CZ298510B6 - Drug delivery device for transdermal administration of a therapeutic agent - Google Patents

Abstract

In the present invention, there is disclosed a drug delivery device for transdermal administration of a therapeutic agent, comprising: a backing; a permeable membrane suitable for placement against the skin of an animal to be treated; and an adsorbent situated between the backing and the permeable membrane, the adsorbent having adsorbed thereon a therapeutic composition comprising a therapeutic agent which is N,N'-dimethylformamide.

Description

Transdermal patch for transdermal administration of a therapeutic agent

Technical field

The present invention relates to a transdermal patch for transdermal administration of a therapeutic agent consisting of polar substances such as Ν, Ν'-dimethylformamide (DMF) for the treatment of animals affected by viral or microbial infections, in particular retroviral infections such as HIV or weight loss syndrome, in particular HIV infection or malignancy.

BACKGROUND OF THE INVENTION

Human immunodeficiency virus

Human immunodeficiency virus (HIV) induces a persistent and progressive infection leading in most cases to the development of acquired immunodeficiency syndrome (AIDS) (Barre-Sinoussi et al., 1983, Science 220: 868-870, Gallo et al., 1984, Science 224: 500-503). There are at least two different types of HTV: HIV-1 (Barre-Sinoussi et al., 1983, Science 220: 868-870, Gallo et al., 1984, Science 224: 500-503) and H1V-2 (Clavel et al. , 1986, Science 223: 343-346, Guyader et al., 1987, Nature 326: 662-669). In humans, HIV replication noticeably occurs in CD4 ⁺ T-lymphocyte populations and HIV infection destroys this cell type and eventually leads to immune deficiency, opportunistic infections, neurological dysfunctions, neoplastic growth, and ultimately death.

HIV belongs to the lentivirus retroviral family (Teich et al., 1984, RNA Tumor Viruses, Weiss et al., Eds., CSH-press, pages 949-956). Retroviruses are small enveloped viruses that contain single-stranded RNA in the genome and replicate via a DNA intermediate consisting of a virus encoded by reverse transcriptase RNA-dependent DNA polymerase (Varmus, H., 1988, Science 240: 1427-1439). Other retroviruses include, for example, oncogenic viruses such as human T-cell leukemia virus (HTLV-1, -II, -III) and feline leukemia virus.

The HIV viral particle consists of a viral nucleus consisting in part of the p24 and p18 labeled capsid proteins together with the viral RNA genome and those enzymes that are essential for early replication processes. The myristylated gag protein forms an outer viral shell around the viral nucleus, which for change is surrounded by a lipid membrane envelope derived from an infected cell membrane. HIV envelope surface glycoproteins are synthesized as a single 160 kDa precursor protein that is cleaved by cellular proteases during the germination of the virus into two glycoproteins, gp41 and gp120. gp41 is a transmembrane glycoprotein and gp120 is an extracellular glycoprotein that remains non-covalently associated with gp41, probably in a trimeric or multimeric form (Hammerskjold, M., A Rekosh, D., 1989, Biochem. Biophys. Acta 989: 269-280).

HIV, like other enveloped viruses, introduces genetic material into the host cell via a viral enveloped-mediated fusion of the viral and target membranes. HIV targets CD4 + cells because the CD4 cell surface protein (CD4) acts as a cell receptor for the HIV virus (Dalgleish et al., 1984, Nature 312: 767-768), Maddon et al., 1986, Cell 47: 333- 348). Viral entry into the cell is dependent on the binding of gp120 to the cellular CD4 receptor (Pal et al., 1993, Virology 194: 833-837, McDougal et al., 1986, Science 231: 382-385), Maddon et al., 1986, Cell 47: 333-348), which explains the tropism of the virus to CD4 cells, while gp41 anchors the envelope glycoprotein complex in the viral membrane. Binding of gp120 to CD4 induces conformational changes in viral glycoproteins, but this binding alone is not sufficient to lead to infection (reviewed in Sattentau and Moore, 1993, Philos. Trans. R. Soc. London (Biol.) 342 : 5966).

Studies of HIV isolates have revealed heterogeneity in their ability to infect various human cell types (reviewed in Miedema et al., 1994, Immunol. Rev. 140: 35-72). Most extensively passaged laboratory strains of HIV clearly infect cultured T-cell lines and primary T-lymphocytes

But not primary monocytes or macrophages. These strains are called T-tropical. HIV-infected individuals are more likely to find T-tropic HIV strains during the late stages of AIDS (Weiss et al., 1996, Science 272: 1885-1886). Most primary HIV isolates (i.e., viruses that have not been extensively passaged in cultures) replicate effectively in primary lymphocytes, monocytes, and macrophages, but poorly grow on established T cell lines. These isolates were designated as M-tropical. The viral determinants of T- and M-tropism map to the third variable region of gp120 (V3 loop) (Choe et al., 1996, Cell 85: 1135-1148, Cheng-Mayer et al., 1991, J. Virol. 65: 6931 -6941, Hwang et al., 1991, Science 253: 71-74, Kim et al., 1995, J. Virol., 69: 1755-1761), and O'Brien et al., 1990, Nature 348: 69 -73). Characterization of HIV isolates of different tropism, together with the observation that binding to CD4 cell surface protein alone is not sufficient to lead to infection, suggest that cell-specific cofactors might be required for CD4 to enter HIV into host cells.

