FIELD OF THE INVENTION The present invention relates to new bisguinolines useful in the treatment of malaria and to processes for the production thereof. The invention also relates to methods for the treatmen of malaria and, in particular, to the treatment of chloroquine resistant strains of malaria. In a preferred embodiment, this invention relates to N,N-bis(7-chloroguinolin-4-yl)alkane diamines which are active against chloroquine-resistant malaria.

BACKGROUND OF THE INVENTION

In the following discussion, a number of citations from professional journals are included for the convenience of the reader. These citations are in abbreviated form in the text by author and year only. The full citation of each is set forth in the References section at the end of the specification.

While these citations more fully describe the state of the art t which the present invention pertains, the inclusion of these citations is not intended to be an admission that any of the cited publications represent prior art with respect to the present invention.

By a large margin, malaria is the most prevalent disease i the world. It is estimated for the year 1986 that some 489 million people contracted malaria, 2.3 million of whom died from the disease (Sturchler, 1989). Whereas effective antimalarial drugs exist, drug-resistance, particularly resistance to

chloroquine (CQ), the most useful antiraalarial drug, has become an enormous problem (Payne, 1987).

Quinoline anti alarials such as quinine, mefloquine, and amodiaquine are active to various extents against CQ-resistant malaria (Geary and Jensen, 1983; Geary et al., 1987; Knowles et al., 1984; Watkins et al., 1984; Cowman and Foote, 1990; Sowunm et al., 1990). Although Cowman and Foote (1990) suggest that C resistance may dispose the parasite to resistance to other quinolines, LeBras et al. (1983), Schmidt et al. (1977), Geary and Jensen (1983) and Oduola et al. (1988) observe a significan lack of cross-resistance among quinoline-containing antimalarial .

It is an object of this invention to provide a new class quinolines which are active against malaria and, in particular, CQ resistant malaria. The compounds of this invention are bis- quinolines.

Many prior art bisquinolines have been reported to be inactive against malaria. These include the succinic acid diester of amodiaquine as a potential repository form (Elslager et al, 1969), and bisquinolines lacking either a 4-amino functi or with a bridge at the 3 rather than the 4-position (Nasr et a 1979; Nasr et al, 1978).

Examples of bisquinolines which have been reported to be active against malaria are shown in Figure 1 and include severa bisquinolylpiperazines such as piperaquine, hydroxypiperaquine, dichloroquinazine, 12494RP (Benazet, 1965; 1967; Lafaix et al., 1967; LeBras et al. , 1983; Li et al. , 1981a; 1981b; 1984; Zhan et al., 1987; Li and Huang, 1988; Chen et al, 1982), and l,4-bis(7-chloro-4-quinolyl-amino)piperazine (Singh et al. , 1971). In general, these bisquinolines are more potent than CQ and are active against CQ-resistant malaria. Both piperaquine (PQ) and hydroxypiperaquine are claimed to be very effective against CQ-resistant malaria in China (Chen et al. , 1982; Li et al., 1981b; 1984; Li and Huang, 1988). Each of these drugs als has a longer duration of action, and less toxicity when compare to CQ (Li et al., 1981a; Lin et al., 1982). Dichloroquinazine active aqainst CQ-resistant falciparum malaria (LeBras et al., 1983), and a mixture of 12,494RP and dichloroquinazine is clinically effective against falciparum malaria and exerts a suppressive effect lasting for 3 weeks (Lafaix et al. 1967; Benazet, 1965). Resistance to dichloroquinazine, however, is noted for a CQ-resistant strain of P. jergr ei (Warhurst, 1966). Although ι,4-bis(7-chloro-4-quinolylamino)piperazine has not be screened against CQ-resistant malaria, it is significantly more effective than is CQ against P. Jbergrhei in mice (Singh et al., 1971).

This invention relates to a new class of bis-4- aminoquinoline antimalarial agents. These agents exhibit potent activity against CQ-resistant malaria in the in vitro and -in. viv tests hereinafter described.

