Liposome Formulations of Hydrophobic Drugs.

Here we report methods of preparation for liposome formulations containing lipophilic drugs. In contrast to the encapsulation of water soluble compounds into the entrapped aqueous volume of a liposome, drugs with lipophilic properties are incorporated into the phospholipid bilayer membrane. Water-soluble molecules, for example cytotoxic or antiviral nucleosides can be transformed into lipophilic compounds by attachment of long alkyl chains, allowing their stable incorporation into liposome membranes and taking advantage of the high loading capacity lipid bilayers provide for lipophilic molecules.

Cytostatic activity of a 5-fluoro-2'-deoxyuridine-alendronate conjugate against gastric adenocarcinoma and non-malignant intestinal and fibroblast cell lines.

Background: 5-Fluoro-2'-deoxyuridine (5-FdU), a drug against gastric cancer, was covalently linked via its nucleobase with the amino-bisphosphonate alendronate (Ale), resulting in a new antimetabolite-bisphosphonate conjugate (5-FdU-Ale), designed for bone-targeting.

Materials And Methods: The cytostatic effect of 5-FdU-Ale was evaluated in vitro compared to monomers and mixtures using CASY Technologies and the human gastric adenocarcinoma cell lines 23132/87 and MKN-45, in comparison to the intestinal CCL-241 and dermal fibroblast NHDF neonatal cell lines.

Results: The adenocarcinoma cell lines demonstrated a slightly higher sensitivity, with respect to the cell lines CCL-241 and NHDF, to incubation with 5-FdU-Ale.

N⁴-[Alkyl-(hydroxyphosphono)phosphonate]-cytidine-new drugs covalently linking antimetabolites (5-FdU, araU or AZT) with bone-targeting bisphosphonates (alendronate or pamidronate).

Amino-bisphosphonates (alendronate, pamidronate) were covalently linked in a three step synthesis, with protected and triazolylated derivatives of therapeutically used nucleoside analogs (5-FdU, araC, AZT) by substitution of their triazolyl residue. From the deprotected and chromatographically purified reaction mixtures N⁴-[alkyl-(hydroxyphosphono) phosphonate]-cytidine combining two differently cytotoxic functions were obtained. This new family of bisphosphonates (BPs) contains as novelty an alkyl side chain with a cytotoxic nucleoside.

Cytostatic activity of the duplex drug linking 2'-deoxy-5-fluorouridine (5FdU) with 3'-C-ethynylcytidine (ECyd) against gastric adenocarcinoma cell lines.

The cytostatic potential of the new duplex drug 2'-deoxy-5-fluorouridylyl-(5'5')-3'-C-ethynylcytidine (5FdU(5'-5')ECyd) was evaluated in comparison to those of 5-fluorouracil (5FU), 2'-deoxy-5-fluorourindine (5FdU), 3'-C-ethynylycytidine (ECyd), cisplatin, an equimolar mixture of 5FdU + ECyd and a three component-mixture of 0.75 μM epirubicin/0.90 μM cisplatin/3.

Liposome formulations of hydrophobic drugs.

Here, we report methods of preparation for liposome formulations containing lipophilic drugs. In contrast to the encapsulation of water-soluble compounds into the entrapped aqueous volume of a liposome, drugs with lipophilic properties are incorporated into the phospholipid bilayer membrane. Water-soluble molecules, for example, cytotoxic or antiviral nucleosides can be transformed into lipophilic compounds by attachment of long alkyl chains, allowing their stable incorporation into liposome membranes and taking advantage of the high loading capacity lipid bilayers provide for lipophilic molecules.

Cellular pharmacology of multi- and duplex drugs consisting of ethynylcytidine and 5-fluoro-2'-deoxyuridine.

Prodrugs can have the advantage over parent drugs in increased activation and cellular uptake. The multidrug ETC-L-FdUrd and the duplex drug ETC-FdUrd are composed of two different monophosphate-nucleosides, 5-fluoro-2'deoxyuridine (FdUrd) and ethynylcytidine (ETC), coupled via a glycerolipid or phosphodiester, respectively. The aim of the study was to determine cytotoxicity levels and mode of drug cleavage.