Treatment of HIV infection

HIV infection is pandemic and HIV-associated diseases are a major global health problem. Although considerable efforts are being made to develop effective therapeutics, there is currently no curative anti-retroviral therapy against AIDS. In attempts to develop such drugs, several stages of the HIV life cycle have been considered as possible targets for therapeutic intervention (Mitsuya, H. et al., 1991, FASEB J. 5: 2369-2381). Many viral targets have been proposed for affecting the HIV life cycle, although the prevailing view is that affecting host cell proteins would have deleterious side effects. For example, virus-encoded reverse transcriptase was one of the focuses of drug development. Many drugs directed to reverse transcriptase have been developed, including 2 ', 3'-dideoxynucleoside analogs such as AZT, dd1, ddC and d4T, and have been shown to be active against HIV infection (Mitsuya, H. et al., 1991, Science 249: 1533-1544.

A new treatment regimen for HIV infection shows that a combination of anti-HIV drugs that target reverse transcriptase (RT) such as azidothymidine (AZT), lemivudine (3TC), dideoxyinosine (ddl), dideoxycitidine (ddC), used in combination inhibitors HIV proteases have a greater effect (decrease by 2 to 3 logs) on the amount of virus compared to AZT alone (decrease by about 1 log). For example, recently, admirable results have been obtained with a combination of AZT, ddl, 3TC and ritonavir (Perelson, AS et al., 1996, Science 15: 1582-1586). However, it is likely that prolonged use of a combination of these chemicals will lead to toxicity, especially bone marrow. Long-term cytotoxic therapy may also lead to suppression of CD8 ⁺ cells, which are necessary for HIV control through cell killing activity (Blazevic, V. et al., 1995, AIDS Res. Hum. Retroviruses 11: 1335-1342) and through release of suppressive factors. , especially the chemokines Rantes, ΜΙΡ-Ια, and MIP1β (Cocchi, FA et al., 1995, Science 270: 1811-1815). Another problem with long-term chemical anti-retroviral therapy is the development of partial or complete resistance HIV mutations (Lange, JM, 1995, AIDS Res. Hum. Retroviruses 10: S77-82). Such mutations are believed to be an inevitable consequence of anti-viral therapy. The nature of the disappearance of wild-type viruses and the emergence of a mutated virus as a result of treatment, combined with a coincidence decrease in CD4 + cell counts, strongly suggest that at least some agents appear to be the major underlying factor in AIDS therapy failure.

There are also attempts to develop drugs that can inhibit the entry of virus into cells, the earliest stage of viral infection. So far, the main focus has been on CD4, the cell surface receptor for HIV. For example, the recombinant soluble CD4 molecule showed inhibition of CD4 ⁺ T cell infection by some HIV-1 strains (Smith, DH et al., 1987, Science 238: 1704-1707). However, certain primary HIV isolates are relatively less sensitive to inhibition by the recombinant CD4 molecule (Daar, E. et al., 1990, Proc. Natl. Acad. Sci. USA 87: 6574-6579). In addition, clinical trials with recombinant soluble CD4 have not given conclusive results (Scholey, R. et al., 1990, Ann. Int. Med. 112: 247-253, Kahn, JO et al., 1990, Ann. Int. Med. 112 : 254261), Yarchoan, R. et al., 1989, Proc. Vth Int. Conf. On AIDS, p. 564, MCP 137).

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Late stages of HIV replication, which include key virus-specific processes of certain virus-encoded proteins, have also been suggested as potential targets for anti-HIV drugs. Late stage processes are dependent on viral protease activity, and drugs that inhibit this protease are being developed (Erickson, J., 1990, Science 249: 527-533).

Recently, chemokines of CD8 + formation by T cells have been included in the inhibition of HIV infection (Paul, W. E. 1994, Cell, 82: 177, Bolognesi, D. P., 1993, Semin. Immunol. 5: 203).

The chemokines Rantes, MIP-1α and MIP-β, which are secreted by CD8 + T cells, have been shown to suppress the production of HIV p24 antigen in cells infected with HIV-1 or HIV-2 isolates in vitro (Cocchi, F. et al. 1995, Science 270: 1811-1815).

Therefore, these and other cytokines could be useful in the treatment of HIV infection. However, the clinical outcome is a question for all these and other candidate drugs.

Attention is also paid to the development of vaccines for the treatment of HIV infection. HIV coat proteins (gp10, gp120, gp41) have been shown to be major antigens for anti-HIV antibodies present in AIDS patients (Barin et al., 1985, Science 228: 1094-1096). Therefore, by this time, these proteins seem to be the most promising candidates to act as antigens for the development of anti-HIV vaccines. Several groups have begun to use different portions of gp110, gp120 and / or gp41 as immunogenic targets for the host immune system. See, for example, Ivanoff, L. et al., US 5,141,867; Saith, G. et al., WO 92/22 654; Shafferman, A., WO 91/09872; Formoso, C. et al., WO 90/07 119. To date, vaccines targeting HIV proteins have been problematic in that viruses mutate rapidly, rendering many of these vaccines ineffective. However, the clinical results of these candidate vaccines are still far away.

Thus, although there is a very strong effort to develop and test anti-retroviral drugs, effective, non-toxic treatment is still needed.