SUMMARY OF THE INVENTION In accordance with the present invention novel N,N-bis(7- substituted-quinolin-4-yl)alkane diaraines are provided. In one embodiment, these compounds can be depicted by the following general formula:

FORMULA I

wherein R is a bivalent radical derived from an acyclic or cycli hydrocarbon by removal of one hydrogen atom from each of two different carbon atoms. R' is hydrogen or lower alkyl (generall containing between about l and about 4 carbon atoms). X is hydrogen (-H), fluoro (-F), chloro (-C1), bro o (-Br), trifluoromethyl (-CF _S ), cyano (-CN) , or methylsulfoxide (-SOCH ₃ ). In its acyclic form, R generally contains at least two, and no more than about 12, carbon atoms and, preferably, is an unsubstituted straight or branched alkane. In its cyclic

form, R contains at least three and, generally, no more than about eight carbon atoms and, preferably, is an unsubstituted cycloalkane.

It is believed that no compound within the scope of the above formula, i.e., N,N-bis(7-substituted-quinolin-4-yl)alkane diamines, has been described in the prior art, aside from N N ² - bis(7-chloroquinolin-4-yl)ethane-l,2-diamine (Pearson et al. , 1946). This compound has the following structural formula:

FORMULA II

The Pearson et al reference does not disclose this bisquinoline to have antimalarial properties. As is shown hereinafter, however, treatment of malaria parasites with this prior art compound, in accordance with the antimalarial methods of this invention, has resulted in antimalarial activity. As will be described more fully hereinafter, however, the next higher homologs of this compound, as well as other analogs thereof, exhibit unexpectedly higher antimalarial activity than the Pearson et al compound. Therefore, referring to Formula I above, an especially preferred embodiment of this invention is where R contains at least 3 carbon atoms.

Compounds of the following formula, wherein R is cyclic, constitute a preferred embodiment of this invention:

FORMULA III

wherein X is as defined above and n is a whole integer from 1 through about 6, 4 being especially preferred because of the exceptional activity exhibited by ±-trans-N ¹ ,N ² -Bis(7-chloro- quinolin-4-yl)cyclohexane-l,2-diaroine (Table I, compound 3).

The following formula, wherein R is acyclic, constitutes another embodiment of this invention:

FORMULA IV

wherein R is selected from the group consisting of CH ₂ CH(CH ₃ ), (CH ₂ ) ₃ , (CH ₂ ) ₄ , (CH ₂ ) ₈ , (CH _a ) ₃ CH(CH ₃ )CH ₂ , (CH ₂ ) _β , (CH ₂ ) ₇ , (CH ₂ )., (CH ₂ )„, CH ₂ ) ₁₀ , (CH.) _U/ and (CH ₂ ) ₁₂ , and other preferably lower- alkyl substituted derivatives thereof.

A method for the treatment of malaria constitutes another embodiment of this invention. This method comprises administering to a host an N,N-bis(7-substituted-quinolin-4- yl)alkane diamine of this invention in a pharmaceutically acceptable dosage form containing an amount of said diamine whic is effective in treating malaria. Ideally, the effective dose for treating malaria is that dose which is toxic to the malaria parasite infecting the host, but below the threshold of significant toxicity to the host. Generally, for the compounds of this invention this dose ranges from about 5 mg to about 100 g per kilogram of host body weight. Because the compounds of this invention exhibit high therapeutic indices, it is possible, but generally not economically practical, to employ higher doses up to even 500 mg/kg and higher. Normally, because the compounds of this invention have such high antimalarial activity, doses of between about 5 and about 50 mg/kg host body weight are employed.

DETAILED DESCRIPTION OF THE INVENTION Che-oistry

The bisquinolines of this invention can be produced via a displacement reaction with 4,7-dichloroquinoline, alkanediamine, and triethylamine in a 2 : 1 : 2 ratio using N-methylpyrrolidinone as solvent. For the production of compounds 1-13 (see Table I) no success resulted with the method of Singh et al. (1971) where K ₂ C0, was used as base with

ethoxyethanol as solvent. Substitution of triethylamine for K.COj, however, gave good results. Furthermore, substitution of N-methylpyrrolidinone as reported by Tyman et al. (1989) was found to be a better solvent than ethoxyethanol for this reaction. For example, yields for compounds 4 and 7 (Table I) more than doubled when N-methylpyrrolidinone, rather than ethoxyethanol was used. Yields for reactions in ethoxyethanol and N-methylpyrrolidinone ranged from 23-85% and 49-87% respectively. Compounds 1-13 (Table I) were isolated by adding wat,er and ethyl ether or ethyl acetate to the cooled reaction mixtures which initiated product precipitation and dissolved an unreacted starting materials.

