LV12490B - Drug delivery devices and methods for treatment of viral and microbial infections and wasting syndromes - Google Patents

Abstract

Novel drug delivery devices, methods and therapeutic compositions are described for treating viral and microbial infections and wasting syndromes in an animal, including a human patient. According to the invention, a polar compound such as dimethylformamide or dimethylsulfoxide is administered to a patient in need of treatment, preferably by a transdermal route. The invention further provides a vaccine prepared from antibodies harvested from the body of a patient treated by the method of the invention for a viral infection.

Description

1 1LV 12490

DRUG DELIVERY DEVICES AND METHODS FOR TREATMENT OF VIRAL AND MICROBIALINFECTIONS AND VVASTING SYNDROMES

1. FIBLD OF THE INVENTION

The present invention is directed to methods of treating an animal a-ffected by a virai or microbial infection, particularly an infection with a retrovirus, 5 such as HIV, or by a wasting syndrome, especially a wasting syndrome asso.ciated with HIV infection or -malignancy/ by administeriņg a polar compound such as N,N'-dimethylformamide (DMF) or dimethylsulfoxide '(DMSO) . — The invention also. provides pharmaceutical preparations and 10 drug delivery devices comprising a polar compound such as DMF or DMSO for treatment of an animal affected by a virai or other microbial infection or a wasting syndrome.·

2. BACKGROUND OF THE INVENTION 2.1 THE HUMAN IMMONODEFICIENCr VIRŪS 15 Human_ imntunodeficiency virus (HIV) inducēs, a persistent and Progressive infection leading, in the vast majority of cases, to the development of the acquired immunodeficiency syndrome (AIDS) (Barre-Sinoussi et al., 1983, Science 220: 868-870; Gallo et al., 1984, Science 20 224:500-503) . There are at least two distinct types of HIV: HIV-i (Barre-Sinoussi et al., 1983, Science 220:868- 870;_Gallo et al., 1984, Science 224:500-503) and HIV-2 (Clajcel et al., 1986, Science'223-.343-346; Guyader et al., 1987, Nature 326:662-669). In humāns, HIV replication 25 occurs prominently in CD4+ τ lymphocyte populations, and HIV infection leads to depletion of this celi type and eventually to imriiune incompetence, opportunistic 2 iņfections, neurological dysfunctions, neoplastic growth, and ultimately death. HIV is a member of the lentivirus family of retroviruses (Teich et al., 1984, RNA Tumor Viruses, Weiss 5 et al.f eds., CSH-press, pp. 949-956). Retroviruses are small enveloped viruses that contain a single-stranded RNA genome, and replicate via a DNA intermediate produced by a virally-encoded reverse transcriptase, an RNA-dependent DNA polymerase (Varmus, H., 1988, Science 240:1427-1439). 10 Other retroviruses include, for example, oncogenic viruses such as human T-cell leukemia viruses (HTLV-l, -II, -III), and feline JLeukemia vīrus.

The HIV virai particle consists of a virai core, composed in part of capsid proteīns designated p24 and pl8, 15 together with the virai RNA genome and those enzymes rēquired for early replicative events. Myristylated gag protein forms an outer virai shell around the virai core, which is, in tum, surrounded by a lipid membrane envelope derived from the infected celi membrane. The HIV envelope 20 siīrface glycoproteins are synthesized as a single 160 kilodalton precursor protein which is cleaved by a cellular protease during virai budding into two glycoproteins, gp41 and gpl20. gp41 is a transmembrane glycoprotein and gp!20 is an extracellular glycoprotein which remains non-25 covalently associated with gp41, possibly in a trimeric or multimeric form (Hammerskjold, M. and Rekosh, D., 1989,

Biochem. Biophys. Acta 989:269-280). r HIV, like other enveloped viruses, introduces virai genetic raaterial _into the host ~čēll through a virai-3 0 envelope mediated fusion of virai and tacrget' raembranes. H-IV is targeted to 004^. celis because a CD4 celi surface protein (CD4) acts as the cellular receptor for the HIV-1 virus (Dalgleish et al., 1984, Nature 312:763-767,-

Klatzmann et al., 1984, Nature 312:767-768, Maddon et al., 1986, Celi 47:333-348). Virai entry into cēlis is 35 3 LV 12490 dependent upon gpl2Q binding the cellular CD4 receptor molecules (Pal et al., 1993, Virology 194:833-837; McDougal et al., 1986, Science 231:382-385, Maddon et al., 1986, Celi 47:333-348), explaining HIV's tropism for CD4· celis, 5 while gp41 anchors the envelope glycoprotein coraplex in the virai membrane. The binding of "gpl20 to CD4 inducēs conformational changes in the virai glycoproteins, but this binding alone is insufficient to lead to infection (reviewed by Sattentau and Moore,-1993, Philos. Trans. R. 10 Soc. London (Biol.) 342:59-66).

Studies of HIV-1 isolates. have revealed a _ heterogeneity in their ability to infect different human celi types (reviewed by Miedema et al·., 1994, Immunol. Rev. 14.0:35-72). The majority of extensively passaged 15 laboratory strains of HIV-l readily infect cultured T celis lines and primary T lymphocytes, but not primary monocytes or macrophages. These strains are termed T-tropic. T-t.ropic HIV-l strains are raore likely to be found in HIV-1. inf ected individuāls during the late_ stages of aids (Weiss 20 et al., 1996, Science 272:1885-1886). The raajority of primary HIV-1 isolates (i.e._, viruses not extensively passaged in culture) replicate efficiently in primary lymphocytes, monocytes and macrophages, but grow poorly in established T celi lines. These isolates have been termed 25 M-tropic. The virai determinant of T- and M- tropism maps to alterations in the third variable region of gpl20 (the V3 loop) (Choe et al., 1996, Celi 85:1135-1148; Cheng-Mayer et al., 1991, J. Virol. 65 ?6931-694l; Hwang et al., 1991, Science 253:71-74; Kim et al.< 1995, J. Virol., 69*1755-30 17£1; and 0' Brien et al., 1990, Nature 348:69-73). The characterization of HIV isolates mth distinct tropisms taken together with the observation that binding to CD4 celi surface protein alone is insufficient to lead to infection, suggest that cell-type specific cofactors might 5 4 be required in addition to CD4 for HIV-1 entrv into the host celi.

2.2 TREATMEMT FOR HIV INEECTION 10 15 HIV infection is pandemic and Hiv-associated diseases represent a major world health problem. Although considerable effort is being put into the design of effective Therapeutics, currently no curative anti-retroviral drugs against AIDS exist. In attempts to develop such drugs, several stages of the HIV life. cycle have been considered as targets for therapeutic intervention (Mitsuya,-H., et al., 1991, F&SEB J. 5:2369-23 81) . Many virai targets for intervention with HIV life cycle have been suggested, as thē prevailing view is that interference with a host celi protein would have deleterious side effects. For example, virally encoded reverse transcriptase. has been one focus l of drug development. A humber of reverse-transcriptase-targeted I ) — drugs, including 2 , 3 -dideoxynucleoside analogs such as 20 AZT, ddl, ddC, and d4T have been developed which have been shown to been active against HIV (Mitsuya, H., et al., 1991, Science 241* 1533-154'4) . 25

The new treatment regimes for Ηΐν-i show that a combination o£ anti-HIV compounds, which target reverse transcriptase (RT) , such as azidothymidine (AZT), lamivudine (3TC) , dideoxyinosine (ddl)', didcuxycytidine (ddC) used in combination with an HIV-1 protease inhibitor have a far greater effect (2 to 3 logs reduction) on virai load compared to AZT alone (about 1 log~ reduction) . For example, irapressive results have recentļy been obtained with a combination .of AZT, ddl, 3TC and ritonavir (Perelson, A.S., et al., 1996, Science 15:1582-1586) . However, it is likely that long-term use of combinations of these Chemicals will lead to toxicity, especially to the bone marrov. Long-term cytotoxic therapy may also lead to 30 5 5LV 12490 suppression o£ CD8 + T celis, which are essential to the control of HIV, via killer celi activity (Blazevic, V., et al., 1995, AIDS Res. Hum.Retroviruses 11:1335-1342) and by the release of suppressive factors, notably the chemokines 5 Rantes, MlP-loc and MIP-IS (Cocchi, F., et al., 1995, Science 270:1811-1815) . Another major concem in long-term Chemical anti-retro virai therapy is the development of HIV mutations with partial or coraplete resistance (Lange, J.M., 1995, AIDS Res. Hum Retroviruses l£:S77-82) . It is thought 10 that such mutations may. be an inevitable consequence of anti-viral therapy. The pattern of disappearance of wild-type virus and appearnace of mutant virus due to treatraentr combined with coincidental decline in CD4+ T celi numbers strongly suggests that, at least with some compounds, the 15 appearance of virai mutants is a major underlying factor in the failure of AIDS therapy.