Weight loss syndrome

Weight loss syndrome is a serious clinical problem characterized by a weight loss of more than 10% compared to baseline body weight and an unbalanced weight loss in relation to body fat (Weinroth et al., 1995, Infectious Agents and Disease 4: 76-94, Kotler and Grunfeld, 1995, AIDS Clin. Rev. 96: 229-275). Therefore, this weight loss is distinguished from starvation, where more body fat is lost than body cell weight (Kotler et al., 1985, Am. J. Clin. Nutr. 42: 1255-1265, Cahill, 1970, N. Engl. J. Med., 282: 668-675). Weight loss is associated with a variety of conditions including HIV infection, other infectious diseases, sepsis, cancer, chronic cardiovascular disease and diarrhea (Kotler et al., 1989, Am. J. Clin. Nutr. 50: 444-447; Heymsfield et al. , 1982, Am J Clin Nutr. 36: 680-690). Importantly, weight loss is an important factor in mortality in patients suffering from infections or cancer. In fact, weight loss has a linear relationship to survival time in AIDS patients (Kotler et al., 1989, Am. J. Clin. Nutr. 50: 444-447).

The cause of weight loss syndrome in AIDS and other conditions is unclear and most likely multifactorial. Metabolic abnormalities, irregular hormone and cytokine levels, and absorption disorders were all involved in weight loss syndrome. Not all AIDS patients suffer from weight loss, which I assume is not the cause of weight loss alone. Most cases of HIV-related weight loss syndromes appear to be due to complications of AIDS, such as secondary infections and gastrointestinal diseases (Kotler and Grundfeld, 1995, AIDS Clin. Rev. 96: 229-275).

Current and potential therapies for weight loss syndrome include nutritional support, appetite enhancers such as dronabinol megestrol acetate, anabolic therapies such as growth hormones and cytokine inhibitors. However, mixed results were obtained with nutritional support and appetite enhancement in those patients who tended to accumulate only fat and not total body mass. The administration of growth hormones and cytokine inhibitors is still being tested and may present a risk of side effects (Kotler and Grundfeld, 1995, AIDS Clin. Rev. 96: 229-275, Weinroth et al., 1995, Infectious Agents and Disease 4: 76-94) .

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Therefore, weight loss treatment is critical to the survival and well-being of patients suffering from serious diseases such as cancer and AIDS. Therefore, there is a need for safe and effective therapy for weight loss syndrome associated with cancer, AIDS, and other infectious diseases.

Properties of dimethylformamide

Ν, Ν'-dimethylformamide (DMF) (molecular formula: C3H7ON) is a colorless, polar, hygroscopic liquid with low volatility and boiling point 152.5 to 153.5 ° C. It is freely miscible with water, alcohols and some hydrocarbons. DMF is generally used as a polar solvent and is readily absorbed through the skin, by inhalation or after oral administration. DMF is rapidly metabolized, mainly in the liver, and excretion occurs mostly in the urine. In rats, mice, hamsters and humans, the major metabolites of DMF are N-hydroxymethyl-N-methylformamide (HMMF), N-methylformamide (NMF) and N-acetyl-S- (N-methylcarbamoyl) cysteine (AMCC) as well as dihydroxymethylformamide (DHMF) and N-hydroxymethylformamide (HMF). Unchanged DMF is excreted in the urine as a small fraction of the administered dose of DMF.

DMF has low acute dermal, oral and inhalation toxicity. It is considered to be a moderate irritant to the skin and eyes and easily penetrates the skin. There is no indication that it has skin sensitizing properties. The major toxic effect of DMF and its metabolites is on the liver; It is well known that DMF causes reversible liver damage associated with typical clinical complications, classical biochemical changes in blood, and the appearance of hepatocyte necrosis in other biopsies. DMF is teratogenic but is not considered to be a mutagen or carcinogen.

VISA et al. have reported that DMF and DMSO inhibit the in vitro replication of HIV and human herpesvirus 6 (HHV-6) in certain cultured cell lines (see Visa et al., 1990 AIDS Res. Hum. Retroviruses 6: 131-132, Visa et al. , 1989 AIDS-FORSCHUNG 4: 349-352, Viza et al., 1992, Antiviral res. 18: 27-38 and erratum at 19: 179).

DMF has been described as an in vitro differentiation agent for certain transformed cells in culture (see Koeffler, 1983, Blood 62: 709-721, Calabresi et al., 1979, Biochem. Pharmacol. 28: 1933-1941). When added to certain malignant cells in vitro, DMF has been reported to reduce their tumorigenicity following subsequent inoculation into nude mice (see Dexter, 1977, Cancer Res. 37: 3136-3140, Dexter et al., 1979, Cancer Res. 39: 1020-1025. After intraperitoneal injection into nude mice, DMF and NMF have been shown to slow the growth of certain human tumor xenografts (see Van Dongen et al., 1989, Int. J. Cancer 43: 285-292, Braakhuis et al., 1989, Head & Neck. 11: 511-515, Van Dongen et al., 1988, Acta Otolaryngol. 105: 488-193; Dexter et al., 1982, Cancer Res. 42: 5018-5022). However, the toxic side effects of formamide and its N-methyl derivatives in the mouse sarcoma allograite model led researchers to conclude that these substances are unlikely to prove therapeutically effective (see Clarke et al., 1953, Proc. Exp. Biol. Med 84: 203-207). Attempts to treat human tumor patients with DMSO led to the conclusion that no objective response was achieved (see Spremulli and Dexter, 1984, J. Clin. Oncol. 2: 227-241). Oral administration of NMF to patients with head and neck cancer has been reported to result in hepatotoxicity without a beneficial response (see Vogel et al., 1987, Invest. New Drugs 5: 203-206) or with minimal activity (see Planting et al. , 1987, Cancer Treat Rep. 71: 1293-1294).