Pharmacology Twelve of the thirteen bisquinolines set forth in Table I had a significantly lower resistance index than did CQ, and compared favorably with PQ in this regard. The resistance ind was apparently unrelated to in vitro or in vivo activity. Eigh bisquinolines were more potent than was either CQ and PQ agains both clones of P. falciparum. Except for compounds 8 and 12, there was a reasonable correlation between in vitro and in vivo antimalarial activities. For example, compounds 2, 3, 6, 7, an 9-11 which had IC _β0 'ε less than 6 nM against P. falciparujπ were either active or curative against P. berghei in vivo. Conversel compounds 1, 4, 5, and 13 which were approximately an order of magnitude less potent in vitro, were also without activity in

vivo. Compound 3, the most potent bisquinoline in vitro, was clearly unique in its in vivo activity; 4/5 and 5/5 mice were cured at 160 and 320 mg/kg, respectively. No other compound was curative at the 160 mg/kg dose.

Methyl substitution in the bridge improved activity, eg. 2 vs. 1 and 7 vs. 6. In the three derivatives (1-3) with a two-carbon bridge, decreased confor ational mobility seemed to increase activity. Compounds 4, 5, and 13 with bridges of three, four or twelve carbon atoms were inactive in both screens.

However, compounds 6-11 with bridges of between 5 and 9 carbon atoms were active. Molecular modeling using MMX suggests that 4 and 5, unlike 6-11, are not able to achieve a conformation similar to that observed for 3 which suggests that the relative orientation of the two quinoline heterocycles is important for activity.

In summary, the data set forth in Table I is consistent with the excellent results observed in China with PQ against CQ-resistant falciparum malaria. PQ had a resistance index of 1.9 compared to that of 11.2 for CQ. From this data, it is also observed that, like PQ, bisquinolines 1-13 have much lower resistance indices than does CQ against CQ-resistant P. falciparum. in vitro. Furthermore, six of the thirteen bisquinolines show superior antimalarial activity (both in vitro and in vivo) to CQ. Thus, it is believed that these results

support the premise that the bisquinolines of this invention are useful agents against malaria and CQ-resistant malaria, in particular.

EXPERIMENTAL Melting points were taken with a Mel-Temp capillary apparatus. IR spectra were run as KBr discs on a Perkin Elmer 1420 spectrophotometer. NMR spectra were obtained with either Varian XL-300 or Bruker AC-200 spectrometers using deuteriated dimethyl sulfoxide with TMS as an internal standard. It was not possible to obtain "C NMR spectra for 1,4, and 5 due to their lo solubilities in DMSO. Microanalyses were performed by M-H-W Laboratories, Phoenix, AZ. The purity of 1-13 was confirmed wit silica gel or alumina TLC. 4,7-Dichloroquinoline and the required dia ines are commercially available from Aldrich Chemical Co. , with the exception of 2-methylpentamethylenediamin and 1,12-dodecanediamine which are available from the Du Pont Company, Petrochemicals Department, Wilmington, Delaware. All reactions were conducted under a positive pressure of N ₂ subsequent to ten purge-cycles using a Firestone valve.

Chemistry. Synthesis of Table I Compounds 1-13. A solution of 4,7-dichloroquinoline (10 mmol, 1.98g), triethyla ine (10 mmol, l.Olg) and diamine (5 mmol) in either ethoxyethanol or

N-methylpyrrolidinone (10 mL) was heated to reflux for 6 to 24 hours under a slight positive N. pressure. After the reaction mixture cooled to room temperature, ether or ethyl acetate (15 mL) and water (15 mL) were added with stirring and the resulting solid was filtered and washed with water and ethyl acetate or

ether to provide 1-13. When required, crystallization of 1-13 was best accomplished from aqueous EtOH.