Attempts are also being made to develop drugs which can inhibit virai entry into the celi,- the earliest stage of HIV infection. Here, the focus has thus far been on 20 CD4, the celi surface receptor for HIV. Recombinant soluble CD4, ±ov example, has been shown to inhibit infection of CD4+ T celis by some HIV-1 strains (Smith, D.H., et al., 1987, Science 238:1704-1707). Certain primary HIV-1 isolates, however, are relatively less 25 sensitive to inhibition by recombinant CD4 (Daar, E., et al., 1990, Proc. Nati. Ācad. Sci. USA 87:5574-6579) . In addition, recombinant soluble CD4 clinical trials have produced inconclusive results (Schooley, R., et. al., 1990, Ann. Int. Med. 2127247-253; Kahn, J.O., et al., 1990, Ann. 30 Int. Med. 112:254^261,- Yarchoan, R-, et al,, 1989, Proc.

Vth Int. Conf. on AIDS, p. 564, MCP 137) .

The late sfeages of HIV replication, which involve crucial vimas-specific processing of certain. virai encoded proteīns, have also been suggested as possible anti-HIV 6 drug targets. Late stage processing is dependent on the activity of a virai protease, and drugs are being developed which inhibit this protease (Erickson, J., 1990 Science 249:527-533) . 5 Recently, chemokines produced by CD8 + T celis have been implicated In suppression of HIV infection (Paul, W.E. , 1994, Celī. 82,1171; Bolognesi, D.P., 1993, Semin.

Inanunol. 5:203) . '

The chemokines RANTES, MIP-l-oc and ΜΙΡ-1β, which are 10 secreted by CD8+;T celis, were shown to suppress HIV-1 p24 antigen praduction in celis·'-infected with HIV-1 or HIV-2 isolates in vitro (Cocchi, F, et al., 1995, Science 270:1811-1815) ..

Thus, these and other chemokines may prove useful in 15 therapies for HIV infection. The clinical outcome, - however, of ali t'hese and other candidate drugs is stili in question.

Attention iš also being given to the development. of vaccines for the treatment of HIV infection. The HIV-1 20 envelope proteīni" (gpl60, gpl20, gp41) have been shown to be the major-antigens for anti-HIV antibodies present in AIDS patients (Bariņ et al.-,- 1985, Science 228:1094-1096) . Thus far, therefore, these proteīns seem to be the most promising candidates to act as antigens for anti-HIV 25 vaccine development. Several groups have begun to usa various portions of gplSO, gpl20, and/or gp41 as immunogenic targets for the host irnmune system. See for example, Ivanoff, L·., et al., U.S. Pat. No. 5,141,867; Saith, G., et al., W092/22,654; ShaffermānT AVI_ 30 WC91/Q9,872; Fcmoso, C-, at al., VĪO90/G7,119 . To thi-s- end, vaccines dīrect against HIV proteins are problematic in that the' virus mutates rapidly rendering many of these vaccines ineffective. Clinical results concerning these candidate vaccines, however, stili remain far in the future. 35 7 7LV 12490

Thus although a great deal of effort is being directed to the design and testing of anti-retroviral drugs, effective, non-toxic treatments are stili needed.

2. WASTING SYDROMES 5 Wasting syndrome is a serious clinical prpblem characterized by a decrease in body mass of more than 10% fram baseline body weight and a disproportionate loās of body mass with respect to body fat (Weinrotb et al.,_1995, Infectious Aģents and Disease 4:76-94; Kotler and Grunfeld, 10 1995, AIDS Clin. Rev. 9£:229-275) . Thus, wasting is distinguished from starvation in which higher Ievels of body fat than body celi mass are depleted (Kotler et al., 1985, Am Clin. Nutr. 42:1255-1265; Cahill, 1970, N. Engrl. J. Med. 282.:568-675) . Wasting is srssociated with a 15 variety of conditions, including HIV infection, bther 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. 15:680-690) . Importantly, wasting is a signiftcant 20 factor in the mortality of patients suffering from infections or cancer. Iņ fact, body celi mass- depletion has a linear relationship to time of survival in AIDS patients (Kotler et al., 1989, Am. J. Clin. Nutr. 50:444-447) . 25 The cause of wasting syndrome in AIDS and other conditions is unclear and is most likely multifactorial. Metabolic abnorraalities, irregular Ievels of hormone-s and cytokines, and malabsorption have ali been implicated in wasting syhdrome. Not ali'AIDS patients suffer from wasting, suggesting'that the cause of the wasting is not HIV itself. Most cases of HIV associated wasting sytrdrome are apparently caused by complications of AIDS, such as secondary infections and gastrointestinal disease (Kotler and Grunfeld, 1995, AIDS Clin. Rev. 96:229-275). 30 8

Current and. potential theraņies for wasting syndromes include nutritional support, appetite enhancers such as dronabinol and raegestral acetate, anabolic therapies, such as growth hormone, and cytokine inhibitors. However, mixed 5 results have been obtained with nutritional support and appetite enhancers in that patients tended to gain only fat and not overall body mass. Administration of growth hormone, and cytokine inhibitors are stili being tested and may pose a risk of side effects (Kotler and Grunfeld, 1995, 10 AIDS Clin. Rev. 96:229-215; Weinroth et al., 1995 Infectious Aģents and Disease 4:76-94).

Thus, treatment of wasting is critical to the survival and well-being of patients suffering from serious diseases suGh as cancer and AIDS; thus, there is a need for safe and 15 effective therapies for wasting syndrome associated witir cancer, AIDS and other infectious diseases. 2.4 PROPERTIES OF DliSSTHTLgORKAMIDE AUD OTHER POLAR-

COMPOUNDS N,N' -Dimethylformamide (DMF) (molecular formula: C3H7ON) 20 is a colourless, polar, hygroscopic liquid with low volatility and a boiling jpoint of 152.5-153 .5?C. It is freely miscible with water, alcohols and some hydrocarbons. DMF is generally used as a polar solvent and is readilv absorbed through the skin, by inhalation, and upon oral 25 ingestion. DMF is rapidlv metabolized, mainlv in the liver, and excretion occurs principally in the urine. In rat, mouse, hamster and man the main metabolites of DMF are N-hydfoxymethyl-N-methylformamide (HMMF) , N-methvlformamide. JŅMF) , and N-acecyl-S- (M-metnyicarbamoyl) cysteine' (AMCC), as well as dihydroxymethyl£ ormamide (DHMF) and īF-hydroxymethylformamide (HMF) . Unchanged DMF is excreted in the urine as a small fraction. of an administered dose of DMF. 30 9 9LV 12490 DMF has low acute dermal, oral and inhalation toxicity. It is considered to be a mild to moderate skin and eye irritant and readily permeates the skin. There is no indication of skin sensitizing properties. The 5 principal toxic effect of DMF and its metabolites is on the liver; DMF is well known to cause reversible hepatic damage associated with typical clinical complaints, classical biochemical changes in the blood, and the appearance of hepatocyte necrosis in liver biopsies. DMF is teratogenic, 10 but is not thought to be a mutagen or a carcinogen.

Viza et al. have -reported that DMF and DMSO inhibit in vitro replication of HIV and Human Herpes Virus 6 (HHV-6) in certain cultured celi lines. (See Viza et al., 1990 AIDS Res. Hum. Retroviruses 6:131-112; Viza et al., 1989, 15 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 differentiating aģent for certain transformed celis in culture (See· Koeffler, 1983, Blood 62:709-721; Calabresi et al., 1979, iū Biochem. Pharmacol. 28:1933-1941). When added to certain malignant celis in vitro, DMF has been -reported' to reduce their tumorigenicity upon subsequent inoculation into nude mice (See Dexter, 1977, Cancer Res. 37:3136-3140; Dexter et al., 1979, Cancer Res. 39:1020-1025). Upon intraperitoneal 25 injection into nude mince, DMF and NMF have been reported to slow the growth of certain human cancer 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 Otolaryņgol. 105:488-493, Dexter et al., 30 1982, Cancer Res. 42:5018-5022). ~However, the toxic side- effects of formaraide and its N-methyl derivatives in a mouse sarcoma allograft modei led investigators to conclude that these aģents were unlikely to prove therapeutically useful (See Clarke et al., 1953, Proc. Sec. Exp. Biol. Med. 35 84:203-207). 10

Attempts at treating human cancer patients with DMSO led to the conclusion that no objective response had been shown (See Spremulli & Dexter, 1984, J. Clin. Oncol. 2:227-241) . Oral administration of NMF to human patients with 5 ' cancer of the head and neck was reported as resulting in hepatotoxicity with no beneficial response (see Vogel et al., 1987, Invest. New Drugs 5:203-206), or with only minimal activity (See Planting et al., 1987, Cancer Treat Rep. 71:1293-1294). 10 U.S. Patent No. 3,551,154 discloses the use of DMF as a penetration enhancer to promote'transdermal absorption of top’ically applied medications. U.S. Patent No. 4,855,294 discloses the use of glycerin to mit-igate the skin irritation arising from the use of DMSO and DMF as 15 _ penetration enhancers to promote transdermal absorption of topically applied medications. f- The use of DMSO as a penetration enhancer to promote transdermal absorption of antiviral aģents is discussed in Noodford & Barry, 1986, j. Toxicol. Cut. & Ocular Toxicol. 5:167-177. 20 Citation or Identification of any reference in Section 2 (or any other section) of this application shall not be construed as an admission that such reference is available as prior art to the present invention.