US 3,551,154 discloses the use of DMF as a penetration enhancer to promote transdermal absorption of topically applied medicinal products. US 4,855,294 discloses the use of glycerin to alleviate skin irritations induced by the use of DMSO and DMF as penetration enhancers to promote transdermal absorption of topically applied medicinal products. The use of DMSO as a penetration enhancer to promote transdermal absorption of antiviral agents is discussed in Woodford and Barry, 1986, J. Toxicol. Cut. & Ocular Toxicol. 5: 167-177.

Citation or identification of any references in this paragraph (or any other paragraph) of this disclosure will not be construed as an admission that such reference is available as prior art to the invention.

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SUMMARY OF THE INVENTION

The transdermal patch for transdermal administration of a therapeutic agent of the invention comprises an external support portion; a permeable membrane suitable for placement against the skin of the animal to be treated; and an adsorbent disposed between the outer support portion and the permeable membrane, the adsorbent having adsorbed therapeutics comprising N, N'-dimethylformamide.

In accordance with the invention, an animal in need of treatment is administered transdermally a compound consisting of Ν, Ν'-dimethylformamide (DMF).

The therapeutic regimen in an HIV-infected patient may advantageously combine a compound of the invention with one or more other agents effective in the treatment of HIV infection, including but not limited to selected from the group consisting of nucleoside analogue reverse transcriptase inhibitors, non-nucleoside inhibitors reverse transcriptase and protease inhibitors in any desired combination.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention may be more clearly understood by reference to the following detailed description of the invention, examples of specific embodiments of the invention, and the accompanying drawings in which:

Figure 1 illustrates plasma DMF concentration as a function of time in 4 patients treated with 2 transdermal patches for 8 hours.

Figure 2 illustrates plasma DMF concentration as a function of time in 3 patients treated with 2 transdermal patches for 6 hours.

Figure 3 shows the viral load of HIV-1 measured by quantitative PCR in patients treated with transdermal DMF. Two DMF patches were placed on the forearm skin for 12 hours on days 0.8 and 13 (indicated by arrows).

Figure 4 shows the overall status of HIV-infected patients before and after treatment with transdermal DMF as assessed by the Kamofsky performance scale. See more details in Example 2.

Detailed description of the invention

The present invention provides a transdermal patch for drug delivery for the treatment of viral and microbial infections. In one embodiment, the infection intended to treat a retrovirus infection such as HIV, including an asymptomatic infection, a latent infection, and an infection accompanied by one or more AIDS-related symptoms and an infection accompanying clinical AIDS. Alternatively, the infection to be treated is any other viral or microbial infection, including Rubella infection, herpesviruses such as Human Herpesvirus 6, Epstein-Barr virus or cytomegalovirus, infection with any virus having a capsule protecting the skin, and any opportunistic infection associated with immune system disease, such as is an opportunistic infection in HIV infected patients.

The present invention also provides a transdermal patch for drug delivery for the treatment or prevention of any disease or disease characterized by weight loss. Particular conditions that can be treated include, but are not limited to, weight loss associated with viral (eg HIV), bacterial or any other infection or sepsis, cachexia associated with malignant disease, chemotherapy or radiation therapy, weight loss associated with chronic cardiovascular disease, weight loss due to exposure of the patient to toxic or radioactive substances, and weight loss associated with diarrhea or other gastrointestinal diseases.

The subject to be treated may be any animal including, but not limited to, monkeys, cows, sheep, cattle, pigs, horses, cats, dogs, chicken, and others, and is preferably a

Even more preferably a primate, and most preferably an adult human or child, e.g., a human child weighing less than 3 kg. Here the term "patient" is used for any animal in need of treatment.

According to the present invention, a transdermal patch is administered with a therapeutic compound consisting of DMF.

The therapeutic regimen in an HIV-infected animal may advantageously include, in addition to DMF, one or more agents effective in the treatment of HIV infection, eg, one or more nucleoside analogue reverse transcriptase inhibitors such as zidovudine (AZT, ZDV), zalcitabine (ddC), didanosine. (ddl), lamivudine (3TC), stavudine (d4T), one or more non-nucleoside reverse transcriptase inhibitors such as nevirapine, delavirdine, loviride, atevirdine, pyridinone, one or more protease inhibitors such as saquinavir, indinavir, ritoonavir, nelfinavir, or any a combination of the above or other anti-HIV therapeutic agents. DMF and other anti-HIV therapeutic agent (s) may be administered simultaneously, sequentially or in treatment cycles according to any desired treatment protocol.

The polar compound DMF is applied to the skin using transdermal patches. The transdermal patch may optimally consist of an outer support portion, a reservoir such as an adsorbent impregnated with a polar compound of the invention and a permeable membrane that is placed in contact with the skin. The support portion may be any material, such as a natural or synthetic polymer, that resists the chemical action of the polar material. Especially preferred is a high density polyethylene support portion. The adsorbent may be a colloidal substance, such as diatomaceous earth or colloidal silica. The permeable membrane may be of any material that chemically resists the polar material and may optionally be supplied with pores. In a preferred embodiment, the Teflon ™ permeable membrane has pores having a diameter of about 0.1 µm, a diameter of about 0.5 µm, or a diameter in the range of about 0.1 to 0.5 µm. The patch may be self-adhesive or may be held in contact with the skin by an applicator such as a wrap or bandage, including, without limitation, an elastic or adhesive bandage; Elatoplast bandage is suitable for this purpose. Preferably, the patch contains a greater amount of a polar substance than is intended to be delivered through the skin of the patient to be treated; for example, the patch may contain about 50% more polar material than is intended to be delivered. The patch may be of any size and shape, and may, for example, be in the form of a disk having a diameter of approximately 9 cm.