N ³ -Bis(7-chloroquinolin-4-yl)ethane-l ,2-diamine (1) : (1.63g , 85%); mp 342-345°C dec. (lit. mp 334.5-337°C (Pearson, et al., 1946)); IR 3460, 3230, 3065, 3020, 2970, 2890, 1610, 1580, 1535 cm-1; Η NMR δ 3.62 (m, 4H) , 6.58 (d, J = 5.4 Hz, 2H) , 7.47 (dd J = 9.0 HZ, J = 2.4 HZ, 2H), 7.48 (t, J = 4.2 Hz, 2H) , 7.79 (d, = 2.4 Hz, 2H), 8.23 (d, J = 9.0 Hz, 2H) , 8.41 (d, J = 5.4 Hz, 2H). Anal. (C _ao H _lβ Cl.N ₄ • 0.5 H ₂ 0) C, H, N. N ¹ ,N ^a -Bis(7-chloroquinolin-4-yl)propane-l,2-diamine (2): (1.43g 72%); mp 287-289°C dec; IR 3440, 3070, 2980, 2930, 1610, 1575, 1535 cm-l; ^X H NMR δ 1.35 (d, J = 6.3 Hz, 3H) , 3.49-3.59 (m, 2H) 4.11-4.20 (m, 1H), 6.55 (d, J = 5.7 Hz, 1H) , 6.61 (d, J = 5.7 Hz, 1H), 7.05 (d, J = 8.1 Hz, 1H) , 7.44 (dd, J = 9.0 Hz, J = 2. Hz, 1H), 7.46 (t, J = 2.4 Hz, 1H) , 7.47 (dd, J = 9.0 Hz, J - 2. HZ, 1H), 7.778 (d, J = 1.8 Hz, 1H , 7.784 (d, J = 1.8 Hz, 1H) , 8.22 (d, J = 9.0 Hz, 1H), 8.34 (d, J = 9.0 Hz, 1H) , 8.35 (d, J 5.7 HZ, 1H) , 8.40 (d, J = 5.7 Hz, 1H) ; "C NMR δ 17.88, 46.86', 46.99, 98.76, 98.93, 117.39, 117.47, 123.86, 124.03, 124.21, 127.43, 127.47, 133.34, 149.05, 149.17, 149.43, 150.10, 151.79, 151.83. Anal. (C _M H _lt Cl,N ₄ ) C, H, N.

±-trans-N ¹ , N*-Bis ( 7-chloroquinolin-4-yl ) cyclohexane-1 , 2-diamine (3): (1.55g, 71%); mp 322-324 ^° C dec; IR 3435, 3250, 3060, 2935, 2860, 1610, 1570, 1535 cm-1; Η NMR δ 1.34-1.70 (m, 4H) , 1.72-1.91 (m, 2H), 2.02-2.21 (m, 2H) , 3.78-3.97 (m, 2H) , 6.74 (d, J = 5.6 HZ, 2H), 6.94-6.98 (m, 2H) , 7.31 (dd, J = 8.9 Hz, J = 2.0 HZ, 2H), 7.63 (d, J = 2.0 Hz, 2H) , 8.11 (d, J - 9.1 Hz, 2H) , 8.28 (d, J = 5.5 Hz, 2H) ; "C NMR δ 24.63, 31.55, 55.50, 99.02, 117.30, 123.54, 124.02, 127.25, 133.09, 149.07, 149.78, 151.55. Anal. C^H _M CI-JU C, H, N. N ^l ,N ³ -Bis(7-chloroquinolin-4-yl)propane-l,3-diauaine (4): (0.73g, 37%); mp 312-314 ^* C dec; IR 3450, 3240, 3070, 2960, 2880, 1610, 1580, 1535 cm-1; Η NMR δ 2.07 (m, 2H) , 3.43 (m, 4H) , 6.51 (d, J = 5.4 HZ, 2H), 7.40 (t, J = 5.3 Hz, 2H) , 7.45 (dd, J = 9.0 Hz , J = 2.4 HZ, 2H), 7.78 (d, J « 2.4 Hz, 2H) , 8.29 (d, J = 9.0 Hz, 2H), 8.37 (d, J = 5.4 Hz, 2H) . Anal. (C-,H _lf Cl,N ₄ ) C, H, N.

N ¹ , N ⁴ -Bis ( 7-chloroquiπolin-4-yl ) butane-1 , -diamine ( 5 ) : ( 1. llg , 54%),* mp 339-341 ^* C dec; IR 3215, 3065, 2960, 1610, 1580, 1550 cm-1; Η NMR δ . Anal. ( C _a -H ₁₀ Cl _a N ₄ ) C, H, N.

N ^λ , N'-Bis ( 7-chloroquinolin-4-yl ) pentane-1 , 5-diamine ( 6 ) : ( 1.07g , 50%); mp 272-274'C; IR 3450, 3250, 3070, 2950, 2880, 1610, 1585, 1535 cm-l; Η NMR δ 1.49-1.56 (m, 2H) , 1.69-1.78 (m, 4H) , 3.25-3.32 (m, 4H), 6.46 (d, J = 5.4 Hz, 2H) , 7.32 (t, J = 5.4 Hz, 2H), 7.44 (dd, J = 9.0 Hz, J = 2.4 Hz, 2H) , 7.78 (d, J = 2.4 Hz, 2H), 8.28 (d, J = 9.0 Hz, 2H) , 8.38 (d, J = 5.4 Hz, 2H) ; "C NMR δ 24.25, 27.55, 42.33, 98.56, 117.42, 123.88, 124.04, 127.45, 133.28, 149.08, 150.03, 151.85. Anal. (C„H--C1.N ₄ ) C, H, N.

