Porphyrin antiviral compositions

Compositions for the treatment or prevention of AIDS or other diseases resulting from infection with the Human Immunodeficiency Virus containing one or more porphyrins. Porphyrins are tetrapyrrole macrocyle compounds with bridges of one carbon joining the pyrroles. Porphyrins occur naturally and are made synthetically. Derivatives of porphyrins include porphyrins with one or more substituents on one or more of the rings, porphyrins in which the conjugation of the ring has been altered by addition of substituents, porphyrins in which one or more center nitrogens is attached to substituents such as metals, liganded metals, and organic moieties, metalloporphyrins and metalloporphyrin-ligand complexes. Examples of natural and synthetic, positively, negatively, and neutrally charged porphyrins and porphyrin derivatives have been found to exhibit selective anti-HIV activity which is not dependent on the presence of light. Most of the compounds did not inhibit replication of herpes simplex virus type 1 or type 2 or Coxsackievirus strain B4. Effective concentrations for inhibition of HIV-1, as measured in vitro by inhibition of replication range from 0.01 to greater than 100 .mu.M. None of the compounds were toxic to unifected PBM cells.

This application relates to the field of anti-viral compounds, specifically 
porphyrin compounds for the treatment of human immunodeficiency virus. 
AIDS, or acquired immunodeficiency disease, is characterized by an 
imbalance in two basic types of immune system cells, helper/inducer T 
lymphocytes and suppressor T lymphocytes, with the ratio of suppressor 
cells to helper/inducer cells greatly elevated. Helper/inducer T cells, 
defined by a surface antigen called CD4, are responsible for the induction 
of most of the functions of the human immune system, including the humoral 
immune response involving the production of antibodies by B lymphocytes 
and the cell-mediated response involving stimulation of cytotoxic T cells. 
A condition associated with HIV is AIDS-related complex, or ARC. Most 
patients suffering from ARC eventually develop AIDS. 
Two related retroviruses can cause AIDS, human immunodeficiency virus type 
1 and type 2 (HIV-1 and HIV-2, generally referred to herein as HIV). The 
genomes of the two viruses are about 50% homologous at the nucleotide 
level, contain the same complement of genes, and appear to attack and kill 
the same human cells by the same mechanism. Also known as LAV 
(lymphadenopathy-associated virus), HTLV-3 (human T-lymphotropic 
virus-type 3), and ARV (AIDS-related virus), HIV-1 was identified in 1983. 
Virtually all AIDS cases in the U.S. are associated with HIV-1 infection. 
HIV-2 was isolated in 1986 from West African AIDS patients. 
Both types of HIV are retroviruses, in which the genetic material is RNA 
rather than DNA. The viruses carry with them a polymerase (reverse 
transcriptase) that catalyzes transcription of viral RNA into 
double-helical DNA. The viral DNA can exist as an unintegrated form in the 
infected cell or be integrated into the genome of the host cell. As 
presently understood, the HIV enters the T4 lyphocyte where it loses its 
outer envelope, releasing viral RNA and reverse transcriptase. The reverse 
transcriptase catalyzes synthesis of a complementary DNA strand from the 
viral RNA template. The DNA helix then inserts into the host genome where 
it is known as the provirus. The integrated DNA may persist as a latent 
infection characterized by little or no production of virus or 
helper/inducer cell death for an indefinite period of time. When it is 
transcribed by the infected lymphocyte, new viral RNA and proteins are 
produced to form new viruses that bud from the cell membrane and infect 
other cells. 
No treatment capable of preventing or reversing the immunodeficiency of 
AIDS or ARC is currently available. All patients with opportunistic 
infections and approximately half of all patients with Kaposi's sarcoma 
have died within two years of diagnosis. Attempts at reviving the immune 
systems in patients with AIDS have been unsuccessful. 
A number of compounds have apparent antiviral activity against this virus, 
including HPA-23, interferons, ribavirin, phosphonoformate, ansamycin, 
suramin, imuthiol, penicillamine, carbovir, 3'-azido-3'-deoxythymidine 
(AZT), and other 2',3'-dideoxynucleosides, such as 2',3'-dideoxycytidine 
(DDC), 2',3'-dideoxyadenosine (DDA), 2',3'-dideoxyinosine (DDI), 
3'-azido-2',3'-dideoxyuridine (CS-87), 
2',3'-dideoxy-2',3'-didehydrocytidine (D4C), 
3'-deoxy-2',3'-didehydrothymidine (D4T) and 
3'-azido-5-ethyl-2',3'-dideoxyuridine (CS-85). 
Inhibitors of cellular processes will often limit viral replication. 
