The present invention relates to a group of new compounds of the general formula I:
Nu-O-Fa
wherein O represents an oxygen, Nu is a nucleoside or nucleoside analogue, and Fa is an acyl group of a mono-unsaturated C18 or C20 fatty acid. The invention also concerns anti-viral pharmaceutical and veterinary compositions comprising a compound of formula I alone or in combination with a pharmaceutically acceptable carrier. A further part of this invention is a method for the treatment of a human or animal patient suffering from a viral infection and for reducing the infectious load by administering a compound of formula I.
A large number of serious diseases, such as AIDS, hepatitis B, herpes and gynecological cancer, as a late result of papilloma warts, are caused by viral infections.
Viruses are small infectious agents which are incapable of independent replication and thus are dependent on a host cell to replicate. The genetic material of the virus is either RNA or DNA.
When infecting an organism, the virus attaches to a specific host cell. The virus penetrates the cytoplasmic membrane after attachment and the viral genome is released from the virus particle. The viral genome is usually transported to the cell nucleus where new viral genomes are replicated. New viral protein is synthesized in the cytoplasm and new particles are formed either close to the cytoplasmic or nuclear membrane.
Some viruses have genomic material which is directly (DNA virus) or indirectly (Reverse transcription of RNA, retrovirus) incorporated in the host cell genomes.
Extracellular viruses are neutralized by circulating antibodies and the cellular immune apparatus may attack and remove infected cells. Viruses within the infected cells escape immune surveillance if viral antigens are not exposed on the surface of the cells.
The immune attack on infected organs contributes to disease by a mechanism commonly called virus induced immuno-pathology.
The mechanisms underlying some of the more important viral diseases differ.
When suffering from a HIV infection, the patient""s T helper cells are invaded and destroyed. This leads to a immunodeficiency condition, which makes the patient very succeptible even to infections which normally are conquered by the immune system without any harmful effects for the patient.
The Hepatitis B virus invades the liver cells, and the patient may become very ill when the immune system tries to rid the body of these infected cells. If the infection is not conquered by the immune system at an early stage, the result will be chronic hepatitis. The patient will thus be infectious throughout his life. For a group of patients the chronic hepatitis will develop into cirrhosis or cancer of the liver.
In herpes simplex infections, the virus enters the epidermal cells originally. The herpes simplex virus travels up to a nerve center where it lies latent to break out at intervals. Although not life threatening in most cases, a herpes infection is painful and the patient will be infectious every time an outbreak occurs.
In the papilloma virus, notably in the genital tract of women, the viral genome is located in the nucleus of epithelial cells, but not integrated in the cell chromosomes. This is a persistent condition and with some tumor promoting strains an integration finally occurs leading to a malignant development. The viral genome in this case has a decisive initiating effect in the process leading to cancer.
If the immune system manages to rid the body of the virus at an early stage, this leads to a life long immunity. On the other hand, if the virus is too aggressive and avoids the immune apparatus, no immunity is achieved and a continuous infectious state is the result.
As a result of the different mechanisms, the therapeutic strategy would be different for these conditions.
The ultimate goal in the treatment of HIV/AIDS would be to free the patient from the infectious virus. This seems to be remote at the present stage. However, much can be obtained by improving the general condition of the patient. A reduction of the virus load would increase the length of the symptom free period and reduce the infectiousness, which is of utmost importance in regard to the epidemiological situation. All currently used anti-viral agents have toxic side effects, which presently makes a sufficiently aggressive treatment impossible.