Synthesis and in vitro activities of new anticancer duplex drugs linking 2'-deoxy-5-fluorouridine (5-FdU) with 3'-C-ethynylcytidine (ECyd) via a phosphodiester bonding.

Two isomeric cytostatic duplex drugs 2'-deoxy-5-fluorouridylyl-(3'-->5')-3'-C-ethynylcytidine [5-FdU(3'-->5')ECyd] and 2'-deoxy-5-fluorouridylyl-(5'-->5')-3'-C-ethynylcytidine [5-FdU(5'-->5')ECyd] were designed and synthesized at gram scale according to the hydrogenphosphonate method in an overall yield of about 40%. The in vitro evaluation of the anticancer effects indicated highly varying sensibilities of the panel of 60 tested tumor cell lines against the duplex drugs. 5-FdU(3'-->5')ECyd had a 50% growth inhibition (IC(50) < or = 10(-8) M) in 44/58 cell lines.

Synthesis and in vitro activities of a new antiviral duplex drug linking Zidovudine (AZT) and Foscarnet (PFA) via an octadecylglycerol residue.

To prepare a new antiviral duplex drug linking Zidovudine (AZT) and Foscarnet (PFA) via a lipophilic octadecylglycerol residue we condensed 1-O-4-monomethoxytrityl-3-O-octadecyl-sn-glycerol-2-hydrogenphosphonate obtained from 3-O-octadecyl-sn-glycerol with AZT by the phosphonate method. The purified condensation product was de-tritylated resulting in 3'-azido-3'-deoxythymidylyl-(5'-->2-O)-3-O-octadecyl-sn-glycerol, followed by treatment with (ethoxycarbonyl)phosphoric dichloride. The resulting 3'-azido-3'-deoxy-thymidylyl-(5'-->2)-3-O-octadecyl-sn-glycerol-1-O-(ethoxycarbonyl)phosphonate was purified by preparative RP-18 column chromatography.

In vivo and in vitro activity and mechanism of action of the multidrug cytarabine-L-glycerylyl-fluorodeoxyuridine.

Multidrugs have the potential to bypass resistance. We investigated the in vitro activity and resistance circumvention of the multidrug cytarabine-L-fluorodeoxyuridine (AraC-L-5FdU), linked via a glycerophospholipid linkage. Cytotoxicity was determined using sensitive (A2780, FM3A/0) and resistant (AG6000, AraC resistant, deoxycytidine kinase deficient; FM3A/TK-, 5FdU resistant, thymidine kinase deficient) cell lines.

5-FdUrd-araC heterodinucleoside re-establishes sensitivity in 5-FdUrd- and AraC-resistant MCF-7 breast cancer cells overexpressing ErbB2.

ErbB2 overexpressing breast tumors have a poor prognosis and a high risk to develop chemoresistance to therapeutic treatment. "Chemoresistance" is a response of cells to toxic stress, and, although it is a common phenomenon, it is still poorly defined. However, a detailed understanding is required to target desensitized pathways and mechanisms for successful reactivation as part of a tailored therapy.

Isolation and characterization of a scFv antibody specific for tumor endothelial marker 1 (TEM1), a new reagent for targeted tumor therapy.

Tumor endothelial marker 1 (TEM1) is a protein predominantly expressed on the cell surface of endothelial cells in newly developing blood vessels and on tumor cells. It is therefore ideally suited as a target for anti-angiogenic tumor therapy. Using phage display technology a single chain antibody fragment (scFv-CM6) was isolated that specifically binds to the extracellular part of TEM1.

Cytotoxic effects of novel amphiphilic dimers consisting of 5-fluorodeoxyuridine and arabinofuranosylcytosine in cross-resistant H9 human lymphoma cells.