Unfortunately, they are also usually toxic for the host and therefore 
cannot be prescribed for a prolonged period of time because of their 
toxicity. Although AZT is the drug of choice at this time for the 
treatment of AIDS, preliminary results indicate that AZT exhibits toxicity 
in a clinical setting, causing bone marrow suppression, resulting in 
anemia and neutropenia. See Yarchoan et al., Lancet 575-580 (1986). 
Efforts to decrease the problem of toxicity have primarily been directed 
towards finding selective, less toxic drugs. Due to the exorbitant cost of 
the nucleoside type drugs, research has also been centered around 
compounds which are relatively easy and economical to manufacture. 
Photolysis using hematoporphyrins has been extensively studied for the 
treatment of tumors, atherosclerotic plaque, and viral infections. 
Hematoporphyrin, a pigmented, iron-free natural derivative of hemoglobin, 
has cytotoxic activity in light. Studies to date indicate that most 
porphyrins and many metalloporphyrins are safe, as summarized by Kappas 
and Drummond, J. Clin. Invest. 77,335-339 (1986). 
Photodynamic therapy (PDT) is the treatment of malignant tumors with 
photosensitizers and light. General reviews of photodynamic therapy are 
Gomer, C. J. et al. Proceedings of the Clayton Foundation Conference on 
Photodynamic Therapy. Photochem. Photobiol. 46, 561-952 (1987) and 
Dougherty, T. J. Photochem. Photobiol. 45, 879-889 (1987). Briefly, 
certain photosensitizers, including porphyrins and metalloporphyrins, 
localize preferentially in tumor cells. Irradiation of the tissue results 
in selective cell death of the cells carrying the photosensitizer. The 
photochemistry and photophysics of porphyrins and metalloporphyrins have 
been studied in detail. Processes observed include radiationless decay to 
ground, loss of an axial ligand, energy transfer, electron transfer, 
formation of singlet oxygen, phosphorescence and fluorescence. The 
photoprocesses observed in each system depend greatly on the central metal 
atom (2H for porphyrin), the oxidation state of the metal and axial ligand 
bound to the metal. A weaker dependence of the photophysical properties on 
the nature of the macrocycle is observed. 
Porphyrins have been reported to have a variety of other biological 
activities. However, relatively little has been done with them with 
respect to in vivo clinical applications other than in photodynamic 
therapy. 
Perlin, et al., Antiviral Res. 7,43-51 (1987), recently reported that, upon 
exposure to light, hematoporphyrin, at concentrations as low as 0.5 
.mu.g/ml, inhibits in vitro replication of influenza A and herpes simplex 
viruses, but not several other viruses. See also Lewin, et al., Proc. Soc. 
Exper. Biol. Med. 163, 81-90 (1980) and Schnipper, et al., J. Clin. 
Invest. 65, 432-438 (1980). Hematoporphyrin in combination with visible 
light also inhibits reverse transcription in vitro by the RNA-dependent 
DNA polymerase of Moloney leukemia virus from an exogenous templete, as 
described by Munson, et al., Res. Commun. Chem. Pathol. Pharmacol. 
16,175-178 (1977). Inhibition does not occur in the absence of exposure of 
the cells or viruses to light. 
Within the last few months, hemin, ferric chloride protoporphyrin IX, has 
been shown to exhibit selective anti-viral activity under certain 
conditions. Tsutsui and Mueller demonstrated in Biochem. Biophys. Res. 
Com. 149(2),628-634 (Dec. 1987) that the reverse transcriptase activity of 
Rauscher murine leukemia virus, but not of avian myeloblastosis virus, was 
inhibited by hemin at a concentration of 10.sup.-4 M. They proposed that 
the hemin inhibited the enzyme activity by reversible, non-covalent 
interaction with the protein, not the template. Bhattacharya, et al., 
Proc. Natl. Acad. Sci. USA 78(5),2683-2687 (1981) showed that hemin 
differentially inhibits three forms of DNA polymerase .alpha., supporting 
the theory that hemin may be an important modulatory protein. None of 
these researchers correlated inhibition with toxicity, however, nor 
examined the compounds as selective antiviral drugs. 
It is therefore an object of the present invention to provide compounds 
having selective activity against Human Immunodeficiency Virus. 
It is a further object of the present invention to provide compounds having 
antiviral activity which are relatively non-toxic. 
It is a still further object of the present invention to provide compounds 
having antiviral activity which are relatively inexpensive and easy to 
manufacture. 
SUMMARY OF THE INVENTION 
Compositions for the treatment of AIDS or other diseases resulting from 
infection with the Human Immunodeficiency Virus containing one or more 
porphyrins, and method for treatment of patients using the compositions. 