It is assumed that there are between 250 and 300 million carriers of hepatitis B world wide. It is known that a great number of these are going to develop hepatomas or liver insufficiencies due to the infections. Promising results in the treatment of the carrier state have been obtained in recent years by induction of an immune response with interferon. Therapies reducing the virus load are important in this regimen as efficient treatment of acute hepatitis B would reduce the number developing into a carrier state. The recently identified hepatitis C virus causes a very great number of cases with hepatitis whereof a large number develop into carriers. Preliminary studies seem to indicate that the carrier state may be broken by similar therapeutic regimens as for hepatitis B. Herpes simplex 1 and 2 frequently infect humans causing a carrier state with recurrencies of local infections. Generalized infections including encephalitis are rare but a catastrophy for the patient. There is a great individual variation in the frequency of local infections. For those patients who are affected either genitally or facially this constitutes a serious health problem physically, mentally and socially. None of the therapeutic regimens developed so far cures the latent infections of cells in the central nervous system. The therapeutic goal is thus to minimize the clinical manifestations of recurrencies both as to symptoms and duration.
The prevalence of genital papilloma virus infections has increased dramatically during the 1980s. It is now established that some genotypes are oncogenic, that is they initiate changes in the cell which after a latency period develop into cancer. Papilloma virus of the genital tract give long standing infections. The factors causing malignant transformation of the lesions are not well understood, but the immune system is assumed to be of importance. It is thought that the lesions showing progression during the months and years are those giving rise to cancer. The genital papillomas called condylomas are at present treated by physical means such as surgical removal, necrotizing means, fluid nitrogen or the like. Genital warts are at the onset benign tumors with altered enzyme patterns affecting among other things the metabolism of nucleoside analogues. Nucleoside prodrugs affect the episomal proliferation of papilloma virus thereby inducing regression of the warts.
Prophylactic vaccination has been very successful in acute infections such as polio, measles, mumps etc., but no effective vaccination has been developed for many of the other serious viral infections.
Even though there have been intensive efforts to produce effective anti-viral chemotherapeutica during the last decades, no satisfactory medical treatment can be offered for most viral diseases today. The efforts have been especially great since the appearance of the HIV and related viral infections, which are spreading throughout the world at an alarming rate, yet the effects obtained with agents such as azidothymidine (AZT) and acyclovir (ACV) in AIDS and herpes can only be characterized as partially successful. These most promising anti-viral agents are thus derivatives of naturally occurring nucleosides, which have been modified either in the base or sugar moiety. They have, however, not had the therapeutic potential hoped for, as they bring forward serious side-effects in some patients or show little or no effect in others. Further, treatment with these agents is extremely expensive. For these reasons only patients suffering from the very serious viral infections such as AIDS receive such treatment. Patients suffering from the less serious, but also very painful viral infections are often left without any treatment to let the infection take its own course.
The untreated patient carries a great infectious load and constitutes a risk to his fellow human beings. If he is treated with an anti-viral agent, the aim is to reduce the infectious load so as to enable the body""s immune system to conquer the infection. A further aim is to reduce the contagiousness and thus the number of new patients and carriers.
Thus the need for compounds having a better therapeutic index is obvious.
The need is especially great in chronic or recurrent viral infections with a dangerous acute phase or long term ill effects on health or well being, such as AIDS, hepatitis B and C, infections of the herpes group and papilloma viral infections. Similarily, there is also a need for anti-viral agents usable in the treatment of animals suffering from viral diseases.
In order to improve the effect, there have been developed derivatives of the nuclosides which are either modified in the base or sugar moiety. Especially fatty acid esters of the nucleosides or of analogues of nucleosides have been developed, in order to improve the lipophility and achieve a better membrane passage.
Thus there are known from EP 56265 (Lxc3x6bering et al.) esters of arabino-furanosyl-thymine (Ara T) with saturated acids having 1-17 C-atoms.
From PCT/WO90/00555 (Hostetler et al.) there are known lipid derivatives linked, epecially through a phosphate group, to the 5xe2x80x2-position of the pentose group of a nucleoside. The purpose of this derivatization was to make the nucleosides more lipophilic so that they could be included into liposomes, which are preferentially taken up by macrophages and monocytes, cells which are found to harbor the HIV virus. It is stated that a targetting effect is thereby achieved.
