Miltefosine (Impavido; HePC)

Akt; PI3K; PKC (IC50 = ~7 μM)

Miltefosine is an alkylphosphocholine medication with demonstrated activity against various parasite species and cancer cells as well as some pathogenic bacteria and fungi. Miltefosine inhibits PKC from NIH3T3 cells in cell-free extracts with a IC50 of about 7 μM.[1] In vivo, macrophages that are HIV-infected serve as long-lived HIV-1 reservoirs, and miltefosine targets these cells. By blocking the PI3K/Akt pathway, miltefosine eliminates infected macrophages from circulation without harming healthy cells.[2] In carcinoma cell lines, miltefosine inhibits the PI3K/Akt survival pathway.[3] Miltefosine interferes with the insulin signaling pathway and prevents insulin-stimulated glucose uptake, which results in skeletal muscle insulin resistance in vitro. With 75% inhibition at 40 M and 98% inhibition at 60 μM, miltefosine inhibits insulin-stimulated Akt phosphorylation in a dose-dependent manner.[4]

Miltefosine inhibits anti-IgE induced histamine release from human skin mast cells. Miltefosine can significantly slow down the esterification of cholesterol as well as lower levels of the cytokines IL-1β, IL-4, and IL-6 in some skin tissue cells. [5]

The ApoAlert Caspase Fluorescent assay kit is used to measure the amounts of enzymatically active caspase-3. In a nutshell, 1 106 BC-1 PEL cells are exposed to vehicle controls, 50 M Miltefosine, 50 M Perifosine, or 20 nM NVP-BEZ235. After 12 hours, cells are collected and lysed. For each sample, an identical amount of cell lysate is incubated with a fluorogenic caspase-3 substrate (DEVD-AFC). With the excitation and emission filter wavelengths set to 400 and 505 nm, respectively, cleavage of DEVD by caspase-3 releases AFC, the fluorescence of which is measured using a FLUOstar OPTIMA fluorometer.

2 × 105 PEL cells are either treated with the therapeutic substances at the recommended doses or with the appropriate vehicle as a negative control. Trypan blue exclusion is performed in quadruplicate to assess cell viability after 96 hours of cell monitoring.

Mice: PEL cells are collected, counted, and diluted in 100 L of PBS combined with 100 L of Matrigel depleted of growth factors after being washed in ice-cold phosphate buffered saline. Subcutaneous injection of 1 105 to 7.5 105 BC-1 cells is made into the right flank of NOD. Alternatively, CB17-Prkdcscid/J mice. On alternate days, the mice are checked for the development of palpable tumors (2 mm3). If this occurs, drug or vehicle treatments are started, and the mice receive either intraperitoneal (Perifosine) or oral gavage (Rosiglitazone, NVP-BEZ235) treatments 5 days a week. PEL tumors are created using groups of 5–7 mice, and either a vehicle or drug cocktail is used to treat them. Multiple replications of every biological experiment are carried out. 30 mg/kg or 60 mg/kg of Rosiglitazone is suspended in 0.25% methylcellulose, which serves as the vehicle for the medication. PBS serves as a vehicle for the drugs Perifosine and Miltefosine, which are dissolved in the solution at a concentration of 50 mg/kg each. In order to dissolve NVP-BEZ235, the substance is combined with polyethylene glycol 300 in a 1:9 vol/vol ratio of 1-methyl-2-pyrrolidone. A dose of 40 mg/kg NVP-BEZ235 or an equivalent volume of the vehicle is given. Digital calipers are used to measure the tumor diameters, and tumor volume is computed. The tumors are removed and then fixed in formalin. With each animal treated as a random effect, statistical analyses are carried out using a linear model fit with the maximum likelihood.
Rats: There are five groups of male Sprague-Dawley rats (n=5), each weighing between 270 and 290 g. Miltefosine (MFS) is given to rats in the treatment groups as a single oral dose of 10 mg/kg as either an aqueous solution or MFS-LNCs dispersion by gastric gavage. This dosage, adjusted for rats, is equivalent to the 20 mg/kg Miltefosine dose given to mice in the preclinical study. Following administration, blood samples are taken through the orbital plexus while the patient is under anesthesia at intervals of 0.5, 1, 2, 4, 7, 10, 24, 48, 72, and 216 hours. The Eppendorf tubes contain EDTA. The next step is an immediate, 10-minute centrifugation of blood samples at 4000 rpm. While awaiting analysis, plasma samples are kept frozen and at -80°C.

[1]. Cancer Res. 1991 Feb 1;51(3):807-12.

[2]. Retrovirology. 2008 Jan 31;5:11.

[3]. Anticancer Drugs. 2003 Feb;14(2):167-73.

C21H46NO4P

407.57

407.3165

C, 61.89; H, 11.38; N, 3.44; O, 15.70; P, 7.60

58066-85-6

58066-85-6

Solid powder

O=P(OCCCCCCCCCCCCCCCC)([O-])OCC[N+](C)(C)C

PQLXHQMOHUQAKB-UHFFFAOYSA-N

InChI=1S/C21H46NO4P/c1-5-6-7-8-9-10-11-12-13-14-15-16-17-18-20-25-27(23,24)26-21-19-22(2,3)4/h5-21H2,1-4H3

hexadecyl (2-(trimethylammonio)ethyl) phosphate

HePC; Hexadecyl phosphocholine; Miltefosin C; HePC; Hexadecylphosphocholine; HDPC; Hexadecylphosphorylcholine; Miltefosinum; mpavido; Miltex; Choline Phosphate Hexadecyl Ester Hydroxide Inner Salt; hexadecylphosphocholine; Miltefosin; Miltefosina; Miltefosinum

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)

Solubility in Formulation 1: 100 mg/mL (245.36 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.

---

Solubility in Formulation 2: Saline: 30mg/mL

---

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