In one embodiment, the transdermal patch comprises DMF and at least one other pharmacologically active agent, eg, an anti-irritant agent such as glycerin. In another embodiment, the patch comprises DMF and no other pharmacologically active agent is present. In another embodiment, the patch comprises DMF and no other pharmacologically active agent is present that is capable of being systemically absorbed through the skin or no pharmacologically active agent is present in an amount that is systemically effective after transdermal absorption. In yet another embodiment, the patch comprises DMF and no other antiviral agent is present, eg, acyclovir or arildone.

Thus, the present invention provides a transdermal patch comprising DMF in an amount effective to treat a viral or microbial infection (eg, HIV infection) in human adult or pediatric patients. The present invention further provides a transdermal patch comprising DMF in an amount that is effective in treating weight loss syndrome in human adult or pediatric patients. In one embodiment, the patch comprises at least 0.25 g DMF, preferably at least 0.5 g, even more preferably at least 1 g, and even more preferably at least 5 g, eg, 5 to 15 g DMF.

In order to minimize the loss of polar material by evaporation, the patch may conveniently be stored in a sealed package such as a sealed polymer bag. If desired, each patch can be individually sealed for convenient use. Advantageously, the patch may be cooled prior to use, e.g. to 4 ° C, to reduce the loss of polar material by evaporation. Preferably, the patches are prepared within 24 hours prior to use and are stored in a sealed package at 4 ° C; however, the patches are stable for one or more weeks when stored at 4 ° C in a sealed package. Before applying the patch, it is advantageous to wash the skin with mild soap and water, rinse,

To remove the soap residue, dry thoroughly and then hydrate with a suitable skin lubricant and moisturizer such as KY ™ jelly. The patch is then removed from the backing material and applied so that the permeable membrane is in contact with the prepared skin surface. The patch may be held in place by the applicator. After the required dosing period, the patch is removed. After removal of the patch, the treatment area may be thoroughly washed with a mild soap to remove residual polar material.

DMF may be administered at any desired interval, for example, every two or three weeks; once, twice or three times a week; every second day; or daily. Preferably, it is administered at a dose that results in a plasma concentration of about 2 to 200 mg / L, more preferably about 100 to 200 mg / L, even more preferably about 150 mg / ml DMF. A plasma level of 100 to 150 mg / l or 150 to 200 mg / l DMF is particularly preferred. As used herein, the term "ppm" refers to a part of a million by weight, and in practice is equivalent to mg / L.

For transdermal administration of a polar substance, the absorption level is determined by the skin of the test subject. After exposure to human skin, DMF is absorbed at a steady rate of approximately 9.4 mg / cm ² / hour (see Mraz and Nohova, Occup. Env. Health 64: 85-92). Similarly, the desired level of absorption can be achieved by controlling the area of the skin area exposed to the drug as well as determining the area of each transdermal patch and the number of transdermal patches applied to the skin. For example, two patches with a diameter of 9 cm will expose the polar substance to a total skin area of 127 cm ² ; for DMF, this will result in an absorption rate of about 1.2 g DMF per hour. A starting dose of 15 mg / kg DMF is particularly preferred.

In one embodiment, a 72 kg patient is initially treated with DMF by administering two transdermal patches 9 cm in diameter for one hour, resulting in an initial absorption dose of about 1.2 g DMF corresponding to a dose of 16.7 mg / kg. The number of transdermal patches may be increased or decreased, and a longer or shorter initial exposure interval may be used, depending on the body weight of the patient being treated. This initial dose may be repeated at any desired interval as described above, and is preferably administered at weekly intervals. Preferably, the patient is monitored for at least 72 hours after DMF administration for signs of toxicity, for example by monitoring serum or plasma levels of enzymes such as aspartate aminotransferase (AST), alanine aminotransferase (ALT), γ-glutamyltransferase (yGT) and alkaline phosphatase, proteins such as albumin. or a substance such as conjugated or unconjugated bilirubin. Preferably, serum or plasma levels of AST and ALT do not exceed five times the upper limit of normal as determined by the reference range of the laboratory and the relevant population; more preferably, serum or plasma levels of AST and ALT do not exceed three times the upper limit of normal, and most preferably serum or plasma levels of AST and ALT are not elevated above normal or above pre-treatment levels. The dose of DMF may be increased by applying the patches for extended periods of time; in one embodiment, the patches are applied for two hours, then four hours, then six hours, etc .; in another embodiment, the exposure time is doubled. Preferably, the patient is monitored prior to any dose escalation to detect any signs of toxicity. If required, the dose may be increased at weekly intervals. The dose may be increased in this manner until the calculated dose is about 150 mg / kg; or until the desired upper plasma level of, for example, 150 to 200 mg / L is reached; or until the exposure time is about six or eight hours. Caution should be exercised before increasing the dose above 240 mg / kg.

DETAILED DESCRIPTION OF THE INVENTION

Example 1: Treatment of HIV infection by transdermal administration of Ν, Ν'-dimethylformamide (DMF)

HIV-1 infected patients were treated with dimethylformamide (DMF) by applying dimethylformamide gel-impregnated skin patches to the patient's body. N-acetyl-cysteine-glutathione and / or necessary phospholipids were administered to the patient (orally or intravenously) at a dose of 250 to

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300 mg a day as a liver function enhancer. Instead or in conjunction with it, glutamine may be administered as a liver function enhancer.