Various amphiphilic heterodinucleoside phosphates have recently been synthesized in order to overcome drug resistance. These agents contain 5-fluorodeoxyuridine (5-FdUrd) and arabinofuranosylcytosine (Ara-C). We now investigated the action of two of these novel dimers (#2 and #10) in sensitive and 5-FdUrd/Ara-C cross-resistant H9 human lymphoma cells.

Synthesis and anticancer activities of amphiphilic 5-fluoro-2'-deoxyuridylic acid prodrugs.

Amphiphilic anticancer prodrugs of 5'-fluoro-2'-deoxyuridine-5'-monophosphate (5-FdUMP) were synthesized according to the hydrogen phosphonate method by coupling lipophilic cytosine derivatives or a phospholipid with 5-fluoro-2'-deoxyuridine (5-FdU). Studies within the in vitro Anticancer Screen Program of the National Cancer Institute have demonstrated high anticancer activities of the heterodinucleoside phosphates: N4-palmitoyl-2'-deoxycytidylyl-(3' --> 5')-3'-O-acetyl-5-fluoro-2'-deoxyuridine (dC(pam)-5-FdU(Ac), N4-palmitoyl-2',3'-dideoxycytidylyl-(5' --> 5')-3'-O-acetyl-5-fluoro-2'-deoxyuridine (ddC(pam)-(5' --> 5')-5-FdU(Ac), 5-fluoro-2'-deoxyuridylyl-(3' --> 5')-5-fluoro-N4-hexadecyl-2'-deoxycytidine (5-FdU-5-FdC(hex)), and of the new liponucleotide 1-O-octadecyl-rac-glycerylyl-(3 --> 5')-5-fluoro-2'-deoxyuridine (Oct1Gro-(3 --> 5')-5-FdU). The anticancer activities of these prodrugs are comparable to those of 5-FdU and the tumor specificities are modulated by their structures.

Heterodinucleoside phosphates of 5-fluorodeoxyuridine and arabinofuranosylcytosine--new drugs in cancer chemotherapy?

The incidence of cancer is rapidly increasing and malignancies have become the number two cause of deaths in the Western world after cardiovascular diseases. In particular, colon cancer represents one of the most frequent types of malignancy. Chemotherapy is, in addition to surgery and irradiation, still one of the main treatment options against this group of diseases.

Lipophilic arabinofuranosyl cytosine derivatives in liposomes.

Highly lipophilic drugs can be used therapeutically only by the addition of possibly toxic solubilizing agents or by development of complex pharmaceutical formulations. One way of overcoming these disadvantages is the incorporation of such drugs into the bilayer matrix of phospholipid liposomes. To this end, we chose the approach of chemical transformation of water-soluble nucleosides of known cytotoxic properties into lipophilic drugs or prodrugs.

In vitro and in vivo antitumor activity of novel amphiphilic dimers consisting of 5-fluorodeoxyuridine and arabinofuranosylcytosine.

Various heterodinucleoside phosphates of 5-fluorodeoxyuridine (5-FdUrd) and arabinofuranosylcytosine (Ara-C) have recently been synthesized as potent chemotherapeutic agents. 5-Fluorodeoxyuridine is being used in patients with colorectal carcinoma, whereas Ara-C is one of the most effective agents in the treatment of hematological malignancies. We now investigated the action of three novel amphiphilic dimers with different structures in various 5-fluorouracil (5-FU) sensitive and resistant human colon tumor cell lines (CCL228, CCL227, 5-FU resistant CCL227 and HT-29) as well as in L1210 murine leukemia cells.

Combination effects of Ara-C and 5-fluorodeoxyuridine against leukemia cells in vitro and in mice.

Background: Ara-C (1-beta-D-arabinofuranosylcytosine) is widely used for treatment of human leukemia. However, due to emergence of resistance, new drug combinations need to be developed.

Materials And Methods: We tested the combination of Ara-C with 5-FdUrd (5-fluorodeoxyuridine) in L1210 and P388D1 mouse leukemia cells in vitro and in vivo by growth inhibition and recovery assay in leukemia-bearing mice.