Porphyrins are tetrapyrrole macrocyle compounds with bridges of one carbon 
joining the pyrroles. Many porphyrins are isolated from nature, for 
example, protoporphyrin, and many porphyrins are made synthetically, for 
example, those synthesized by condensation of aldehydes and pyrroles such 
as tetraphenylporphyrin. Derivatives of porphyrins include porphyrins with 
one or more substituents on one or more of the rings, porphyrins in which 
the conjugation of the ring has been altered by addition of substituents, 
porphyrins in which one or more center nitrogens is attached to 
substituents such as metals, liganded metals, and organic moieties, 
metalloporphyrins and metalloporphyrin-ligand complexes. 
Examples of natural and synthetic, positively charged, negatively charged, 
and neutrally charged porphyrins and porphyrin derivatives, have been 
found to exhibit selective anti-HIV activity. The activity is not 
dependent on the presence of light. The compounds did not inhibit 
replication of herpes simplex type 1 or type 2 or Coxsackievirus strain 
B4. Effective concentrations for inhibition of HIV-1, as measured in vitro 
by inhibition of replication range from 0.01 to greater than 100 .mu.M. 
Compounds which are useful are those having a therapeutic index of greater 
than 10, toxicity at levels of greater than or equal to 100 .mu.M, and 
antiviral activity at concentrations of less than or equal to 10 .mu.M. 
None of the compounds were toxic to uninfected PBM cells at 100 .mu.M.

DETAILED DESCRIPTION OF THE INVENTION 
Porphyrins are tetrapyrrole macrocycle, compounds with bridges of one 
carbon joining the pyrroles (FIG. 1A). There are many different classes of 
porphyrins. Some porphyrins are isolated from nature, for example, 
protoporphyrin IX, which is the organic portion of hemin. Many derivatives 
of natural porphyrins are known. Many porphyrins are synthesized in the 
laboratory. These include those made via the condensation of aldehydes and 
pyrroles, such as tetraphenylporphyrin (FIG. 1B). They also include 
porphyrins built up from smaller organic fragments. All porphyrins can 
have substituents off any of the positions of the ring periphery, 
including the pyrrole positions and the meso (bridging one carbon) 
positions as well as the central nitrogens. There can be one or more 
substituents, and combinations of one or more different substituents. The 
substituents can be symmetrically or unsymmetrically located. The 
substituents, as well as the overall structure, can be neutral, positively 
charged or negatively charged. Charged structures have counterions, and 
many counterions and combinations of counterions are possible. Porphyrins 
can be covalently attached to other molecules, for example a cyclodextrin 
(Gonzalez, M. C.; Weedon, A. C. Can. J. Chem. 63, 602-608 (1985)). They 
can have an attached molecular superstructure. The conjugation of the ring 
can be altered by addition of one or more substituents. Metals can be 
inserted into the porphyrin. Metals include but are not limited to Fe, Co, 
Zn, Mo, Ti, Mn, Cr, Ni, Mg, Cu, Tl, In, Ru, V and Au. Additional ligands 
can be attached to the metal. 
A variety of porphyrins have been found to have selective activity against 
HIV-1 when tested in cell culture. Both natural and synthetic porphyrins 
and metalloporphyrins were tested for inhibition of reverse transcriptase. 
Compounds tested included 
5,10-Diphenyl-15,20-di(N-methyl-3-pyridyl)-porphyrin; 
5,10Diphenyl-15,20-di(N-methyl-4-pyridyl)-porphyrin (FIG. 3A); 
5,15-Diphenyl-10,20-di(N-methyl-3-pyridyl)-porphyrin (FIG. 3B); Hemin; 
Protoporphyrin (FIG. 4); Tetra-(N-methyl-4-pyridyl)-porphyrin; 
Meso-tetraphenylporphine; Protoporphyrin IX dimethyl ester; 
Tetra-(4-carboxyphenyl)-porphyrin (FIG. 2B); 
Tetra-(4-methylphenyl)-porphyrin; Tetra-(3-methylphenyl)-porphyrin; 
Tetra-(4-hydroxyphenyl)-porphyrin (FIG. 2A); 
Fe(III)-tetraphenyl-porphyrin; Tetra-(4-chlorophenyl)-porphyrin; 
Fe(III)-tetra-(4-methylphenyl)-porphyrin; 
Fe(III)-tetra-(N-methyl-4-pyridyl)-porphyrin; and Fe(III)-mu-oxo-dimer of 
tetraphenylporphyrin. 