Two skin patches have been applied to different parts of the body, eg the forearm. Each skin patch contained about 7.06 g of DMF gel (92.5% w / w) and colloidal silica (7.5% w / w). The gel was used to prevent leakage of liquid DMF through the patch. The patches were made no more than 12 hours before use, as DMF evaporates rapidly. The intended level of DMF in the patient's blood was 100 ppm. For a patient weighing about 60 kg, an amount of about 14 g is required over a 12 hour period to reach a level of 100 ppm. Based on studies by Marz and Foot, the area to absorb this amount is about 127.2 cm ² . To achieve a blood level of 100 ppm, about 1.272 g of DMF must be absorbed per hour, so each patch must contain about 7.064 g of DMF to deliver the required amount at a surface area of 6.36 cm ² (each patch). The absorption rate of DMF is 9.4 mg / cm ² / hour. Theoretically, this treatment delivers 125 to 135 ppm, but because of the DMF evaporation, 100 ppm has been achieved. Absorption capacity varies from patient to patient depending on factors such as skin type and skin thickness. In order to achieve the desired DMF level in the patient, plasma DMF levels were monitored in each patient and the treatment was adjusted according to the DMF levels in each patient. Giant. 1 shows plasma DMF levels in 4 patients treated with two transdermal DMF patches for 8 hours. Giant. 2 shows plasma DMF levels in 3 patients treated with 2 transdermal patches for 6 hours.

Thus, the patches were impregnated with about 7.064 g of a gel with DMF and silica. Each patch was applied for 12 hours, either weekly for 12 weeks, or twice weekly for 6 weeks.

Blood tests showed an increase in CD4 T-cell counts from 350 to 1000 in certain patients and a rapid reduction in PCR (polymerase chain reaction) (viral load) from 120,000 to 500 / ml in three weeks after only three treatment cycles. Giant. 3 shows a serial quantitative PCR measurement of HIV-1 viral load in a patient before treatment and after three cycles of treatment with two transdermal patches with DMF for 12 hours. The PCR tests performed were a Roche Amplicor HIV monitor. A viral load of less than 500 / ml plasma is considered undetectable.

Some patients undergoing treatment had severe acne or had rubella symptoms prior to treatment. When treated with DMFF so that DMF levels in the patient's blood were 50-100 ppm, rubella and severe acne symptoms disappeared within 7 days.

Prior to dimethylformamide treatment, the patient underwent an overall baseline clinical and psychological evaluation. The evaluation provided basal levels of the patient's biochemical and hermatological data. Detailed virological serological tests (HIV-1) were also performed to determine the total number of viruses in the patient and were performed weekly or by treatment.

The concentration of DMF in the patient's blood was determined every hour during the treatment period. An intravenous cannula was taken every morning to collect blood samples and left open with an infusion of normal saline at a rate of 20 ml per hour, and daily monitoring of the active metabolite AMCC (eg, by urine sampling after 4 hours) derived from DMF was also performed. Subsequent DMF applications were adjusted in accordance with the measured changes in blood DMF levels that resulted from changes in absorption variables, and complete haematological and biochemical profiles were performed daily to detect any changes in liver function. Full clinical and psychological evaluations were also performed daily.

Virological serological calculations were also performed daily to determine the total number of virus in the body and to monitor the improvement in the patient's immune status (CD4 helper T-cells) and in prognostic factors. Serological calculations were performed based on p24 antigens and quantitative PCR or preferably other methods. CD4 counts were also evaluated weekly

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Tests included:

a) serum: Na, K, Cl, CO ₂ , urea, urate, creatinine, Ca, Mg, phosphate, total and conjugated bilirubin

b) hematology: hemoglobin, red blood cell count, hematocrit, MCV, MCH, MCHC, RDW

c) serum protein electrophoresis: total proteins, albumin, total globulin, alpha globulin, alpha2 globulin, beta globulin, gamma globulin

d) white blood cell analysis: white blood cell count, absolute neutrophil count, lymphocyte, monocyte, eosinophil and basophil count

e) liver enzymes: alkaline phosphatase, gamma GT, ALT (SGPT), AST (SGOT), LDH

f) Others: cellular traits, PCR, beta2-macroglobulin, p24 antigen, C-reactive protein, CK-MB concentration

g) Analysis of blood for DMFF levels

h) Urine analysis for AMCC levels

DMF appears to act as at least one of reverse transcriptase inhibitors and protease inhibitors. In vitro assays have been performed and the dissolution properties of DMF appear to break down viral particles, eg capsid.

Example 2: Treatment of HIV infection by transdermal administration of Ν, Ν'-dimethylformamide (DMF)

A preliminary study was conducted to determine the efficacy of transdermal DMF in the treatment of HIV infected patients. Informed consent was obtained from each patient. The seropositive condition was examined by Western blot using a commercial kit for the detection of anti-p24 antibodies (Abbott Diagnostics) and the presence of HIV-1 was documented by quantitative PCR using a commercial kit (Roche Amplicor).

Commercially available DMF (Sigma-Aldrich) and plasma samples were analyzed by mass spectroscopy / gas chromatography (GC / MS), using a Varian 9600 gas chromatograph, an OV351 column, a capillary carbos wax-PEG column, and an ion trapping Finnegan Mat ITS40 detector. The operating parameters were as follows: GC temperature program: 60 ° C for 1 minute followed by a 9.4 ° C / min temperature gradient. for 20 minutes. MS ionization method: electron impact, mass range: 40 to 80 mass units: 1 image / sec, peak threshold: 3 calculations / sec, background mass: 69 mass units. Dimethylacetamide stock solution was used as internal standard. All samples were extracted with an organic solvent containing an internal standard, allowed to precipitate in the refrigerator for 30 minutes, and centrifuged at 3000g for 5 minutes before being transferred to GC ampoules for injection. The retention time of the DMF peak was 3.26 minutes relative to the internal standard; the calibration curve showed a correlation of 0.98. The DMF quantification appeared linear up to 100 mg / ml and the lower limit of detection was set at 0.5 mg / ml based on a signal to background ratio of 3: 1.