3 . STJMMARY OF THE INVENTION 25 The present invention is directed to methcds, compositions and drug dalivery. devices for treating an animal affected by a virai or other microbial infection, especially an infection with a rētrovirus such as HIV. The invention also provides methods, compositions _and drug delivery devices for treating an animal affected by a wasting syndrome, such as ^-wasting associated with KIV infection or malignancy.

According to the present invention, there is administered to an animal in need of treatment a 30 11 11LV 12490 composition comprising N,N'dimethylformaraide (dimethylformamide, DMF) ; N-hydroxymethyl N-methylformamide ( HMMF ) ; N-hydroxymethy1formamide (HM.F) ; dihydraxymethylf ormamide (DHMF) ; N-acetyl, S-(N-5 methylcarbamoyl)cysteine (AMCC); N-methylformamide (NMF),· dimethylsufoxide (DMSO); formamide; acetamide; methylacetamide, dimethylacetamide ,* diethylacetamide; isopropylacetamide; diisopropylacetamide; N-acetylpiperidine; N-(|8-hydroxyethyl) acetamide; N,N'-di(S-10 hydroxyethyl) acetamide,· N-acetylmorpholine, acrylamide; propionamide; N-fluoromethyl-N-methyl-formamide; pyridine-N-oxide; or any aģent selected from the group consisting of amides of the general formula Rļ-CO-NRļS^, in which Rļ and R2 are independently selected from the group consisting of 15 H, methyl, halomethyl, saturated and unsaturated C2-C3 alkyl groups, and hydroxylated alkyl groups; or Rļ and R2 are together selected from the group consisting of (¢312)4, ^^2^5' and 5111(1 R3 is selected from the group consisting of H, methyl, 20 and saturated and unsaturated C2-C3 alkyl groups. The thērapeutic composition may comprise a mixture of any two or more of the aforementioned compounds.

In a patient infected with HIV, the therapeutic regimen may optionally combine a composition of the present 25 invention with one or more additional aģents effective for treating HIV infection, including but not limited to aģents selected from the group consisting of nucleoside analog reverse transcriptase inhibitors, non-nutfleoside reverse ~transcriptase inhibitors, and 'protease inhibitors, in any 30 desired combination.

The invention also extends to a vaccine prepared from antibodies that are obtained from the body of an animal after treatment with a composition of the invention for a virai infection. 12

4. BRIEF DESCRIPTION OF THE FIGŪRES

The present invention may be raore fully understood by reference to the following detailed description of the invention, examples of specific embodiments of the invention and the appended figurēs in which: FIGURĒ 1 illustrates plasma concentrations of DMF as a function of tirae in 4 patients treated with 2 DMF derraal patches for 8 hours. FIGURĒ 2 illustrates plasma concentrations of DMF as a function of time~in 3 patients treated with 2 DMF dermal patches f or S hours FIGURĒ 3 illustrates HIV-1 virai load as measured by quantitative PCR in a patient treated with transdermal DMF. Two DMF patches were-placed against the skin of the forearm for 12 hours on days 0, 8, and 13 (indicated by arrows). FIGURĒ 4 illustrates the general condition of HIV iūfected patients before and after. treatment with transdermal DMF, as assessed according to. the. .Kamofsky performance scale. See text, Section 7, for details.

5. DETAILED DESCRIPTION OF THE INVENTION

The present invention provides methods, compositions and drug delivery devices for treating virai and microbial infections. In one embodiment, the infection to be treated is an infection with a retrovirus such as HIV, including an asymptomatic infection, a latent infection, an infection accompanred by one or mora symptoms of the AIDS-related ccmplex, and an infection acccmpanied by clinical AIDS-Altematively, the infection to be treated is any other virai or itiicrobial' infection; including infection with rubella, a herpesvirus such as Human Herpes virus 6, the Epstein-Barr virus “or cytomegalovirus, infection with any virus having a capsid protective coating, and any opportunistic infection associated with disease of the 13 13LV 12490 immune system, such as an opportunistic infection in a patient infected with HIV.

The present invention also provides methods, compositions and drug delivery devices for treatment or 5 prevention of any disease or disorder characterized by a loss of body mass. Particular conditions that can be treated by the methods and compositions of the invention include, but are not limited to, wasting associated v/ith virai' (e.g. HIV), bacterial, or any other types of 10 infection or sepsis, cachexia associated with malignancy, chemotherapy or radiation therapy, wasting associated with chronic cardiovascular disease, wasting caused by exposnre to toxic or radioactive substances, and wasting associated with diarrhea_and other gastrointestinal disorders. 15 The subject to be treated may be any animal, including but not limited to a monkey, cow, sheep, ox, pig, horse, cat, dog chicken and the like, and is preferably a mammai, more pre£erably a primate, and most preferably a human adult or child, for instance a human child weighing at 20 least 3 kg. As used herein, the term "patient" refera-to any animal in need of treatment according to the methods or compositions of the present invention.

According to the present invention, there is administered a therapeutic composition comprising DMF; 25 HMMF; HMF; DHMF; AMCC; NMF; DMSO; formamide; acetamide; methvlacetamide/ dimethylacetamide; diethylacetamide/ isopropylacetamide; diisopropylace tamide ,· N-acetylpiperidine, N-(fi-hydroxyethyl) acetamide; N,N'-di_(S-hydroxye thy 1) acetamide; N - acety lmorpholine; acrylamide ; 3 0 propionamide^I-f luoromethyl-N-methyl-f ormamide;pyridine-N- oxide; any aģent selected from the group consisting of amides of the general formula Rj-CO-NRjI^, in whiq.h Rļ and R2 are independently selected from the group consisting of H, methyl, halomethyl, saturated and 5 14 unsaturated C2-C3 alkyl groups, and hydroxylated alkyl groups; and R3 is selected frora the group consisting of K, methyl, and saturated and unsaturated C2-C3 alkyl groups; or any aģent selected from the group consisting of araides of the general formula R3-CO-NR1R2, in which Rļ and R2 are independently selected from the group consisting of H, methyl, halomethyl, saturated and unsaturated C2-C3 alkyl groups, and hydroxylated alkyl 10 groups,· or Rļ and R2 are together selected from the group consisting of (012)4» (^¾) 5, and ((1¾) 20 (^2)2- R3 is selected from the group consisting of H, methyl, and saturated and unsaturated Cļ -C3 alkyl groups. 15 20

In one specific embodiment, at least one of Rļ and R2 is a methyl group. In another specific embodiment, at least one of Rļ and R2 is a fluorinated Cļ - C3 alkyl group. The therapeutic composition may comprise a mixture of any two or more. of the- aforementioned compounds. Especially preferred is a composition comprising DMF. 25

For "treatment of an animal infected with HIV, the therapeutic regimen may optionallv include, in addition to a composition of the present invention, one or more other aģents effective for treating HIV infection, for instance one or more nucleoside analog reverss transcriptase inhibitors such as zidovudine (AZT, ZDV), zalcitabine (ddC), didanosine (ddl), lamivudine (3TC), stavudine (d4T); one"or more ncn-nuclecside _rev3rst transcriptase irhibitcrs such as nevirapine, delavirdine, loviride, atevirdine, pyridinone,· one or more protease inhibitors such as saguinavir, indinavir, ritonavir, nelfinavir; or any combination of the aforesaid or other anti-HIV therapeutic aģents. The composition of the present invention and the 30 15 15LV 12490 additional anti-HIV therapeutic aģent or aģents may be administered simultaneously, sequentially, or in cycles of treatment accordlng to any desired therapeutic protocol.

The compositions of the present invention may be 5 administered by any desired enteral or parenteral route, including but not limited to transdermal, intradermal, subcutaneous, intramuscular, intraperitoneal, intravenous, intranasal, epidural, intralymphatic and oral routes. The compounds may be administered by any convenient raethod, for 10 exatnple by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g:, oral, - gastric, intestinal or rectal raucoša, etc.) and may be administered together with other biologically active aģents. 15 - Ādministration can be systemic or local. In addition, it may be desirable to introduce the pharmaceutical compositions of the invention into the central nervous system by any suitable route, including intraventricular and intrathecal injection; intraventricular injection may’ 20 be facilitated by an intraventricular catheter, for example, attached to a reservoir, such - as an Ommaya reservoir. Pulmonary administration can also be enq?loyed, e.g., by use of an inhaler or nebulizer, and the formulation may include an aerosolizing aģent. If desired, 25 any two or more routes of administration may be employed siraultaneously, sequentially, or in crycles of treatment according to any therapeutic protocol.

In a specific embodiment, it may be desirable to ādminister a composition of the invention -loca.lly to the area in need—of treatment; this may" be achieved, for example and hot by way of limitation, by topical application, by injection, by raeans of a catheter, by means of a suppository, or by means- of an implant, said implant being of a porous, non-porous, or gelatinous .30 16 material, including membranes, such as sialastic metnbranes, or fibres.