Protohemin was obtained from Aldrich Chemical Co., Milwaukee, Wis. Fe(III) 
tetraphenylporphyrin derivatives were either purchased from Midcentury 
Chemicals or synthesized by pyrrole-benzaldehyde condensation in a 
propionic acid reflux, by the method of A. D. Adler, F. R. Longo, J. D. 
Finarelli, J. Goldmacher, J. Assour, and L. Korsakoff, J. Org. Chem., 32, 
476 (1967). Iron was inserted using FeCl.sub.2 in dimethylformamide, as 
taught by A. D. Adler, F. R. Longo, and V. Varadi, Inorg. Syn., 16, 
213-220 (1976). 
General synthetic references are Dolphin, D. Ed., "The Porphyrins", Vol. 6, 
Chap 3-10, pp. 290-339 (Academic Press: New York, 1979); Morgan, B., 
Dolphin, D. Struct. Bonding (Berlin), 64 (Met. Complexes Tetrapyrrole 
Ligands I), pp. 115-203 (1987); Smith, Kevin M.; Cavaleiro, Jose A. S. 
Heterocycles, 26(7), 1947-63 (1987). 
Still other synthetic techniques include the recent advances by Lindsey, et 
al., J. Org. Chem. 52, 827-836 (1987); Momenteau, M.; Loock, B.; Huel, C.; 
Lhoste, J. M. J. Chem. Soc., Perkin Trans. I, 283 (1988); Morgan, B.; 
Dolphin, D. J. Org. Chem. 52, 5364-5374 (1987); Smith, K. M.; Parish, D. 
W.; Inouye, W. S. J. Org. Chem. 51, 666-671 (1986); and Smith, K. M.; 
Minnetian, O. M. J. Chem. Soc., Perkin Trans. I, 277-280 (1986). Other 
references to metal insertion include Buchler, J. E., "The Porphyrins", 
vol. 1, ch. 10, Dolphin, D., ed. (Academic Press, N.Y. 1979); Lavallee, D. 
K. Coord. Chem. Rev. 61, 55-96 (1985); Lavallee, D. K. Comments Inorg. 
Chem. 5, 155-174 (1986). 
Porphyrins may also be obtained from commercial sources including Aldrich 
Chemical Co., Milwaukee, Wis., Porphyrin Products, Logan, Utah, and 
Midcentury Chemicals, Posen, Ill. 
Another group of porphyrins which may be useful in the present invention 
are those in which the ring conjugation ha been interrupted, e.g., 
chlorins, as described by Oseroff, et al., Proc. Natl. Acad. Sci. USA, 83, 
8744-8748 (1986). 
One can screen the present compositions for inhibition of HIV by various 
experimental techniques. One technique involves the inhibition of viral 
replication in human peripheral blood mononuclear cells. The amount of 
virus produced is determined by measuring the quantity of virus-coded 
reverse transcriptase (an enzyme found in retroviruses) which is present 
in the culture medium. Another technique involves measuring inhibition of 
purified reverse transcriptase in a cell free system. 
Methodology for Testing Antiviral Drugs for Inhibition of Replication of 
HIV-1 in Human Peripheral Blood Mononuclear (PBM) Cells 
Virus Strain and Cells 
PBM cells from healthy HIV-1 and hepatitis B virus seronegative donors were 
isolated by Ficoll-Hypaque discontinuous gradient centrifugation at 
1,000.times.g for 30 minutes, washed twice in PBS and pelleted at 
300.times.g for 10 minutes. Before infection, the cells were stimulated by 
phytohemagglutinin (PHA) at a concentration of 16.7 .mu.g/ml for three 
days in RPMI 1640 medium supplemented with 15% heat-inactivated fetal calf 
serum, 1.5 mM L-glutamine, penicillin (100 U/ml), streptomycin (100 
.mu.g/ml), and 4 mM sodium bicarbonate buffer. 
HIV-1 (strain LAV-1) was obtained from the Center for Disease Control, 
Atlanta, and propagated in PHA-stimulated human PBM cells using RPMI 1640 
medium as above without PHA and supplemented with 7% interleuken-2 
(Advanced Biotechnologies, Silver Spring, Md.), 7 .mu.g/ml DEAE-dextran 
(Pharmacia, Uppsala, Sweden), and 370 U/ml anti-human leukocyte (alpha) 
interferon (ICN, Lisle, Ill.). Virus was obtained from cell-free culture 
supernatant and stored in aliquots at -70.degree. C. until used. 