N ¹ ,N ^s -Bis(7-chloroquinolin-4-yl)-2-Biethylpentane-l,5-diami ne (7)

(0.50g, 23%); mp 228-230°C; IR 3450, 3065, 2960, 1610, 1580, 15 cm-1; *H NMR δ 0.97 (d, J = 6.6 HZ, 3H) , 1.23-1.36 (m, 1H) , 1.55-2.02 (m, 4H) , 3.03-3.51 (m, 4H) , 6.44 (d, J = 5.4 Hz, 1H) , 6.45 (d, J = 5.4 Hz, 1H) , 7.29 (t, J = 5.1 Hz, 1H) , 7.36 (t, J 5.7 HZ, 1H), 7.42 (dd, J = 9.0 Hz, J = 2.1 Hz, 2H) , 7.77 (d, J 2.4 HZ, 2H) , 8.25 (d, J = 9.0 Hz, 1H) , 8.29 (d, J = 9.0 Hz, 1H) 8.357 (d, J = 5.4 Hz, 1H) , 8.364 (d, J = 5.4 Hz, 1H); "C NMR δ 17.71, 25.18, 31.26, 31.57, 42.64, 48.66, 98.50, 98.58, 117.39, 123 _. .82, 123.90, 123.97, 127.41, 133,26, 149.04, 149.08, 149.99, 150.14, 151.75, 151.79. Anal. (C-.H^Cl-U C, H, N. N ¹ N*-Bis(7-chloroguinolin-4-yl)hexane-l,6-diaB-ine (8): (l.55g, 71%); mp 284-286 ^* C dec; IR 3450, 3300, 3105, 3065, 3010, 2930, 2830, 1610, 1570, 1535 cm-1; ^l H NMR δ 1.42-1.53 (m, 4H), 1.63-1.74 (m, 4H), 3.23-3.29 (m, 4H) , 6.45 (d, J = 5.4 Hz, 2H) , 7.31 (t, J = 5.1 Hz, 2H), 7.43 (dd, J = 9.0 Hz, J = 2.1 Hz, 2H) 7.77 (d, J = 2.1 Hz, 2H), 8.27 (d, J = 9.0 Hz, 2H) , 8.37 (d, J 5.4 Hz, 2H) ; "C NMR δ 26.37, 27.71, 42.29, 98.52, 117.39, - 123.86, 124.02, 127.41, 133.25, 149.06, 150.01, 151.83. Anal. (C _M H^CI-NJ C, H, N.

N ¹ ,IT-Bis(7-chloroquinolin-4-yl)heptane-1,7-diamine (9) : (1.96g 87%); mp 218-220"C; IR 3450, 3060, 2935, 2860, 1610, 1580, 1535 cm-1; *H NMR δ 1.39 (br s, 6H) ,1.53-1.77 (m, 4H) , 3.16-3.32 (m, 4H), 6.44 (d, J = 5.5 Hz, 2H) , 7.31 (t, J = 5.1 Hz, 2H) , 7.44 (dd, J - 9.0 HZ, J = 2.2 Hz, 2H) , 7.79 (d, J = 2.2 Hz, 2H) , 8.2 (d, J = 9.1 HZ, 2H) , 8.39 (d, J = 5.4 Hz, 2H) ; "C NMR δ 26.64,