In another embodiment, a coraposition of the invention can be delivered in a vesicle, in particular a liposome 5 . (see Langer, Science 249:1527-1533 (1990); treat et al., in Lipasomes in the Therapy of Infectious Disease and Cancer, Lopez-Berestein and Fidler (eds.), Liss, New York, pp. 353-365 (1989); Lopez-Berestein, ibid., pp. 317-327; see generally ibid.) 10 In yet another embodiment, — a composition of the invention can be delivered in a controlled release system.

In one embodiment, a pump may- be used (see Langer, - stipra; Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987); Buchwald et al., Surgery 88:507 (19~80) ; Saudek et al, N. 15 Engl. J. Med^ 321:547 (1989) ). In another embodiment, polymeric materiāls can be used (se~e Medical Applications _of Controlled Release, Langer and Wise (eds.), CRC Pres., Boca Rāton, Florida (1974) Controlled Drug

Bioavailability, Drug Product Design and Performance, -20 Smolen and Bail (eds.) Wiley, New Ygrk (1984); Ranger and Peppas, J. Macromol. Sci. Rev. Macromol. Chem. 23,:61 (1983) : see also Levy et al., Science 228:190 (1985)·;

During et al., Ann. NeuroĪ. 25:351 (1989); Howard et al., J. Neurosurg. 71:105 (1989)). In yet another embodiment, 25 a controlled release system can be placed in proximity of the therapeutic taraet, thus requiring only a fraction of the svstemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, stipra, vel. 2, pp. 115-138 (1984)). 30 Other controlled release systerus are discussed in the r'eview by Langer (Science 249:1527-1533 (1990)).

The present invention also provides pharmaceutical preparations. Such preparations comprise a therapeutically effective amount of a composition of the invention and a pharmaceutically acceptable carrier. In a specific 35 17 17LV 12490 embodiment, the term "pharinaceutically acceptable" means approved by a regulatory agency of the Federal or a State ar other government or listed in the U.S. Pharraacopoeia or other generally recognized phartnacopoeia for use in 5 animals, and more particularly in humāns. The term "carrier" refers to a diluent, adjuvant, excipient, or vehicle with which the composition of the present invention is ādministered. Such pharmaceutical carriers can be - sterile liguids, such as water and oils, including those of 10 petroleum, animal, veģetārie or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water is a preferred carrier when the pharmaceutical preparation is administered 'intravenously. Saline

Solutions and aqueous dextrose and glycerol Solutions can 15 also be employed as liquid carriers, particularly for- injectable Solutions. Suitable pharmaceutical excipients include starch, ‘ glucose, lactose',—sucrose, gelatin, malt, rice, flour, chalk, silica. gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, 20 glycerol, propylene glycol, water, ethanol and the like.

The pharmaceutical preparation, if desired, can also contain minor amounts of wetting or eraulsifying aģents, or pH buffering aģents. These preparations can take the form of Solutions, suspensions, emulsion, tablets, pilis, 25 capsules, powders, sustained-release formulations and the like. The preparation can be formulated as a suppository, with traditional binders and carriers such as triglycerides. Oral formulation can include Standard carriers . such as pharmaceutical .grades of mannitol, 30 lactose, starch, magnesium' stearate, sodium saccharine, cellulose, magnesium carbonate, etc~. “Examples of suitable pharmaceutical carriers are described in "Reminton' s Pharmaceutical Sciences" by E.W. Martin. Such preparations will ' contain a therapeutically effective amount of the 35 . therapeutic, preferably in purified form, together with a 18 suitable amount of carrier so as to provide the form for proper administration to the patient. The formulation should suit the mode of administration.

The compositions of the present invention may be 5 administered transdermally. In one embodiment, a composition of the invention is directly applied to the skin. In another embodiment, a composition of the invention is applied to a reservoir (e.g. a cotton wool pad, a synthetic polymer such as Teflon^^, or any suitable 10 adsorbant) that is applied to the skin, preferably under an occlusive dressing. In s .specific embodiment, a composition of the present invention is applied to the skin_ by means of a dermal patch. The concentration of the activ-e therapeutic aģent in-the composition that is applied to the 15 skin, contained in or adsorbed onto the reservoir, or contained in the dermal patch may be about 10-100%, preferably at least 50%, more preferably at least 90%.-

In a preferred embodiment, a polar eompound such as DMF is administered transdermally using any suitable drug 20 delivery device, for example by applying one or more dermal patches. The dermal -patch may optionally comprise a ~ backing, a reservoir such as an adsorbant impregnated with a polar eompound of the invention, and a perraeable membrane that is placed in contact with the skin. The backing may 25 be of any material, such as a natūrai or synthetic polyraer, that resists Chemical attack bv the polar eompound. Especially preferred is a backing of high density polyethylene. The adsorbant may be a colloidal substance, for instance diatcmaceous earth or colloidal Silicon 3Ό* aioxide. The permeable membrane may be - of any material that chemically resists the polar eompound and may optionally be provided with pores. In a preferred embodiment, the permeable membrane is a Teflon^ membrane having pores with a diameter of about 0.1 μτη,' a diameter of 35 about 0.5 /xm, or a diameter within the range of frem about 19 19LV 12490 0.1 /*m to about 0.5 μιη. The patch may be self-adhesive or tnay be held in contact with the skin by an applicator, such as a vrapping or bandage, including without limitation an elastic bandage or an adhesive bandage; an Elastoplast bandage is suitable for this purpose. Preferably, the patch contains a greater quantity of polar compound than is intended to be delivered through the skin of the patient to be treated; for instance, the patch may contain about 50% more of the polar compound than is intended to be delivered. The patch may be of any desired size and shape, and may for instance take the form of a disk approximately 9cm in diameter.

In one_ etnbodiment, the patch cotnprises a polar compound, such as DMF, and at least one other pharmacologičally active aģent, for instance an local anti-irritant such as glycerine. In another etnbodiment, the patch cotnprises a polar compound, such as DMF, and has no other pharm'acologically active aģent present. In a further : etnbodiment, the patch comprises a polar compound, such as DMF, ' and has present no other pharmacologically active aģent that is capable of being systemically absorbed through" the skin, or has-present no other pharmacologically active aģent in an mount that is systemically effective after transdermal absorption. In yet another etnbodiment, the patch comprises a polar compound, such as DMF, and has present no other systemically active pharmacological aģent. In yet a further etnbodiment, the patch comprises a polar compound, such as DMF, and has present no other antiviral aģent, for instance acyclovir or arildone.

The present invention thus provides a dermal _patch-comprising a polar compound, such as DMF, in an amouht that is effective for treating a virai or microbial infection (for example an infection with HIV) in a human adult or a human child. The present invention further provides a dermal patch comprising a polar compound, such as DMF, in 5 20 an mount that is effective for treating a wasting syndrome in a human adult or a human child. In one embodiment, the patch contains at least 0.25g of a polar compound such as DMF, preferably at ieast 0.5g, more preferably at least lg, and stili more preferably at least 5g, for instance 5-15g of a polar compound such as DMF. 10 15 20

To minimizē evaporative loss of the polar compound, the patch may optionally be stored in a sealed Container, such as a sealed polymer bag. If desired, each patch mav be individually sealed for convenience of use. Optionally, the patches may be rafrigerated prior to use, for instance _at about 4°C, so as to reduce evaporative loss of the polar compound. Preferably, the patches are prepared within 24 hours of use and are stored in a sealed Container at 4°C; however, patches are stable for one or more weeks when stored in a sealed Container at _4°C. Prior to applying a patch, an arear of skin is preferably washed with-a mild soap and water, rinsed to remove soap residue, thoroughly dried, and then hyarated with a suitable skin lubricant and moistening aģent, such as jelly. The patch is then 25 removed from the packing material and applied so that the permeable metribrane contacts the prepared skin surface. The patch may be held in place with an applicator. After a desired dosage period, the patch is removed. After removal of the patch, the treated area may be thoroughly washed with a mild scap and water to remove any residue of the polar compound. 30 35 such

The compositions of the present invention -may be administered at any desired - interval, for instance cnce everv two or three weeks; once, twice or three times per week; every šecond day,· or daily. Preferablv, a compcsition of the invention is administered in a dose that results in a peak ņlasma Ievel of about 2-200 mg/1, more pre£erably about 100-200 mg/1, stili more pre£erably about 150 mg/1 of the active ingredient of the compcsition, as DMF. Especially preferred is a peak plasma Ievel of 100-150 mg/1 or 150-200 mg/1 of DMF. As used herein, the tertn "ppm" refers to parts per million by weight, and is in practice equivalent to mg/1.

For transdermal administration of a polar compound, the rāte of absorption is determined by the skin of the subject. Upon exposure to the human skin, liguid DMF is absorbed at a steady-state rāte of approximately 9.4 mg/cm^/hour (see Mraz and Nohova, 1992, Occup. Env. Health 64:85-92) . Accordingly, the desired rāte of absorption may be achieved by controlling the surface area of the skin exposed to the drug, as by determining the area of each patch and the number of patches applied to the skin. For example, two patches of diameter 9 cm will expose a total skin surface area of 127 cm? to the polar compound; for DMF, this will result in an absorption rāte of about 1.2 g of DMF per hour. An initial dose of about 15 mg/kg of DMT is especially preferred.