Addition of Compounds 
Uninfected PHA-stimulated human PBM cells were uniformly distributed among 
25 cm.sup.2 flasks to give a 5 ml suspension containing about 
1.times.10.sup.6 cells/ml. Suitable dilutions of HIV were added to infect 
the cultures. The mean reverse transcriptase (RT) activity of the inocula 
was 50,000 dpm/ml which was equivalent to about 100 TCID.sub.50, 
determined as described in AIDS Res. Human Retro. 3,71-85 (1987). The 
drugs, at twice their final concentrations in 5 ml of RPMI 1640 medium, 
supplemented as described above, were added to the cultures. Uninfected 
and untreated PBM cells were grown in parallel as controls. The cultures 
were maintained in a humidified 5% CO.sub.2 -95% air incubator at 
37.degree. C. for five days after infection at which point all cultures 
were sampled for supernatant RT activity. Preliminary studies had 
indicated that the maximum RT levels were obtained at that time. 
The RT assay was performed by a modification of the method of Spira, et al, 
in J. Clin. Microbiol. 25,97-99 (1987) in 96-well microtiter plates. The 
radioactive cocktail (180 .mu.l) which contained 50 mM Tris-HCl pH 7.8, 9 
mM MgCl.sub.2, 5 mM dithiothreitol 4.7 .mu.g/ml (rA).sub.n.(dT).sub.12-18, 
140 .mu.M dATP and 0.22 .mu.M[.sup.3 H]dTTP (specific activity 78.0 
Ci/mmol, equivalent to 17,300 cpm/pmol; NEN Research Products, Boston, 
Mass.) was added to each well. The sample (20 .mu.l) was added to the 
reaction mixture and incubated at 37.degree. C. for two hours. The 
reaction was terminated by the addition of 100 .mu.l 10% trichloroacetic 
acid (TCA) containing 0.45 mM sodium pyrophosphate. The acid insoluble 
nucleic acid which precipitated was collected on glass filters using a 
Skatron semi-automatic harvester (setting 9). The filters were washed with 
5% TCA and 70% ethanol, dried, and placed in scintillation vials. Four ml 
of scintillation fluid (Econofluor, NEN Research Products, Boston, Mass.) 
was added and the amount of radioactivity in each sample determined using 
a Packard Tri-Carb liquid scintillation analyzer (model 2,000CA). The 
results were expressed in dpm/ml of original clarified supernatant. 
Methodology for Testing Antiviral Drugs for Toxicity and Inhibition of Cell 
Proliferation 
The drugs were evaluated for their potential toxic effects on uninfected 
PHA-stimulated human PBM cells. Flasks were seeded so that the final cell 
concentration was 3.times.10.sup.5 cells/ml. The cells were cultured with 
and without drug for 6 days at which time aliquots were counted for cell 
viability. 
Methodology for Testing Antiviral Drugs for Inhibition of HIV-1 Reverse 
Transcriptase 
Cells and viruses are cultured as described above. Reverse transcriptase is 
purified from detergent disrupted HIV-1 infected cells using DEAE and 
phosphocellulose column chromatography, according to the method of Abrell 
and Gallo, J. Virol. 12,431-439 (1973). Cocktail, template, compound to be 
tested and reverse transcriptase are mixed together on ice, then incubated 
for one hour at 37.degree. C. The DNA synthesized is acid precipitated and 
the radioactivity measured, as described by Eriksson, B., et al., 
Antimicrobial Agents and Chemotherapy 31, 600-604 (1977). 
Methodology for Testing Antiviral Drugs for Inhibition of Replication of 
HSV-1 and HSV-2 
Compounds were tested for inhibition of herpes simplex virus type 1 and 
herpes simplex virus type 2 using the method of Schinazi, et al., 
Antimicrobial Agents and Chemotherapy 22, 499-507 (1982). This is a plaque 
assay using Vero cells (African green monkey cells) infected with HSV. 
Examples of Compounds Having Anti-viral Activity and Low Toxicity 
EC.sub.50 is the median effective concentration of the compound as 
determined from inhibition of HIV replication in PBM cells. The 
therapeutic index of a compound is determined by dividing the inhibitory 
or lethal dose for 50% of the population (IC.sub.50 or LD.sub.50) by the 
effective dose for 50% of the population (EC.sub.50). 
A variety of porphyrins were tested for anti-HIV activity. Inhibition of 
replication of virus in cell culture is shown in Table I. Inhibition of 
reverse transcriptase is described in Table II. None of these compounds 
were toxic at concentrations of 1 .mu.M, 10 .mu.M and 100 .mu.M as 
determined by trypan blue exclusion by uninfected human PBM cells. As a 
result, all of the compounds active at less than 10 .mu.M have very 
favorable therapeutic indices. 
A variety of porphyrins were also tested for inhibition of herpes simplex 
type 1 and type 2 (HSV-1 and HSV-2) activity. The results are described in 
Table III. With few exceptions, no significant inhibition of HSV-1 or 
HSV-2 was noted. Similar results were obtained with Coxcksackievirus 
strain B4. 