9 cm diameter transdermal patches were used within 12 hours of manufacture. Each patch had a high density polyethylene backing (0.245 g), a Teflon permeable membrane (0.2 µm pore size; 0.268 g) for placement against the skin and containing 0.573 g of colloidal silica impregnated with 7.067 g DMF between the backing and permeable membrane. The patches were visually inspected to confirm the absence of penetration and weighed on an analytical balance to confirm less than 10% deviation from a total weight of 8.153 g.

-9EN 298510 B6

The patches were applied to the forearm skin and secured with an Elatoplast ™ bandage. Either one or two patches were used at the initial dose, as determined from an assumed skin patency rate of 9.4 mg / cm ² / g patient body weight.

A phased dose of DMF was sampled every two hours to determine peak plasma levels of DMF. Patients were clinically monitored for any toxic side effects and kept under overall biochemical control while the dose was increased as necessary to achieve a maximum plasma level of 100 to 120 ppm. Once the correct dose was established, transdermal DMF was used once a week. The initial evaluation included a daily determination of:

(a) vital signs and body weight

(b) the clinical list and the Kamofsky score

c) total blood cell count and erythrocyte sedimentation rate

d) serum urea, creatinine, glucose, sodium, potassium, ALT, AST, alkaline phosphatase and total bilirubin

e) analysis of CD4 + and CD8 + cells and CD4 + / CD8 + ratio by Coulter

f) quantitative estimates of HIV-1 load using PCR (Roche Amplicor) and analysis of anti-p24 antibodies (Abbott)

(g) Urinalysis (Dipstix).

At each weekly visit, the patient was evaluated for adverse reactions, nearly all of these tests were repeated, and blood and urine were collected to determine DMF and its metabolites.

Patient 1 (ADF) started the protocol in a relatively good state of health, complaining basically about pain in the hands and feet and inability to sleep. He was given two DMF patches once a week with an average exposure time of 8 hours. The average weekly dose of DMF was 6.11 g and the resulting mean maximum plasma DMF level of 75 mg / l. After 9 weeks, the number of CD4 + T cells increased from 140 to 640 cells / μΙ and the viral load measured by PCR was reduced from 250,000 to 50,000 copies / ml. After 10 weeks, the patient's weight increased from 81.9 to 96.0 kg, the patient felt clinically comfortable and no longer complained of limb pain.

Patient 2 (AM) entered the protocol with complaints of loss of strength, inability to sleep, pain in the hands and feet and had herpetic sores in his mouth. He received one DMF patch per week with an average exposure time of 8 hours. The mean weekly dose of DMF was 7.12 g and the resulting mean maximum plasma DMF level of 125 mg / l. After 9 weeks, the CD4 + T cell count increased from 460 to 720 cells / μΙ and the viral load measured by PCR decreased from 29,000 to 13,000 copies / ml and the patient's weight increased from 58.4 to 63.0 kg. Herpetic sores cured the patient, limb pain disappeared and the patient appeared clinically good.

Patient 3 (SM) started a protocol with overt clinical AIDS and complained of respiratory problems. He was given two DMF patches once a week with an average exposure time of 8 hours. The average weekly dose of DMF was 8.97 g and the resulting mean maximum plasma DMF level was 121 mg / l. After 7 weeks, the CD4 + T cell count increased from 39 to 138 cells / μΙ and the viral load measured by PCR decreased from 222,000 to 16,000 copies / ml and the patient's weight increased from 74.2 to 100 kg. The patient's appetite improved, respiratory problems resolved, and the patient appeared clinically good and began to exercise again.

Patient 4 (EM) began a protocol with secondary infection (including herpes), anemia, diarrhea and acne. He was given two DMF patches once a week with an average exposure time of 8 hours. The average weekly dose of DMF was 7.33 g and the resulting mean maximum plasma DMF level of 90 mg / l. After 18 weeks, the number of CD4 + T cells increased from 249 to 450 cells / μΙ and by PCR

- The measured viral load decreased from 13,000 to 4000 copies / ml and the patient's weight increased from 81.5 to 90.4 kg. The patient appeared clinically good and did not have active health problems.

Patient 5 (SV) began the protocol by complaining of poor appetite, forgetfulness, abdominal pain, and severe fatigue that prompted him to consider selling his stake in the business. He was given two DMF patches once a week with an average exposure time of 6 hours. The mean weekly dose of DMF was 3.75 g and the resulting mean maximum plasma DMF level of 67 mg / l. After 5 weeks, the number of CD4 + T cells increased from 354 to 396 cells / μΙ and the viral load measured by PCR decreased from 156,000 to 13,000 copies / ml and the patient's weight increased from 56.0 to 58.0 kg. The patient appeared clinically good, acquired the shares of his business partner, and started to manage the business himself.