In one embodiment, a patient weighing about 72 kg is initially treated with DMF by administering two 9 cm diameter dermal patches for one hour, resulting in an initial adsorbed dose of about 1.2 g of DMF, equivalent to about 16.7 mg/kg. The number of patches may be scaled up or down, and a longer or shorter initial period of exposure used, according to the body weight of the patient being treated. This starting dose may be repeated at any desired interval, as described above, and preferably is given at weekly intervāls. Preferably, the patient.is monitored for at least 72 hours after administration of DMF for signs of toxicity, such as liver toxicity, for example by monitoring the serum or plasma Ievels of enzymes such as aspartate aminotransferase (AST), alanine arainotransferase (ALT) , γ-glutamyl transferase (γ(?Γ) and alkaline phosphatase, proteīns such as albumin, or substances such as conjugated or_unconjugated bilirubin. Preferably, serum or plasma 22

Ievels of AST and ALT do not exceea five times the upper limit of normai as determined by the reference range of the laboratory and population in cruestion; more preferably serum or plasma Ievels of AST and ALT do not exceed three 5 times the upper limit of normai, most preferably AST and ALT are not elevated above normai Ievels or above pre-: treatment Ievels. The dose of DMF may be escalated by applying the patches for successively longer periods; in one embodiment the patches are applied for two hours-, then ļLO four hours, then six hours, and so on; in another embodiment, the period of exposure is successively doubled. Preferably, the patient is monitored prior to any dose escalation, in order to detect any signs of toxicity. If desired, the dose may be escalated at weekly intervāls. ;15 The dose may be escalated in this manner until the dose is calculated to be about 150 mg/kg/dose; or until a desired peak plasma Ievel, for instance 100-150 or 150-200 mg/1 is achieved; or until the period of exposure is about 6 hours or "about 8 hours. One should be cautious before increasing ~20 the dose above 240 mg/kg/dose.

6. -- EZAMPLE: TREATMENT O.F HIV INFECTION BY TRANSDERKAL 25 ĀDMINISTRATION OF Ν,Ν' -DIMETHYL?QRMAMIDE (DMF) Patients infected with HIV-1 were treated with dimethylformamide (DMF) by the application of skin patches imņregnated with a dimethvlfarmamide cel to the patient's body. N-acetyl-cysteine-glutathione and/or essential phospholipids were administered (orally or intravenously) to the patients at a dcsage-of 250 mg to 300 mg daily as a liver booster. Instead or in addition. glutamine may also be administered to the patient as a liver booster.

Twa skin patches were applied to different parts of the patient's body, for example to the forearm. Each skin patch contained about 7,06g of gel comprising DMF (92,5 % m/m) and colloidal silicone dioxide (7,5 % ra/m). The gel 30 23 LV 12490 served to prevent leakage of liquid DMF from the patches. The patches were manufactured at most 12 hours prior to use as DMF evaporates rapidly. The intended Ievel of DMF in the patient's blood was 1Q0 ppm. For a patient weighing 5 about 60 kg, an amount of about 14g over a period o£ 12 hours is required to producē a Ievel of lOOppm. Based on studies by Marz and Nohova the surface area required to n absorb this amount xn about 127,2 cm. To obtain a blood Ievel of 100 ,ρρτη about 1,272 g of DMF must be absorbed per -10 hour, thus each sticker reguires about 7,064 g DMF to deliver the reguired amount, having a surface area of 6,36 cm? (each*sticker) . DMF absorption rāte is 9,4 mg/cm?/hour.

In theory this treatment will deliver 125 - 135 ppm, but due to evaporation of the DMF, 100 ppm was obtained. 15 Absorption capability variēs from patient to patient depending on factors such as skin-type and skin thickness. To obtain the desired Ievels of DMF in the patients, plasma DMF concentrations were monitored for each patient and treatment adjusted accordingly depending on the DMF Ievel 20 of each patient. Figurē 1 shows the plasma DMF Ievels of 4 patients treated with 2 dermal DMF patches for 8 hours. Figurē 2 shows the plasma DMF Ievels of 3 patients treated with 2 dermal DMF patches for 6 hours.

The stickers were thus each loaded with about 7,064 g 25 of the gel of DMF and silicone dioxide. Each patch was applied for a period of 12 hours, either once per week over a period of 12 weeks, or twice per week over a period of 6 weeks.

Blood tests in certain patients indicate an increase 30. in CD4 T-cell counts from 350 to 1000, and a rapid reduction of PCR (Polymerase Chain reaction) (virai load) of 120,000 to 500/ml within three weeks with as little as three treatments. Figurē 3 shows serial quantitative PCR measurements of HIV-l virai load in a patient prior to treatment and following three treatments with 2 dermal DMF 35 24 patches f or 12 hours. The PCR tēst conducted was the Roche amplicor HIV monitor. A virai load of < 500/ml plasma is considered to be undetectable.

Some patients undergoing treatment had severe acne or 5 displayed German measles symptoms before treatment. When treated with DMF so that the DMF Ievel in the patient's blood was 50 - 10 0 ppm, the German measles syitvptoms and the severe acne cleared up or disappeared within 7 days.

Prior to the treatment with dimethylformamide; a 10 comprehensive base-line clinical and psychological evaluation of the patient was conducted. The evaluation provided baseline biochemical and hematological data on the patient. Detailed virologičal serology (HIV-l) tests were also conducted to determine the patient's total body virus 15 count, and these tests were conducted on a weekly basis, or as per treatment.

The concentration of DMF in the patient's blood was determined hourly during the_ period of treatment. An intravenous line was introduced each morning to take blood 20 samples and was ķept open with an infusion of Normai Saline at a rāte of 20ml/hour and daily monitoring of the active metabolite AMCC (eg by 4-hourly urine sampling) derived from the DMF was also conducted. Subsequent applications of DMF were adjusted in accordance with measured changes in 25 blood Ievel DMF concentration resulting from changes in absorption variables and daily full haematological and biochemical profilēs were conducted to detect any changes in liver function. Daily full clinical and psychological evaluations_were also conducted. 30 A—daily virologičal serology workout to establish total body virus count and to monitor improvements in the immune status of the patient (CD4- T-helper celis) and prognostic factors were also conducted. The serology workout was based on p24 antigen and ouantitative PCR or, optionally, by other methods. A weekly determination of 35 25 5 10 15 20 25 LV 12490 CD4 counts and Beta-2-macroglobulin was also conducted specifically to monitor improvements in the patient's immune status and prognosis. Ali clinical and laboratory data were fed into a centralized data system to facilitate rapid response to any detrimental change so as to curtail fereatment to maximize clinical effect and minimizē potential side effects. The tests included a) Serum: Na, K, Cl, C02, Urea, Urate, Creatinine, Ca, Mg, Phosphate, Total and Conjugated Bilirubin,· b) · Hematology: Hemoglobiņ, Red celi count, Hematocrit, MCV, MCH, MCHC, RDW; c) Serum Protein Electrophoresis: Total Proteins, Albumin, Total Globulin, Alphal Globulin, Alpha2 Globulin, Beta Globulin, Gamma Globulin; d) White celi arialysis: Differential white celi count, Absolute Neutrophil, Lymphocyte, Monocyte, Eosinophil and Basophil counts; e) Liver Enzymes: Alk. Phos, Gamma GT, ALT (SGPT) , AST (SGOT)LDH; f) Others Celi markers, PCR, Beta2-Mocroglobulin, p24 Antigen, C-Reactive protein, CK-MB concentration,· g) Blood analysis for DMF Ievels; h) Urine analysis for AMCC Ievels

It appears as if DMF acts as at least one of a reverse -transcriptase inhibitor and a protease inhibitor. In vitro tests were conducted and it appears “that the solvent 30 =- ^properties of DMF dissolve the virus pārticies, e.g. the capsid. _ 7.