To insure that light is not required for antiviral activity, compounds were 
tested for inhibition of HIV, HSV-1 and HSV-2 replication in cell culture 
both in the dark and in the light. No significant differences in activity 
in inhibition of HIV by 
5,10-Diphenyl-15,20-di(N-methyl-4-pyridyl)-porphyrin Cl-, nor of HSV-1 or 
HSV-2 by any of several compounds, were observed. 
The results demonstrate that the non-metalloporphyrins are generally more 
active than the metalloporphyrins. However, porphyrin derivatives 
containing iron were the least active of the metalloporphyrins. Copper and 
nickel substituted porphyrins were significantly more active. Porphyrins 
having a positive charge, negative charge, and neutral charge, under 
physiological conditions, were also tested and examples found to have HIV 
inhibitory activity. As a general rule, the positively charged porphyrins 
are more active under the assay conditions. 
TABLE I 
______________________________________ 
Code Drug Name EC.sub.50 (.mu.M) 
______________________________________ 
Inhibition of HIV Replication in PBM Cells 
Natural Porphyrins 
PPIX Protoporphyrin, disodium salt 
3.62, 0.48 
PPIXDME Protoporphyrin IX dimethyl ester 
1.01, &gt;10 
Synthetic Porphyrins 
CP-3 5,10-Diphenyl-15,20-di(N-methyl-3- 
2.28 
pyridyl)-porphyrin Cl-- 
CP-4 5,10-Diphenyl-15,20-di(N-methyl-4- 
0.38, 0.43* 
pyridyl)-porphyrin Cl-- 
TP-3 5,15-Diphenyl-10,20-di(N-methyl-4- 
0.34, 1.87 
pyridyl)-porphyrin Cl-- 
TP-4 5,15-Diphenyl-10,20-di(N-methyl-3- 
&lt;1, 0.76 
pyridyl)-porphyrin Cl-- 
TMPyP Tetra-(N-methyl-4-pyridyl)-porphyrin 
3.23, 0.66 
tosylate salt 
TPP Meso-tetraphenylporphine 
13.7, &gt;10 
TPP(4- Tetra-(4-carboxyphenyl)-porphyrin 
1.11, 10.7 
CO.sub.2 H).sub.4 
TPP(4- Tetra-(4-methylphenyl)-porphyrin 
0.754, 10 
Me).sub.4 
TPP(3- Tetra-(3-methylphenyl)-porphyrin 
0.05, 10 
Me).sub.4 
TPP(4- Tetra-(4-hydroxyphenyl)-porphyrin 
0.6, 0.7 
OH).sub.4 
TPP Tetra-(4-chlorophenyl)-porphyrin 
.sup..about. 100 
(4-Cl).sub.4 
Inhibition of HIV Production continued 
Natural Metalloporphyrins 
FePPIXCl 
Hemin, bovine, (chloro 49.3 
protoporphyrin IX Fe(III)) 
Synthetic Metalloporphyrins 
FeTPPCl Fe(III)-tetraphenylporphyrin chloride 
.sup..about. 100 
FeTPP Fe(III)-tetra-(4-methylphenyl)- 
&gt;100 
(4-Cl).sub.4 
porphyrin chloride 
FeTMPyP Fe(III)-tetra-(N-methyl-4-pyridyl)- 
78.1 
porphyrin chloride 
.mu.-oxo-TPP 
Fe(III)-mu-oxo-dimer of 
&gt;100 
tetraphenyl-porphyrin 
Cu-- CP4 
Cu(II)-5,10-diphenyl-15,20- 
4.76, 5.78 
di(N-methyl-4-pyridyl)-porphyrin 
Ni--CP4 Ni(II)-5,10-diphenyl-15,20- 
3.44, 6.54 
di(N-methyl-4-pyridyl)-porphyrin 
______________________________________ 
*Tested in darkness. 
