B is relatively high, and most patients receiving this drug will experience potentially serious adverse reactions during treatment. Acute infusion reactions (e.g., fever, chills, headache, nausea, vomiting) and nephrotoxicity are the most common adverse reactions to conventional intravenous amphotericin B. Although clinical experience with amphotericin B cholesterol sulfate complex, amphotericin B lipid complex, and amphotericin B liposomes is currently limited, these drugs appear to be better tolerated than conventional intravenous amphotericin B. As with conventional intravenous amphotericin B, the most common adverse reaction to amphotericin B cholesterol sulfate complex, amphotericin B lipid complex, or amphotericin B liposomes is acute infusion reaction; however, accumulated data to date suggest that liposomal and liposomal amphotericin B formulations may have a lower overall incidence of adverse reactions and a lower incidence of hematologic and nephrotoxicity compared to conventional formulations. Acute infusion reactions may occur within 1–3 hours after intravenous infusion of conventional amphotericin B, amphotericin B cholesterol sulfate, amphotericin B lipid complex, or amphotericin B liposomes, including fever, chills, hypotension, anorexia, nausea, vomiting, headache, dyspnea, and tachypnea. These reactions are most severe and frequent at the initial dose and usually decrease with increasing doses. Fever (with or without chills) may occur within 15–20 minutes after intravenous infusion of conventional amphotericin B. In patients receiving routine intravenous amphotericin B treatment, most (50–90%) experience some degree of intolerance to the initial dose, even at lower initial doses. Although these reactions occur less frequently with subsequent or every-other-day administration, they recur if routine intravenous amphotericin B treatment is interrupted and then restarted. For more complete data on drug warnings for amphotericin B (18 in total), please visit the HSDB record page. Pharmacodynamics: Amphotericin B exhibits high in vitro activity against a variety of fungi. Histoplasma capsulatum, Coccidioides immitis, Candida species, Blastomyces dermatitidis, Rhodotorula, Cryptococcus neoformans, Sporothrix schenckii, Mucor mucedo, and Aspergillus fumigatus can all be inhibited in vitro by amphotericin B at concentrations ranging from 0.03 to 1.0 mcg/mL. While Candida albicans is generally highly susceptible to amphotericin B, other non-Candida strains may be less susceptible. Pseudallescheria boydii and Fusarium sp. may also be susceptible to amphotericin B. They are generally resistant to amphotericin B. This antibiotic is ineffective against bacteria, rickettsiae, and viruses.

923.487

C, 61.09; H, 7.96; N, 1.52; O, 29.43

1397-89-3

Amphotericin B trihydrate;1202017-46-6;Amphotericin B-13C6

5280965

Light yellow to yellow solid

1.3±0.1 g/cm3

1140.4±65.0 °C at 760 mmHg

>170°C

643.5±34.3 °C

0.0±0.6 mmHg at 25°C

1.614

1.16

12

18

3

65

1670

19

C[C@H]1/C=C/C=C/C=C/C=C/C=C/C=C/C=C/[C@@H](C[C@H]2[C@@H]([C@H](C[C@](O2)(C[C@H](C[C@H]([C@@H](CC[C@H](C[C@H](CC(=O)O[C@H]([C@@H]([C@@H]1O)C)C)O)O)O)O)O)O)O)C(=O)O)O[C@H]3[C@H]([C@H]([C@@H]([C@H](O3)C)O)N)O

APKFDSVGJQXUKY-INPOYWNPSA-N

InChI=1S/C47H73NO17/c1-27-17-15-13-11-9-7-5-6-8-10-12-14-16-18-34(64-46-44(58)41(48)43(57)30(4)63-46)24-38-40(45(59)60)37(54)26-47(61,65-38)25-33(51)22-36(53)35(52)20-19-31(49)21-32(50)23-39(55)62-29(3)28(2)42(27)56/h5-18,27-38,40-44,46,49-54,56-58,61H,19-26,48H2,1-4H3,(H,59,60)/b6-5+,9-7+,10-8+,13-11+,14-12+,17-15+,18-16+/t27-,28-,29-,30+,31+,32+,33-,34-,35+,36+,37-,38-,40+,41-,42+,43+,44-,46-,47+/m0/s1

(1R,3S,5R,6R,9R,11R,15S,16R,17R,18S,19E,21E,23E,25E,27E,29E,31E,33R,35S,36R,37S)-33-(((2R,3S,4S,5S,6R)-4-amino-3,5-dihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)-1,3,5,6,9,11,17,37-octahydroxy-15,16,18-trimethyl-13-oxo-14,39-dioxabicyclo[33.3.1]nonatriaconta-19,21,23,25,27,29,31-heptaene-36-carboxylic acid

Amphotericin B;NSC 527017;Ambisome NSC527017;Amphozone FungilinFungizoneAMPH-B Fungizone Liposomal Amphotericin B NSC-527017

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

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

DMSO :~50 mg/mL (~54.11 mM)

Solubility in Formulation 1: 10 mg/mL (10.82 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with heating and sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 100.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: 10 mg/mL (10.82 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with heating and sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 100.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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