EXAMPLE: TREATMENT OF HIV INFECTION BY TRANSDERMAL 26 ADMINISTRATION OF N. N1-DIMETHYLFORMAMIDE (DMF) A pilot studv was conducted to evaluate the sfficacy of transdermal DMF in the treatment of patients infected with HIV. Informed consent was obtained frora each patient. 5 Seropositive status was verified by 'viestam blot using a commercial kit for detecting antibodies to p24 (Abbott Diagnostics) and the presence of HIV-1 was docutnented by quantitative PCR using a commercial kit (Roche Amplicor). DMF as obtained commercially (Sigraa-Aldrich) and in 10 plasma samples was analysed by mass —spectroscopy/ gas chromatography (GC/MS), using a Vārian 9600 gas chromatograph, OV 351 column, carbowax-PEG capillary column, and Finnegan Mat ITS40 ion trap detector. Operational parameters were as follov/s-: GC temperatūra 15 program: 60°C for 1 min. followed by a temperature gradient of 9.4°C/min. for 20 min. MS ionizationr method: electron impact; mass range: 40-80 mass units; 1 scan/sec; peak thresholds 3 counts/sec; background mass·. 69 mass units. A stock solution of dimethylacetamide was used as an 20 internai Standard. Ali samples were extracted with an organic solvent containing the internai Standard, allowed to precipitate for 30 min. in a ref rigerator, and centrifuged at 3000g for 5’ min. befcre transfer to GC vials for injection. Retention time of the DMF peak relative to 25 the internai Standard was 3.26 minūtes; the calibration curve showed a correlation of 0.98. Quantitation of DMF appeared linear up to 100 mg/1 and the lower limit cf detection was estimate at 0.5 mg/1 based on a signal-to-background ratio of 3:1. 3 0 _ Dermal patches 9 cm in diameter were used v/ithin 12_ hours' CLf manufacture. Each patch had a backing of high density polyethylene (0.245 g) , a permeable membrane of Teflon^^ (pore size 0.2 μτη; 0.268 g) for placement agair.st the skin, and containing 0.573 g of colloidai Silicon dioxide impregnated with 7.067 g DMF between the backing 35 27 10 15 20 25 30 LV 12490 and the permeable membrane. The patches were visually inspected to verify the absence of leakage and were weighed on an analytical balance to verify less than 10% deviation from a total mass of 8.153 g. The patches were applied to the skin of the forearm and secured with Elastoplast'^ bandages. Either one or two patches were used f or the initlal dose, as determined from the predicted skin transfer rāte of 9.4 mg/cm^/hr and the patient's body weight. A staged dose of DMF was used, with two-hourly blood and urine sampling to determine peak plasraa DMF Ievels. Patients were clinically monitored for any toxic side-effects and were ķept under comprehensive biochemical surveillance while the dosage was increased as necessary to achieve a peak plastna DMF Ievel of 100-120 ppm. Once the correct dosage was established, transdermal DMF was giveh once a week. Initial evaluation includeā daily — determination of t a) vital signs and body weight; b) clinical checklist and Kamofsky score,- a) full blood count and erythrocyte sedimentation rāte; b) serum urea, creatinine, glucose, sodium, potassium, ALT, AST, alkyline phosphatase and total bilirubin; c) Coulter analysis of CD4+ and CD8+ counts and CD4/CD8 ratio; d) Quantitative estimates of HIV-1 load by PCR (RoChe Amplicor) and analysis of antibody to p24 (Abbott); e) Urinalysis (Dipstix). At each weekly vīsit, the patients were evaluated for adverse effects, virtually ali these tests were repeated, ēind .blood and urine were collected for measurement of DMF and its metabolites. -55 28

Patient 1 [ADF] began the protocol in relatively good health, complaining principally of pains in the arms and legs and inability to sleep. Two DMF patch.es were ādministered once a week, for an average eacposure pericd of 5 8 hours . The average weekly dose of DMF was 6.11 g and the resulting peak plasma DMF Ievel average 75 mg/1. After 9 weeks, the patient's CD4+ T celi count had increased from 140 to 640 cells/μΐ and the PCR-measured virai load had decreased from 250,000 to 50,000 copies/ml. After 10 weeks 10 the patient's weight had increased from 81.9 to 96.0 kg, the patient was clinically well and no longer coraplained of pain in the limbs.

Patient 2 [AM] began the protocol complaining of loss of strength, inability to sleep, pains in the arms and 15 legs, and had herpes sores in the mouth. One DMF patch was administered per week, for an average exposure period of 8 hours. The average weekly dose of DMF v/as 7.12 g and the resulting peak plasma DMF Ievel averaged 125 m/f. After 9 weeks, the patient's CD4+ T celi count has increased from 20 460 to 720 cells/jtif, the PCR-measured virai load had decreased from 29,000 to 13,000 copies/mf, and the patient's weight had increased from 58.4 to 63.0 kg. The patient's herpetic sores had resolved, the limb pains had disappeared and the patient appearea clinicallv well. 25 Patient 3 [SM] began the protocol with overt clinical AIDS, and complained of respiratory difficulty. Two DMF patches were administered once a week, for an average exposure period of 8 hours. The average weekly dose of DMF was 8.97 g and the resulting peak plasma DMF Ievel averaged 121 mg/t. After 7 w.eeks, the patient's CD4+ T celi count had increased from 39 to 13a cells/μ?, the PCR-measured virai load had decreased from 222,000 to 160,000 copies/mč, and the patient's weight had increased from 74.2 to 100 kg. The patient's appetite had improved, the rsspiratcrv 30 29 29LV 12490 difficulties had resolved, the patient appeared clinically well and started exercising again.

Patient 4 [EM] began the protocol with secondary infections (including herpes), anemia, diarrhea and acne. 5 Two DMF patches were ādministered once weekly, for an average exposure period of 8 hours. The average weekly dose of DMF was-7.33 g and the resulting peak plasma DMF Ievel averaged 90 mg/£. After 18 weeks, the patient's CD4+ T celi count had increased from 249 to 450 cells/μ?, the 10 PCR-measured vijral load had decreased from 13,000 to 4.000 copies/mf, and the patient' s weight~had increased from 81.5 to 90.4 kg. The patient appeared clinically well and had no active medical complaints.

Patient 5 [SV] began the protocol complaining of poor" 15 appetite, forgetfulness, abdominal pain, and severe fatigue that prompted him to consider selling his interest in his business. Two DMF patches were administered once weekly, for an average ~exposure period of 6 hours. The average weekly dose of DMF was 3.75 g and the resulting peak plasma 20 DMF Ievel average 67 mg/1. After 5 weeks, the patient's CD4+ T celi'count had increased from 354 to 396 celis/μΐ, the PCR-measured virai load had decreased from 156,000 to 13.000 copies/mi, and the patient's weight had increased from 56.0 to 58.0 kg. The patient was clinically well, had 25 acquired his business partner's share, and was running the business himself.

Patent 6 [W] began the protocol complaining of secondary infections (including herpes), ataxia, and numbness of the left arra and left side of the faee. Two 30 DMF patches were administered weekly, for an average exposure period_ of 8 hours. The average weekly dose of DMF was 8.25 g-and the resulting peak plasma DMF Ievel averaged 110 mgfl. After 19 weeks, the patient's CD4+ celi count had increased from 260 to 450 celis/μ.^, the PCR-measured 35 virai load had decreased from 120,000 to 24,000 copies/m£. 30 and the patient's weight had increased from 75.4 to 34.5 kg. The secondary infections had resolved and tha patient appeared clinicallv well.

Patient 7 [AJF] began the protocol severely ill. The patient's initial CD4+ T celi count was 29 cell/μί, and the initial PCR-measured virai load was 1,156,000 copies/mč. Treatment was with 1 patch, for an average exposure period of 4 hours. The average weekly dose of DMF was 4.60 g and the resulting peak plasma DMF Ievel averaged 100 mg/č. After the first treatment, the patient's CE4+ T celi count decreased to 14 cells/μΖ. Treatment was given daily for five days and then continued at weekly intervāls. After 9 treatments, the'patient's CD4+ τ celi count had increased to 35 cells/41?, the PCR-measured virai load had decreased to 9,000 copies/m£,'and the patient's weight had increased from 46.5 kg (at commencement of DMF therapy) to 49.0 kg. The patient felt well and had returned to full--time employment.

Patient 8 [MS] began the protocol with severe herpetic lesions in the lower back and genitalia. Two DMF patehes were given once weekiy, for an average exposure period of 8 hours. . The average weekly dose of DMF was 6.24-g and the resulting peak plasma DMF Ievel averaged 130 mg/£. After 3 weeks, the patient's CD4+ T celi count had increased from 200 to 240 cells/μζ, the PCR-measured virai load had decreased from 1,200,000 to 250,000 ccpies/mŽ. The Datient' s weight had increzcod from 43.1 to 52.2 kg, and the Herpes lesions had entirely resolved-

Two patients were excluded from the trial, one due to alcohol abuse and one due to virai hepatitis B.

Most patients. esperienced mild local skin irritation at the area of application after removal of the patches; the skin at the application site had a maculopapular appearance, probably due to incense hydration under the patch. In one case, there v/as slight blistering that 31 31LV 12490 resolved within 24 hours and Chat did not cause the patient significant discomfort. Most patients experienced mild transient nausea, usually on day fchree after treatment, which gradually decreased during the treatment protocol; 5 one patient reported moderate transient nausea. Four patients showed transient elevation of liver enzymes, which never exceeded three times the upper limit of normai and which in most cases retumed to pre-treatment Ievels prior to the next dose-of DMF. Most instances of elevated liver 10 enzymes were assočiated with at least one factor unrelated to the treatment protocol (alcohol consumption, hepatitis, and previous anti-HIV therapy with other aģents) .