From EP 393920 there are known unsaturated, preferably polyunsaturated, C16 or higher fatty acid esters or amides of nucleoside or nucleoside analogues. It is stated that the fatty acid portion of these molecules preferably are made up of poly-unsaturated fatty acids, such as xcex3-linolenic or linoleic acid.
It has now surprisingly been found that a selected group of fatty acid, esters of anti-viral nucleosides or nucleoside analogues, wherein the fatty acid is a mono-unsaturated C18 or C20 acid gives a much improved effect. Also the compounds according to the present invention seem to be relatively nontoxic as judged by growth experiments with young mice and also have a low cytotoxicity according to observations made in tissue culture experiments.
Although it is known that both nucleosides and nucleoside analogues, by themselves, and also some unsaturated fatty acids, by themselves, exhibit anti-viral effects, the magnitude of the effects achieved with the compounds according to this invention indicates that this is not an additive, but rather a synergistic activity which is special for the compounds of formula I.
The mechanism behind these effects is at present not known, but they are of an order of magnitude better than the closest related compounds of the prior art. Thus it is not considered likely that they arise only due to a membrane effect or targetting effect.
Further it is also clear, as will appear from the biological examples included herein, that effects are achieved with these compounds in systems where no effects may be achieved with the mother nucleoside compound.
The compounds of this invention can be characterized by the general formula I:
Nu-O-Fa
wherein O is an oxygen, Nu is a nucleoside or nucleoside analogue and Fa is an acyl group of a mono-unsaturated C18 or C20 fatty acid.
Nucleosides are molecules comprising a heterocyclic base, such as cytosine, uracil, adenine or guanine, linked to a ribose unit. In nucleoside analogues, either the base or the ribose unit has been modified. For example, the ribose unit may be replaced by another sugar unit or by a noncyclic chain.
The fatty acid is esterified with a hydroxyl group in the 5-position of the sugar moiety of the nucleoside or with a hydroxyl group on the non-cyclic group of the nucleoside analogue.
The nucleoside or nucleoside analogues which may be chosen as Nu in the compounds of formula I may preferably be represented by the formula II:
Sxe2x80x94Bxe2x80x83xe2x80x83(II)
wherein S is either a mono-saccharide-derivative selected from: 1-xcex2-D-arabinofuranose or 2,3-di-deoxy-3-azido-1-xcex2-D-ribofuranose, or selected from the group of 2-hydroxy-ethoxy-methyl, 4-hydroxy-3-(hydroxymethyl)-butyl, 2-hydroxy-1-(hydroxy-methyl)-ethoxy-methyl or 2,3-di-hydroxy-propoxy; and B is a nitrogen base selected from adenine, guanine, cytosine, uracil, thymine or a thymine derivative of the following formula: 
Wherein X is deuterium or fluorine.
Examples of these nucleosides or nucleoside analogues are: 
Examples of the group R in different nucleoside analogues appear below: 
There exist several systems for denomination of the position of double bonds in fatty acids. In the present application the xcfx89-system is used, wherein the position of the double bond in the unsaturated fatty acids is counted from the terminal methyl group. Eicosenic acid (C20:1 xcfx89-9), for example, has 20 carbon atoms in the chain, and the double bond is found between carbon atom 9 and 10 counting from the end of the chain.
The fatty acids which are reacted with the nucleosides or nucleoside analogues to form the esters according to this invention with the pronounced activity are the xcfx89-9 C18 or C20 monounsaturated fatty acids. Further, even though the effect observed differs somewhat between acids of the same chain length when the double bond is in cis or trans configuration, both show a strong activity. Also the position of the double bond is of importance, as it appears that especially the esters with a C18 or C20 fatty acid having the unsaturation in xcfx89-9 position will have the surprisingly elevated activity.