TABLE II 
______________________________________ 
Inhibition of HIV Reverse Transcriptase 
Code Drug Name EC.sub.50 (.mu.M) 
______________________________________ 
Natural Porphyrins 
PPIX Protoporphyrin, disodium salt 
&gt;100 
PPIXDME Protoporphyrin IX dimethyl ester 
&gt;100 
Synthetic Porphyrins 
CP-3 5,10-Diphenyl-15,20-di(N-methyl-3- 
41.6 
pyridyl)-porphyrin Cl-- 
CP-4 5,10-Diphenyl-15,20-di(N-methyl-4- 
26.9 
pyridyl)-porphyrin Cl-- 
TP-3 5,15-Diphenyl-10,20-di(N-methyl-4- 
7.3 
pyridyl)-porphyrin Cl-- 
TP-4 5,15-Diphenyl-10,20-di(N-methyl-3- 
6.9 
pyridyl)-porphyrin Cl-- 
TPP Meso-tetraphenylporphine 
&gt;100 
TPP(4- Tetra-(4-carboxyphenyl)-porphyrin 
1.1 
CO.sub.2 H).sub.4 
TPP(4- Tetra-(4-methylphenyl)-porphyrin 
&gt;100 
Me).sub.4 
Natural Metalloporphyrins 
FePPIXCl 
Hemin, (chloroprotoporphyrin IX Fe(III)) 
20.5 
Synthetic Metalloporphyrins 
Cu--CP4 Cu(II)-5,10-diphenyl-15,20- 
57.4 
di(N-methyl-4-pyridyl)-porphyrin 
Ni--CP4 Ni(II)-5,10-diphenyl-15,20- 
24.8 
di(N-methyl-4-pyridyl)-porphyrin 
______________________________________ 
TABLE III 
__________________________________________________________________________ 
Inhibition of HSV Type 1 and HSV Type 2 
EC.sub.50 (.mu.M) 
HSV type 1 HSV type 2 
Code Compound Light Dark Light Dark 
__________________________________________________________________________ 
Natural Porphyrins 
PPIX Protoporphyrin, 
&lt;0.01 0.05 10 
disodium salt 
TMPyP Tetra-(N-methyl- 
0.03 0.04 &gt;1 &gt;10 
4-pyridyl)-porphyrin 
tosylate salt 
TPP Meso-tetraphenyl- 
10 &gt;10 &gt;10 &gt;10 
porphine 
PPIXDME 
Protoporphyrin IX 
10 &gt;10 &gt;10 &gt;10 
dimethyl ester 
TPP(4- 
Tetra-(4-carboxy- 
10 2.5 &gt;10 &gt;10 
CO.sub.2 H).sub.4 
phenyl)-porphyrin 
TPP(4- 
Tetra-(4-methyl- 
1 &gt;10 &gt;10 3.95 
Me).sub.4 
phenyl)-porphyrin 
TPP(3- 
Tetra-(3-methyl- 
&gt;10 &gt;10 &gt;10 
Me).sub.4 
phenyl)-porphyrin 
TPP(4- 
Tetra-(4-hydroxy- 
&gt;10 &gt;10 &gt;10 &gt;10 
OH).sub.4 
phenyl)-porphyrin 
__________________________________________________________________________ 
Humans suffering from infections caused by HIV can be treated by 
administering to the patient a pharmaceutically effective amount of the 
above described compounds having effective concentrations in vitro of less 
than or equal to 10 .mu.M in the presence of a pharmaceutically acceptable 
carrier or diluent. Some porphyrins are water soluble and may be 
administered in sterile water or physiological saline or phosphate 
buffered saline (PBS). Many porphyrins are not water soluble and are 
preferably administered in pharmaceutically acceptable non-aqueous 
carriers including oils and liposomes. Solubility of the porphyrins can be 
increased by techniques known to those skilled in the art including 
introducing hydroxyl groups and changing the counter ions. 
The compounds according to the present invention are included in the 
pharmaceutically acceptable carrier or diluent in an amount sufficient to 
exert a therapeutically useful inhibitory effect on HIV in vivo without 
exhibiting adverse toxic effects on the patient treated. By "HIV 
inhibitory amount" is meant an amount of active ingredient sufficient to 
exert an HIV inhibitory effect as measured by, for example, an assay such 
as the ones described herein. 
There may also be included as part of the composition pharmaceutically 
compatible binding agents, and/or adjuvant materials. The active materials 
can also be mixed with other active materials including antibiotics, 
antifungals, other antivirals and immunostimulants which do not impair the 
desired action and/or supplement the desired action. The active materials 
according to the present invention can be administered by any route, for 
example, orally, parenterally, intravenously, intradermally, 
subcutaneously, or topically, in liquid or solid form. 
A preferred mode of administration of the compounds of this invention is 
oral. Oral compositions will generally include an inert diluent or an 
edible carrier. They may be enclosed in gelatin capsules or compressed 
into tablets. For the purpose of oral therapeutic administration, the 
aforesaid compounds may be incorporated with excipients and used in the 
form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, 
chewing gums and the like. These preparations should produce a serum 
concentration of active ingredient of from about 0.2 to 40 .mu.M. A 
preferred concentration range is from 0.2 to 20 .mu.M and most preferably 
about 1 to 10 .mu.M. However, the concentration of active ingredient in 
the drug composition itself will depend on bioavailability of the drug and 
other factors known to those of skill in the art. 