Virtually ali patients showed clinical improvement -after 2-3 weeks of treatment. As shown by Figurē 4, every 15 patient showed itnproved general condition after treatment with DMF when assessed according to the Karnofsky performance scale, in which a patient's general status is assigned a numerical'value as follows: 100 = normai," no complaints; 90 = able to carry on normai activities, minor 20 signs or symptoms of disease; 80 normai activity with effūrt; 70 « cares for self, unable to carry on normai activity or to do active. work; 60 = requires occasional assistance, but is able to care for most needs; 50 = reguires considerable assistance and frecjuent medical care; 25 40 » disabled, requires special care; 30 * severely disabled, hospitalization indicated although death not iraminent; 20 = very sick, hospitalization and active supportive treatment necessary,* 10 = moribund, fatal process progressing. rapidly; 0 = dead. (See Kanofsky. et 30 _al., 1984, Cancer 1:634-656). Improvement in neurological symptoms and Herpes virai infections was remarkablē. Additional anti-microbial therapy for secondary infections ~ was rarely needed. Viithin the first 14 days of DMF treatment, there was marked improvement in general fatigue and in appetite. Ali the patients gained weight. Clinical 35 32

improvement correlated well with disease status as assessed by virai load and CD4+ T celi count. For five out of eight patients, the relative PCR-measured virai load could be fitted to Gompertz curves; this analysis revealed an 33.3% 5 decline in PCR-measured virai load after 42 days of DMF treatment. For seven out of eight patients, the relative CD4+ T celi count could be fitted to Gompertz curves; this analysis showed a 73.4% increase in CD4+ T celi counts after 42 days of DMF treatment. 10 - The present invention is not to be limited in scope by the exemplified embodimeiits, which are intended as illustrations of single aspects of the invention. Indeed, various modifications of the invention in addition to those shov/n and described herein will become" apparent to those 15 skilled in the art from the foregoing description and accompanying drawings. Such modif ications are intended to fall within the scope of the appended claims.

Ali publications cited herein are incorporated by reference in their "entirety.

CLAIMS 1. A drug delivery device for transdermal administration of a therapeutic aģent, comprising: a reservoir containing or having absorbed thereon a therapeutic composition comprising N-hydroxymethyl-N-methylformanu.de (HMMF), N-hydroxymethylformamide (HMF) , dihydroxymethylformamide (DHMF), N-acetyl-S-(N- methylcarbamoyl) cysteine (AMCC)-, dimethylsulfoxide (DMSO) or pyridine-N-oxide. 2. A drug delivery device for transdermal administration of a therapeutic aģent, comprising: a backing; a permeable membrane suitable for placement against the sjcia of an animal to be treated; and an adsorbant situated between the backing and the permeable membrane, the adsorbant having -adsorbed thereon a therapeutic conņosition comprising N,N' -dimethylformamide (DMF) , N-hydroxymethyl-N-methylformamide (HMMF) , N-hydroxymethylformamide (-HMF), dihydroxymethylformamide (DHMF), N-acetyl-S-(N-methylcarbamoyl) cysteine (AMCC), N-methylformamide (HMF) , dimethylsulfoxide (DMSO) , formamide, acetamide, methylacetamide, dimethylacetamide, diethylacetamide, isopropylacetamide, diisopropylacetamide, N-acetylpiperidine, N- (0-hydroxyethyl) acetamide, N, N'-di(£-hydroxyethyl) acetamide, N-acetylmorpholine, acrylamide, propionamide, N-fluoromethyl-N-methyl-formamide, pyridine-N-oxide, or an amide of thē general formula R^-CO-NRļR^, in which R] and R2 are independently selected from the group consisting of H, methyl, halomethyl, saturated and 34 unsaturated C2-C- aikvi groups, and hydroxyIated alkvl groups or Rļ and R2 are together selected frcm the grouo consisting of (CK2>4., (C^)^, and (CE2) 20 (CH2) 2; and R3 is selected from the group consisting of K, methyl, and saturated and unsaturated C2-C3 aikyi groups; the drug delivery device adapted to be placed in contact with the skin of an animal to be treated. 3. The drug delivery device according to Ciaim. 2, in which the therapeutic composition comprises DMF. 4. The drug delivery device according to Ciaim 3, in which the therapeutic composition comprises at least 0.25 g of DMF.

The drug deliverv device according to Claim 4, in therapeutic’ composition comprises at least 5 g of 5. - which the DMF. 6. The drug delivery device according to any one of

Claims 3 to 5 iaciusiva, in «hich the therapeutic composition comprises DMF in an amounfc that is effective for treating a virai or microbiai infection or a wasting svndrome in a human aauit ar a human child.

~r .-i ^ J • -------------3 t. O SUV 3112 of inclusive, havinc* OlTSSeilt. no ’e aoent other than DM -? rrU — -Ϊ i · 1UC W.,

Claims 3 to 5 oharmacolcaicallv ac 8 . The drug delivery device according to any one of Claims 3 to inclusive, navinc prasent nc pnarmacologicallv activa aģent, other fchan DMF, that is capable of being svstemicallv absorbed thrcugh the skin. 35 LV 12490 9. The drug delivery device according to any one of Claims 3 to 8 inclusive, in which the adsorbant is colloidal Silicon dioxide and the permeable membrane is a Teflon™ membrane having pores of diameter from about 5 0.1/,um to about 0.5fim. 10. A therapeutic composition comprising N,N'-dimethylformamide (DMF) , N-hydroxymethyl Krītiet hylformamide_ (HMMF) , N-hydroxymethy 1 formamide (HMF) , 10 dihydroxymethylformamide (DHMF), N-acetyl-S-(N- methylcarbamoyl)cysteine (AMCC), M-methylformamide (NMF), dimethylsulfoxide (DMSO), formamide, acetamide, methylacetamide, dimethylacetamide, diethylacetamide, isopropylacetamide, diisopropylacetamide, N- ^ acetylpiperidine, N-(P~hydroxyethyl) acetamide, N,N'- di(p-hydroxyethyl)acetamide, N-acetylmorpholine, acrylamide, propionamide, N-fluoromethyl-N-methyl-formamide, pyridine-N-oxide, or an amide of the general formula R3-CO-NR]R2, in which 20 R, and R2 are independently selected from the group consisting of H, methyl, halomethyl, saturated and unsaturated C2 - C3 alkyl groups, and hydroxylated alkyl groups; or

Rj_ and R2 are together selected from the group consisting of (CH2)4, (CH2)j, and (CH2)20(CH2)2; and R3 is selected from the group consisting of H, methyl, and saturated and unsaturated C2 - C3 alkyl groups; for use in the treatment of a wasting syndrome in animal. 3 0

The therapeutic composition according to Claim 10, in which said therapeutic composition comprises an amide of the general formula R3-CO-NRiR2, in which 36

Ri and R2 are independently selected from the group consisting of H, methyl, halomethyl, saturated and unsaturated C2 - C3 alkyl groups, and hydroxylated alkyl groups; or 5 R: and R2 are toget’ner selected from the group consisting of (CH2)4, (CH2)5 and (CH2) 20 (CH2) 2; and

Rj is selected from the group consisting cf H, methyl, and saturated and unsaturated C: - C3 alkyl groups. 10 12. The therapeutic composition according to Claim 10 or Claim 11, in which said therapeutic composition comprises an amide of the general formula R3-CO-NRiR2< in which Rļ and R2 are independently selected from the group 15 consisting of H, methyl, halomethyl, saturated and unsaturated C2 - C3 alkyl groups, and hydroxylated alkyl groups; and R3 is selected from the group consisting of H, methyl, and saturated and unsaturated C2 - C3 alkyl 20 groups, and hydroxylated alkyl groups. 13. The therapeutic composition according to any one of Claims 10 to 12 inclusive, in which said therapeutic composition comprises DMF, HMMF, HMF, DHMF, AMCC, NMF, 25 DMSC, formamide, acetamiae, methyiacetamide,

Cti.rr 0 - 'n'f J_ 3. C S12.rai CL0 , 0.i. Θ tlrvi3.C0 “ 3.iril.Q.0 , 2. S Ο Ο I3.C B13Π1.19 , diiscpropvlacetamide, N-acetyipiperidine, Ν-β- acetamide, M-acetyimorphcline, acrvlamide, propicnamide, " ' M-fluorGmethyl-N-methyl-formamide, or pyridine-U-oxide. 14. The therapeutic composition according to Claim 13, in which said therapeutic composition comprises DMF, ΗΪΊΗ?, HMF, DHI4F, AMCC, or NI4F. 37 LV 12490 15. The therapeutic composition according to any of • Claims 10 to 14 inclusive, further comprising at least one other aģent effectiva for treating HIV infection. 5 16. The therapeutic composition according to Claim 15, further comprising at least one nucleoside analog reverse transcriptase inhibitor, at least, one non-nucleoside reverse transcriptase inhibitor, or at least one protease inhibitor. 10 17. The therapeutic composition according to any one of Claims 14 to 16 inclusive, in wnich said therapeutic composition comprises DMF. 15 18. The therapeutic composition according to any one of

