Synthesis And Characterization Of Miltefosine Derivatives And Photolabeling Probes Of Miltefosine

Miltefosine (HePC, hexadecylphosphocholine), an alkyllysophosphocholine, shows remarkable cytostatic and cytotoxic activity and has been approved as an anticancer drug for topical treatment of certain forms of cutaneous and breast cancers. Recently, miltefosine acquires increasing interest as a new antileishmanial drug and has been registered for oral treatment of visceral leishmaniasis in India, with cure rate reaching to 97%.However, due to its gastrointestinal toxicity and its haemolytic side effects, the application of miltefosine is limited to topical treatment. So it is very important to design and synthesize new generations of miltefosine as more efficient and safer drugs. For this purpose, we designed and synthesized a series of miltefosine analogues.The alkyl chains of alkylphosphocholine are partly responsible for the physical and chemical properties of the drugs. We have synthesized a series of long chain alcohol with one or more methylene groups substituted with oxygen atoms as intermediates of miltefosine derivatives.We have synthesized 12-(propoxy)dodecylphosphocholine and 12-(ethoxy) dodecylphosphocholine, analogues of hexadecylphosphocholine. Due to the introduction of the ether bond, the hydrophilicity of miltefosine has been increased, which make it easier for miltefosine to permeate the membrane and maybe reduce the required effective concentration.Results from studies of structure-activity relationships of miltefosine showed that the phosphate moiety is not always necessary for cytostatic action. Some non-phosphorus alkyl ether glycerolipids also exhibite cytostatic properties. Thus, we have synthesized a series of miltefosine derivatives, in which the phosphate moity isreplaced by carbonate. These compounds lack the negatively charged phosphate and can resist to catabolic phospholipases, therefore they will have improved biological stability, and better membrane permeability.Although miltefosine has gained clinical success, little is known about the mechanism of action in either tumor cell or parasites. Due to its amphiphilic properties, miltefosine primarily interact with cellular membranes in cancer cells as well as in protozoa. Many hypotheses have been proposed to explain the anticancer activity of miltefosine, there is still no definitive mechanism. The mechanisms proposed so far include modulating membrane permeability and membrane lipid composition, influencing phospholipids metabolism, alternating proliferation signal transduction pathway, and inducing apoptosis.In order to study the molecular mechanisms responsible for the biological activity of miltefosine, we propose the photoaffinity labeling approach. Photoaffinity labeling is an efficient method of studying the interactions between drugs and their biological targets. It allows us to identify the targets of drugs, to determine the affinity and selectivity of the drug-target interaction, and to identify the binding sites on the targets.We designed and synthesized two photolabeling probes of miltefosine. Probe 1 has the fluorinated arylazido group at the polar head of miltefosine, while probe 2 has the fluorinated arylazido group at the alkyl chain. Photochemical studies of these two probes showed that both the probes undergo a fast, clean process of photodecomposition upon irradiation at >300nm. Therefore, probe 1 and 2 can both constitute promising tools for studying the mechanisms underlying the biological effects of miltefosine using the photolabeling approach.