The C18 or C20 xcfx89-9 fatty acids, which when bound to the nucleosides give the surprisingly elevated effect, are the following: oleic acid (C18:1, xcfx89-9, cis), elaidic acid (C18:1, xcfx89-9, trans) eicosenic acid, (c20:1, xcfx89-9, cis) and (C20:1, xcfx89-9, trans)
Preferred representatives of the compounds according to this invention are listed below.: Ara A-oleic acid, Ara A-elaidic acid, Ara A-eicosenic acid, cis, Ara A-eicosenic acid, trans, Ara T-oleic acid, Ara T-elaidic acid, Ara T-eicosenic acid, cis, Ara T-eicosenic acid, trans, ACV-oleic acid, ACV-elaidic acid, ACV-eicosenic acid, cis, ACV-eicosenic acid, trans, AZT-oleic acid, AZT-elaidic acid, AZT-eicosenic acid, cis, AZT-eicosenic acid, trans. Their formulas will appear from FIG. 4.
The compounds according to this invention exhibit anti-viral effects, and the present invention thus includes pharmaceutical or veterinary compositions comprising at least one compound of formula I alone or in combination with a pharmaceutically acceptable carrier or excipient. In the remainder of the text and in the claims, a pharmaceutical composition will be used for compositions usable in the treatment of both human and animal patients.
Further, it appears that certain of the monounsaturated fatty acid nucleosides or nucleoside analogues will be especially suitable for the treatment of certain viral infections. Thus it appears that the fatty acid derivatives of AZT are especially useful for the treatment of AIDS.
Similarly, it appears that the fatty acid derivatives of Ara T and Ara A are especially suitable for the treatment of hepatitis B. It appears also that the Ara T esters will be efficient in agents suitable for treatment of papilloma viral infections.
Further, it appears that the fatty acid esters according to this invention of ACV are especially suitable for the treatment of herpes infections.
As mentioned, the production of the necessary immune response in order to conquer a viral infection, such as hepatitis, can be induced in some cases by the co-administration of interferon.
Depending on which viral infection is to be treated and at what stage the infection is, or if the patient is a human being or an animal, both a systemic and a local administration of the compounds can take place.
For local administration, the compounds can be formulated as known in the art for administration to the skin or mucosa in any suitable form.
When administered topically the compounds of formula I may be formulated as an ointment, cream, gel, tincture, spray, lotion or the like containing the compounds of formula I in admixture with inert, solid or liquid carriers which are usual in topical preparations. It is especially suitable to use a formulation which protects the active ingredient against oxidation or degradation.
The pharmaceutical preparations comprising the compounds of formula I may also be administered systemically, either enterally or parenterally.
When administered enterally, the compounds of formula I may be formulated e.g. as soft or hard gelatine capsules, tablets, granules, grains or powders, dragees, syrups, suspensions or solutions.
When administered parenterally, preparations of the compounds of formula I as injection or infusion solutions, suspensions or emulsions are suitable.
The preparations can contain inert or pharmacodynamically active additives. Tablets or granulates e.g. can contain a series of binding agents, filler materials, carrier substances or diluents. Liquid preparations may be present, for example, in the form of a sterile solution. Capsules can contain a filler material or thickening agent in addition to the active ingredient. Furthermore, flavor-improving additives as well as the substances usually used as preserving, stabilizing, moisture-retaining and emulsifying agents, salts for varying the osmotic pressure, buffers and other additives may also be present.
The dosages in which the preparations according to this invention are administered will vary according to the mode of use and the route of use, as well as to the requirements of the patient. In general a daily dosage for a systemic therapy for an adult average patient will be about 0.1-100 mg/kg body weight/day, preferably 1-20 mg/kg/day. For topical administration, the suitable ointment can contain from 0.1-10% by weight of the pharmaceutical formulation, especially 0.5-5% by weight.
If desired, the pharmaceutical preparation of the compound of formula I can contain an antioxidant, e.g. tocopherol, N-methyl-tocopheramine, butylated hydroxyanisole, ascorbic acid or butylated hydroxytoluene.
The invention further discloses a method for the treatment of viral infections, which comprises administering at least one compound of formula I to a human or animal patient in need of such treatment.
Further, the invention also comprises a method for the treatment of a patient in need of such treatment with a combination of a compound of formula I and an interferon.