Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Reexamination Certificate
2001-09-21
2003-04-29
Wilson, James O. (Department: 1623)
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
C536S026710, C536S026720, C536S026740, C536S026800, C536S124000, C536S027140, C536S027210, C536S027800, C536S027810, C536S028100, C536S028200, C536S028500
Reexamination Certificate
active
06555676
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to the bioreversible functionalization of phosphate or phosphonate groups of biologically active compounds.
The present invention relates more particularly to phosphotriester-type biologically active compounds bearing phosphate or phosphonate groups which are protected by protecting groups that are bioreversible in an intracellular medium.
BACKGROUND OF THE INVENTION
Compounds bearing a phosphate or phosphonate group have a negatively charged ionic nature and a physiological pH. As a result, the therapeutic activity of such compounds is limited by the low diffusion of negatively charged compounds across biological lipid membranes. Moreover, compounds bearing phosphate groups are readily dephosphorylated by the action of phosphatase enzymes in the blood or on cell membranes, which enzymes dephosphorylate substrate compounds. In general, charged phosphate or phosphonate compounds are poorly absorbed via oral administration, and do not diffuse efficiently across cell membranes or even the cerebral barrier, which are lipidic in nature.
Certain compounds, such as nucleoside derivatives or analogs, are active agents that are administered in non-phosphorylated form, but are phosphorylated in vivo in the form of metabolic monophosphate or triphosphate to become active.
Thus, nucleoside derivatives having antitumor activity, such as 5-fluorouridine, 5-fluoro-2′-deoxyuridine or 1-O-D-arabinofuranosylcytosine, exert their activity in phosphorylated form.
Similarly, in order to exert their antiproliferative activity, certain nucleoside or phosphononucleoside analogs need to be phosphorylated into the corresponding triphosphate thereof by cellular or viral enzymes; this triphosphate is then capable of inhibiting the viral and/or cellular polymerases.
Among the various structural classes of antiviral agents, 2′, 3′-dideoxynucleosides are among the most effective compounds in the treatment of AIDS. However, these nucleoside analogs must undergo a biotransformation by cell kinases in order to exert their activity on the replication of HIV, the etiological agent of AIDS. This metabolization occurs via the dideoxynucleoside 5′-monophosphate and then the 5′-diphosphate to lead to the 5′-triphosphate, which is an inhibitor of HIV reverse transcriptase and which thereby interferes with the biosynthesis of viral DNA.
Despite their great therapeutic potential, 2′,3′-dideoxynucleosides suffer from limitations, in particular the low metabolizability of some of them by kinases into triphosphate. 2′,3′-Dideoxyuridine 5′-triphosphate, for example, is an excellent inhibitor of reverse transcriptase (Z. Hao et al., Proc. Am., Assoc. Cancer Res., 1988, 29, 348, E. Matthes et al., Biochem. Biophys. Res. Commun, 1987, 148, 78-85). However, the nucleoside thereof is able to inhibit the replication of HIV in vitro. Studies have shown that this result is linked to the low metabolizability of the nucleoside into its monophosphate by cell kinases (Z. Hao et al. Mol. Pharmacol. 1990, 37, 157-153).
Thus, AZT is successively metabolized into the triphosphate thereof (AZTP), which is a potent inhibitor of HIV reverse transcriptase. Similarly, Acyclovir (ACV) is converted into the triphosphate thereof (ACVTP) which selectively inhibits herpesvirus DNA polymerase. The first step in the activation of the nucleosides (Nu) consists of a monophosphorylation, leading to the corresponding monophosphate (NUMP). It is this first step which is the most selective.
In order to circumvent this key step of enzymatic monophosphorylation, it has already been proposed to administer NuMPs directly, but their use for therapeutic purposes was contraried by the abovementioned limitations and drawbacks.