It is to be noted that dosage values will also vary with the specific 
severity of the disease condition to be alleviated. It is to be further 
understood that for any particular subject, specific dosage regimens 
should be adjusted to the individual need and the professional judgment of 
the person administering or supervising the administration of the 
aforesaid compositions. It is to be further understood that the 
concentration ranges set forth herein are exemplary only and they do not 
limit the scope or practice of the invention. The active ingredient may be 
administered at once, or may be divided into a number of smaller doses to 
be administered at varying intervals of time. 
The tablets, pills, capsules, troches and the like may contain the 
following ingredients: a binder such as microcrystalline cellulose, gum 
tragacanth or gelatin; an excipient such as starch or lactose, a 
disintegrating agent such as alginic acid, Primogel, corn starch and the 
like; a lubricant such as magnesium stearate or Sterotes; a glidant such 
as colloidal silicon dioxide; and a sweetening agent such as sucrose or 
saccharin or flavoring agent such as peppermint, methyl salicylate, or 
orange flavoring may be added. When the dosage unit form is a capsule, it 
may contain, in addition to material of the above type, a liquid carrier 
such as a fatty oil. Other dosage unit forms may contain other various 
materials which modify the physical form of the dosage unit, for example, 
as coatings. Thus tablets or pills may be coated with sugar, shellac, or 
other enteric coating agents. A syrup may contain, in addition to the 
active compounds, sucrose as a sweetening agent and certain preservatives, 
dyes and colorings and flavors. Materials used in preparing these various 
compositions should be pharmaceutically pure and non-toxic in the amounts 
used. 
The solutions or suspensions may also include the following components: a 
sterile diluent such as water for injection, saline solution, fixed oils, 
polyethylene glycols, glycerine, propylene glycol or other synthetic 
solvents; antibacterial agents such as benzyl alcohol or methylparabens; 
antioxidants such as ascorbic acid or sodium bisulfite; chelating agents 
such as ethylenediaminetetraacetic acid; buffers such as acetates, 
citrates or phosphates and agents for the adjustment of tonicity such as 
sodium chloride or dextrose. The parental preparation can be enclosed in 
ampoules, disposable syringes or multiple dose vials made of glass or 
plastic. 
The compositions of the present invention are prepared as formulations with 
pharmaceutically acceptable carriers. Preferred are those carriers that 
will protect the active compound against rapid elimination from the body, 
such as a controlled release formulation, including implants and 
microencapsulated delivery systems. Biodegradable, biocompatable polymers 
can be used, such as polyanhydrides, polyglycolic acid, collagen, and 
polylactic acid. Methods for preparation of such formulations will be 
apparent to those skilled in the art. 
Liposomal suspensions (including liposomes targeted to infected cells with 
monoclonal antibodies to viral antigens) are also preferred as 
pharmaceutically acceptable carriers. Methods for encapsulation or 
incorporation of porphyrins into liposomes are described by Cozzani, I.; 
Jori, G.; Bertoloni, G.; Milanesi, C.; Sicuro, T. Chem. Biol. Interact. 
53, 131-143 (1985) and by Jori, G.; Tomio, L.; Reddi, E.; Rossi, E. Br. J. 
Cancer 48, 307-309 (1983). These may also be prepared according to methods 
known to those skilled in the art, for example, as described in U.S. Pat. 
No. 4,522,811 (which is incorporated herein by reference in its entirety). 
For example, liposome formulations may be prepared by dissolving 
appropriate lipid(s) (such as stearoyl phosphatidyl ethanolamine, stearoyl 
phosphatidyl choline, arachadoyl phosphatidyl choline, and cholesterol) in 
an inorganic solvent that is then evaporated, leaving behind a thin film 
of dried lipid on the surface of the container. An aqueous solution of the 
active compound is then introduced into the container. The container is 
then swirled by hand to free lipid material from the sides of the 
container and to disperse lipid aggregates, thereby forming the liposomal 
suspension. 
Other methods for encapsulating porphyrins within liposomes and targeting 
areas of the body are described by Sicuro, T.; Scarcelli, V.; Vigna, M. 
F.; Cozzani, I. Med. Biol. Environ. 15(1), 67-70 (1987) and Jori, G.; 
Reddi, E.; Cozzani, I.; Tomio, L. Br. J. Cancer, 53(5), 615-21 (1986). 
Modifications and variations of the present invention, compositions 
containing porphyrins, and methods for using the compositions, for 
treating patients with HIV infections, will be obvious to those skilled in 
the art from the foregoing detailed description of the invention. Such 
modifications and variations are intended to come within the scope of the 
appended claims.