Patient 6 (VV) began the protocol by complaining of secondary infections (including herpes), ataxia and numbness of the left hand and left side of the face. He was given two DMF patches once a week with an average exposure time of 8 hours. The mean weekly dose of DMF was 8.25 g and the resulting mean maximum plasma DMF level of 110 mg / l. After 19 weeks, the CD4 + T cell count increased from 260 to 450 cells / μΙ and the viral load measured by PCR was reduced from 120,000 to 24,000 copies / ml and the patient's weight increased from 75.4 to 84.6 kg. Secondary infections disappeared and the patient appeared clinically good.

Patient 7 (AJF) started the protocol seriously ill. The patient's initial CD4 + T-cell status was 29 cells / μΙ and the initial PCR measured viral load was 1,156,000 copies / ml. Treatment took one patch with an average exposure time of 4 hours. The mean weekly dose of DMF was 4.60 g and the resulting mean maximum plasma DMF level of 100 mg / l. After the first treatment, the number of patient CD4 + T cells decreased to 14 cells / μΙ. Treatment was given daily for the first five days and then continued at weekly intervals. After 9 treatment cycles, the patient CD4 + T cell count increased to 39 cells / μΙ and the viral load measured by PCR was reduced to 9000 copies / ml and the patient's weight increased from 46.5 kg (at the start of DMF therapy) to 49.0 kg. The patient felt comfortable and returned to full-time employment.

Patient 8 (MS) began a protocol with severe herpetic lesions in the lower back and genitals. He was given two DMF patches once a week with an average exposure time of 8 hours. The average weekly DMF dose was 6.24 g and the resulting mean maximum plasma DMF level was 130 mg / L. After 8 weeks, the number of CD4 + T cells increased from 200 to 240 cells / μΙ and the viral load measured by PCR was reduced from 1,200,000 to 250,000 copies / ml and the patient's weight increased from 48.1 to 52.2 kg. Herpetic lesions have completely disappeared.

Two patients were excluded from the trials, one because of alcohol dependence, the other because of viral hepatitis B.

Most patients have experienced mild local irritation in the application area after patch removal; the skin at the application site had a maculapapular appearance, probably due to intense hydration under the patch. In one case, slight blisters were formed which resolved within 24 hours and did not cause significant discomfort to the patient. Most patients experienced mild transient nausea, usually on the third day of treatment, which gradually decreased during the treatment protocol; one patient reported moderate transient nausea. Four patients showed a transient increase in liver enzymes, which never exceeded three times the upper limit of normal and which in most cases returned to pre-treatment levels before the next dose of DMF. Most cases of elevated liver enzymes were associated with at least one factor not related to the treatment protocol (alcohol consumption, hepatitis and previous antiHIV therapy with other agents).

Almost all patients showed clinical improvement after 2-3 weeks of treatment. As shown in FIG. 4, each patient showed an improved overall condition after DMF treatment when assessed by Kamovsky's performance scale, in which the patient's overall condition is determined by a numerical value

As follows: 100 = normal, no problems, 90 = able to perform normal activities, mild signs or symptoms of the disease, 80 = normal activity with exertion, 70 = self-care, unable to perform normal activity or active work, 60 = requires occasional help but is able to cater for most needs, 50 = requires significant help frequent medical care, 40 = not able, requires special care, 30 = serious incapacity, indicated hospitalization although there is no immediate risk of death, 20 = very 10 = moribund, fatal progression progresses rapidly, 0 = dead (see Kamofsky et al., 1984, Cancer 1: 634-656). The improvement in neurological symptoms and herpes viral infections was remarkable. Additional anti-microbial therapy for secondary infections was seldom needed. Overall fatigue and appetite improved significantly during the first 14 days of DMF treatment. All patients increased weight. Clinical improvement correlated well with disease status as assessed by viral load and CD4 + cell count. In five of the eight patients, the relative viral load measured by PCR may fit Gompertz curves; this analysis revealed a 88.8% reduction in viral load measured by PCR after 42 days of DMF treatment. In seven out of eight patients, the relative number of CD4 + T-cells may conform to Gompertz curves; this analysis showed a 73.4% increase in the number of CD4 + T cells after 42 days of DMF treatment.

The present invention is not limited to the scope of the disclosed embodiments, which are intended to illustrate particular aspects of the invention. Indeed, various modifications of the invention in addition to those described herein will be apparent to those skilled in the art from the foregoing descriptions and accompanying drawings. Such modifications are intended to fall within the scope of the appended claims.

All publications cited herein are hereby incorporated by reference in their entirety.

Claims (6)

PATENT CLAIMS A transdermal patch for transdermal administration of a therapeutic agent, comprising an external support portion; a permeable membrane suitable for placement against the skin of the animal to be treated; and an adsorbent disposed between the outer support portion and the permeable membrane, the adsorbent having adsorbed therapeutics comprising N, N'-dimethylformamide. The transdermal patch of claim 1, wherein the therapeutic composition comprises at least 0.25 g of Ν, Ν'-dimethylformamide per transdermal patch. The transdermal patch of claim 2, wherein the therapeutic composition comprises at least 5 g of Ν, Ν'-dimethylformamide per transdermal patch. The transdermal patch of any one of claims 1 to 3 inclusive, wherein the therapeutic composition comprises Ν, Ν'-dimethylformamide in an amount effective to treat viral or microbial infections or slimming syndrome in human adults or human children. Transdermal patch according to any one of claims 1 to 4 inclusive, characterized in that no pharmacologically active substance other than Ν, Ν'-dimethylformamide is present. Transdermal patch according to any one of claims 1 to 6 inclusive, characterized in that the adsorbent is colloidal silica and the permeable membrane is a polytetrafluoroethylene membrane having a pore diameter of from 0.1 µm to 0.5 µm.