Compounds bearing a phosphate or phosphonate group have a negatively charged ionic nature at physiological pH. The therapeutic activity of such compounds is consequently limited, on account of the low diffusion of negatively charged compounds across biological lipid membranes. In particular, charged compounds do not diffuse efficiently across cell membranes, or indeed across the cerebral barrier, which are lipidic in nature. Moreover, such compounds are readily dephosphorylated by the action of phosphatase enzymes in the blood or on the cell membranes, which enzymes dephosphorylate the substrate compounds thereof. In general, charged phosphate or phosphonate compounds are poorly absorbed via oral administration.
It has been sought to convert mononucleotides into neutral phosphotriesters capable of crossing the cell membrane and of intracellular delivery of the corresponding mononucleotide phosphotriester (NUMP). Such an approach has been adopted by various authors for a number of years, but has proved to be disappointing. The derivatives obtained were in general either excessively toxic or of insufficient extracellular stability, and did not in the end result provide any enhancement of the biological activity.
Thus, the use of phosphorylated nucleoside structures comprising bioreversible protecting groups of acyloxymethyl or acyloxybenzyl type has been proposed, for antitumor nucleoside derivatives such as 5-fluorouracil, in WO patents No. 9,008,155 and 9,119,721. However, these compounds are of limited chemical stability, and generate toxic formaldehyde metabolites in vivo. Furthermore., they are sparingly soluble and the yield of their chemical preparation is low.
The aim of the present invention is thus to provide other types of bioreversible groups which may be combined especially with mononucleotide or other structures such that the biological activity thereof is enhanced, in particular as regards compounds derived from or analogous to nucleosides having antiviral activity, and which reversible groups do not have the abovementioned drawbacks.
The present invention proposes to use novel groups, characterized by the presence of —SIS— and/or —S/C═Z enzymelabile bonds which lead, after enzymatic activation, to the formation of unstable intermediates that selectively release the corresponding monophosphate or monophosphonate.
More precisely, the subject of the present invention is the compound corresponding to the general formula I:
RO—P(═O)(OR)—Nu (I)
in which:
R is a radical —(CH
2
)n—S—X, where X represents a radical —C(═Z)(Y) or —S—U, and Z is O or S;
Y and U represent an alkyl, aryl or saccharide radical which is optionally substituted, in particular with an OH, SH or NH group; and
n is equal to 1 to 4, preferably 1 or 2;
Nu is a radical consisting of a residue of a biologically active compound or the dephosphorylated residue of a compound which is biologically active when it bears a phosphate or phosphonate group.
Moreover, the present invention also relates to the compound corresponding to the general formula Ia:
RS—P(═O)(QR)—Nu (Ia)
in which:
R is a radical —(CH
2
)n—W—X, where X represents a radical —C(═Z)(Y) or —S—U, and Z is O or S;
Q is O or S;
W is O or S;
Y and U represent an alkyl, aryl or saccharide radical which is optionally substituted, in particular with an OH, SH or NH group;
n is equal to 1 to 4, preferably 1 or 2; and
Nu is a radical consisting of a residue of a biologically active compound or the dephosphorylated residue of a compound which is biologically active when it bears a phosphate or phosphonate group.
When, in the formulas (I) and (Ia), Nu is linked to the phosphorus by a P—O bond, the compound of formulas (I) and (Ia) according to the invention bears a phosphate group and thus constitutes a phosphotriester compound.
When Nu is linked to the phosphorus by a P—C bond, the compound of formulas (I) and (Ia) according to the invention bears a phosphonate group.
The mechanisms of bioreversibility of the radicals R take place via enzymatic cleavage of the S—X or O—X bonds and release of the (CH
2
)
2
—S residues, according to the mechanisms which are illustrated by the examples represented FIG.
1
and FIG.
9
.
For Y and U there are especially mentio
Gosselin Gilles
Imbach Jean-Louis
Perigaud Christian
Centre National de la Recherche Scientifique
Lewis Patrick
Wilson James O.
Woodcock & Washburn LLP
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