Claims 13, 15 or 16, in which the therapeutic composition comprises DMSO. 19. The therapeutic composition according to any one of 20 Claims 10 to 18 inclusive, in which said therapeutic composition is administered transdermally. 20. A therapeutic composition comprising N-hydroxymethyi-N-methylformamide (HMMF) , N-hydroxymethylformamide (HMF) , dihydroxymethylformamide (DHMF), N-acetyl-S-M-meohvlcarbamovijrvsteine (AMCC), dimechylsulfoxide (DMSO) or pyridir.e-N-oxide for use in the treatment of a virai cr micrcbial infection in animal. 21. A therapeutic composition as claimed in Claim 20, in which said therapeutic compositions is administered for treatment of a retroviral infection. 38 22. A therapeutic composition as claimed in Claim 21, ir which the retroviral infection is an HIV infection. 23. A therapeutic composition acccrding to ar.v cne c* Glaims 20 to 22 inclusive, further comprising at least cne other aģent effective for treating KIV infection. 24. A therapeutic composition as claimed in any cne cf Claims 20 to 23 inclusive, further comprising at least one nucleoside analog reverse transcriptase inhibitcr, at least one non-nucleoside reverse transcriptase inhibitcr, or at least one protease inhibitor. 25. A therapeutic composition as claimed in any cne of Claims 20 to 24 inclusive, in which said cherapeutic aģent is administered transdermallv. 26. A vaccine prepared from antibodies harvested from the body of an animal treated for a virai infection according to the therapeutic composition of Claim 20. LV 12490 1/4 PLASMA GMF CONCENTRATION ' patches per patient = 2 for 8 hours

FIG 1 LV 12490 2/4 PLASMA DMF CONCENTRATIOM patches per 'patient =* 2 for B hours

FIG 2 LV 12490 3/4

HGcJ 120.000 LV 12490 4/4

g LL

Claims (17)

1. A medicament delivery device for percutaneous administration of a therapeutic agent comprising: a reservoir (matrix) containing or on which the surface (matrix) is adsorbed a therapeutic composition comprising N-oxymethyl-N-methylformamide (HMMF), N -oxymethylformamide (HMF), dioxymethylformamide (DHMF), N-acetyl-S- (N-methylcarbamoyl) cysteine (AMCC), dimethylsulfoxide (DMSO) or pyridine N-oxide. A drug delivery device for percutaneous administration of a therapeutic agent comprising: a base (layer); a permeable membrane suitable for being placed on the skin of the treated animal; The adsorbent between the substrate and the permeable membrane is adsorbed onto the adsorbent surface by a therapeutic composition comprising N, N'-dimethylformamide (DMF), N-oxymethyl-N-methylformamide (HMMF), N-oxymethylformamide (HMF), dioxymethylformamide (DHMF ), N-acetyl-S- (N-methylcarbamoyl) cysteine (AMCC), N-methylformamide (NMF), dimethylsulfoxide (DMSO), formamide, acetamide, methylacetamide, dimethylacetamide, diethylacetamide, isopropylacetamide, diisopropylacetamide, N-acetylpiperidine, N- (p-oxyethyl) acetamide, Ν, Ν < Η (β-oxyethyl) acetamide, N-acetylmorpholine, acrylamide, propionamide, N-fluoromethyl-N-methylformamide, pyridine N-oxide or amide of general formula R3-CO- NR1R2, where: R1 and R2 are independently selected from the group consisting of hydrogen, methyl, halogenyl, saturated and unsaturated C2-C3 alkyl and hydroxylated alkyl: or R1 and R2 taken together are selected from the group consisting of - (CH2) 4 -, - (CH2) 5- and - (CH2 ) 20 (CH2) 2-groups; and R3 is selected from the group consisting of hydrogen, methyl and saturated and unsaturated C2-C3 alkyl; drug delivery product is suitable for contact with the skin of the treated animal. 3. The drug delivery product of claim 2, wherein the therapeutic composition comprises DMF. The drug delivery product of claim 3, wherein the therapeutic composition comprises at least 6.25 g of DMF. 5. The drug delivery product according to claim 4, wherein the therapeutic composition comprises at least 3 g of DMF. The drug delivery product according to one of claims 3 to 5. characterized in that the therapeutic composition comprises DMF in an amount effective for treating viral and microbial infections or debilitating syndromes in adults and children. The drug delivery product according to one of claims 3 to 6. with no pharmacologically active agent other than DMF. Product of administration according to one of claims 3 to 7. with no pharmacologically active agent other than DMF; wherein the DMF is able to systematically absorb through the skin. 9. The drug delivery product according to one of claims 3 to 8. characterized in that the adsorbent is a colloidal silicon dioxide and the permeable membrane is a Teflon Teflon ™ membrane with a pore diameter of 0.1 μιτι to 0.5 μιτι. A therapeutic composition comprising Ν, Ν'-dimethylformamide (DMF), N-oxymethyl-N-methylformamide (HMMF), N-oxymethylformamide (HMF), dioxymethylformamide (DHMF), N-acetyl-S- (N-methylcarbamoyl) ) cysteine (AMCC), N-methylformamide (NMF), dimethylsulfoxide (DMSO), formamide, s, acetamide, methylacetamide, dimethylacetamide, diethylacetamide, isopropylacetamide, diisopropylacetamide, N-acetylpiperidine, N- (P-oxyethyl) acetamide, N, N '-di (p-oxyethyl) acetamide, N-acetylmorpholine, acrylamide, propionamide, N-fluoromethyl-N-methylformamide, pyridine N-oxide or amide of the general formula R3-CO-NR1R2, wherein: selected from the group consisting of hydrogen, methyl, halo, saturated and unsaturated C2-C3 alkyl and hydroxylated alkyl: or R1 and R2 taken together are selected from the group consisting of - (CH2) 4-, - (CH2) s- and - (CH 2) 20 (CH 2) 2 -; and R3 is selected from the group consisting of hydrogen, methyl and saturated and unsaturated C2-C3 alkyl; used in animals for the treatment of slimming syndrome. 11. A therapeutic composition according to claim 10, wherein said therapeutic composition comprises an amide of the general formula R3-CO-NR1R2, wherein: R1 and R2 are independently selected from the group consisting of hydrogen, methyl, halogenomethyl , saturated and unsaturated C 2 -C 3 alkyl groups and hydroxylated alkyl groups: or R 1 and R 2 taken together are selected from the group consisting of - (CH 2) 4 -, - (CH 2) s - and - (CH 2) 20 (CH 2) 2 -; and R3 is selected from the group consisting of hydrogen, methyl and saturated and unsaturated C2-C3 alkyl. 12. A therapeutic composition according to claim 10 or 11, wherein the composition is: that this therapeutic composition contains an amide of the general formula R3-CO-NR1R2, wherein: R1 and R2 are independently selected from the group consisting of hydrogen, methyl halogenmethyl, saturated and unsaturated C2-C3 alkyl and hydroxylated alkyl; and R3 is selected from the group consisting of hydrogen, methyl and saturated and unsaturated C2-C3 alkyl and hydroxylated alkyl. 13. A therapeutic composition according to any one of claims 10-12. characterized in that the therapeutic composition comprises DMF, HMMF, HMF, DHMF, AMCC, NMF, DMSO, formamide, azetamide, methylacetamide, dimethylacetamide, diethylacetamide, isopropylacetamide, diisopropylacetamide, N-acetylpiperidine, N- (P-oxyethyl) ) acetamide, N, N'-di (p-oxyethyl) acetamide, N-acetylmorpholine, acrylamide, propionamide, 4 N-fluoromethyl-N-methylformamide or pyridine N-oxide. 14. The therapeutic composition of claim 13, wherein said therapeutic composition comprises DMF, HMMF, HMF, DHMF, AMCC or NMF. 15. A therapeutic composition according to any one of claims 10-14. further comprising at least one other effective agent for treating HIV infection. 16. The therapeutic composition of claim 15, further comprising at least one nucleoside analogue reverse transcriptase inhibitor, at least one non-nucleoside reverse transcriptase inhibitor, or at least one protease inhibitor. 1 A therapeutic composition according to any one of claims 14-16. characterized in that the therapeutic composition comprises DMF. 18. A therapeutic composition according to claim 13, 15 or 16, wherein the therapeutic composition comprises DMSO. 19. A therapeutic composition according to any one of claims 10-18. characterized in that the therapeutic composition is administered percutaneously. A therapeutic composition comprising N-oxymethyl-N-methylformamide (HMMF), N-oxymethylformamide (HMF), dioxymethylformamide (DHMF), N-acetyl-S- (N-methylcarbamoyl) cysteine (AMCC), dimethylsulfoxide (DMSO) or pyridine N-oxide used to treat viruses or microbial infections in animals. 21. A therapeutic composition according to claim 20, wherein said therapeutic composition is administered to treat a retroviral infection. 22. A therapeutic composition according to claim 21, wherein the retroviral infection is an HIV infection. 5 LV 12490 23. A therapeutic composition according to any one of claims 20-22. additionally containing at least one other agent for treating an effective HIV infection. 24. A therapeutic composition according to any one of claims 20-23. containing at least one nucleoside analogue reverse transcriptase inhibitor, at least one non-nucleoside reverse transcriptase inhibitor or at least one protease inhibitor. 25. A therapeutic composition according to any one of claims 20-24. characterized in that the therapeutic composition is administered percutaneously. 26. A vaccine obtained from an antibody of an animal organism is treated with a viral infection with a therapeutic composition according to claim 20.