27.73, 28.64, 42.35, 98.54, 117.43, 123.91, 124.08, 127.46, 133.30, 149.09, 150.04, 151.87. Anal. (C _2S H ₂₆ C1-N ₄ ) C, H, N. N ¹ , N*-Bis ( 7-chloroquinolin-4-yl ) octane-1 , 8-diamine ( 10 ) : ( l .8 Og , 77%); mp 216-219"C; IR 3450, 3350, 3070, 2940, 2865, 1610, 1580, 1540 cm-1; Η NMR δ 1.35 (br s, 8H) ,1.57-1.75 (m, 4H) , 3.15-3.31 (m, 4H), 6.44 (d, J = 5.5 Hz , 2H) , 7.28 (t, J = 5.1 Hz , 2H) ,.7.43 (dd, J = 9.0 HZ, J = 2.3 HZ, 2H), 7.77 (d, J = 2.2 Hz, 2H) , 8.27 (d, J = 9.1 Hz, 2H) , 8.38 (d, J = 5.4 Hz, 2H) ; "C NMR δ 26.58, 27.73, 28.79, 42.36, 98.55, 117.42, 123.92, 124.08, 127.44, 133.30, 149.08, 150.04, 151.87. Anal. ( C _2<s H _2β Cl ₂ N ₄ ) C, H, N.

N ¹ ,N ^β -Bis(7-chloroquinolin-4-yl)nonane-l _f 9-diamine (11): (1.82g, 76%); mp 161-164 ^* C; IR 3455, 3370, 3065, 2930, 2860, 1610, 1575, 1540, 1535 cm-1; Η NMR δ 1.14-1.50 (br s, 10H) ,1.54-1.73 (m, 4H), 3.13-3.32 (m, 4H) , 6.44 (d, J = 5.5 Hz, 2H) , 7.28 (t, J = 5.1 HZ, 2H), 7.43 (dd, J = 9.0 Hz, J = 2.3 Hz, 2H) , 7.77 (d, J = 2.2 HZ, 2H), 8.28 (d, J = 9.0 HZ, 2H) , 8.38 (d, J = 5.4 Hz, 2H) ; "C NMR δ 26.61, 27.74, 28.78, 28.98, 42.37, 98.53, 117.43, 123.90, 124.08, 127.46, 133.30, 149.09, 150.04, 151.87. Ana-1. (C _M H ₃₀ C1 ₂ N ₄ ) C, H, N. N ^r ,N ¹⁰ -Bis(7-chloroquinolin-4-yl)decane-l,10-diaHine (12):

(2.15g, 87%); mp 200-204 ^β C; IR 3445, 3285, 3060, 2930, 2855, 1610, 1580, 1535 cm-1; Η NMR δ 1.28 (br s, 12H) ,1.52-1.74 (m, 4H), 3.15-3.31 (m, 4H) , 6.44 (d, J = 5.4 Hz, 2H) , 7.28 (t, J «* 5.1 Hz, 2H), 7.43 (dd, J = 8.9 Hz, J = 2.3 Hz, 2H) , 7.70 (d, J = 2.2 H2, 2H), 8.27 (d, J = 7.0 Hz, 2H) , 8.38 (d, J = 5.4 Hz, 2H) ; "C NMR δ 26.70, 27.72, 28.80, 28.94, 42.37, 98.53, 117.44,

123.89, 124.08, 127.46, 133.29, 149.10, 150.04, 151.86. Anal. (C ₂ ,H ₃₂ C1 ₂ N ₄ ) C, H, N.

N ¹ ,N"-Bis(7-chloroquinolin-4-yl)dodecane-l,12-diaaine (13) : (1.70g, 65%); mp 188-190°C; IR 3460, 3070, 2930, 2860, 1610, 1580, 1540 cm-1; ^X H NMR δ 1.24-1.35 (m, 16H) ,1.59-1.69 (m, 4H) ,

3.22-3.28 (m, 4H), 6.46 (d, J = 5.7 Hz , 2H) , 7.57 (t, J = 5.3 Hz 2H), 7.44 (dd, J = 9.0 Hz, J = 2.1 Hz, 2H) , 1.IS, (d, J = 2.1 Hz, 2H), 8.29 (d, J = 9.0 Hz, 2H) , 8.39 (d, J = 5.7 Hz , 2H) ; "C NM rS 18.49, 25.42, 26.55, 27.69, 28.74, 28.91, 42.34, 55.97 98.52, 117.41, 123.86, 124.05, 127.41, 133.26, 149.07, 150.03, 151.83. Anal. (C ₃₀ H, _t ClJI _< ♦ H ₂ 0) C, H, N. Analytical Data

<1> Anal. Calcd. for . 0.5 H.O l C, 61.33; E, 4.37) It, 14.2S. pound: C, 61.36, B, 4.46; H, 14.26. <2) Anal. Calcd. Cor C^E^Cl-M,: C, 63.4a; B, 4.B7, K, 14.10. roundr C, 63.49; B, 4.71; B, 14. S. O) Anal. Calcd. for C ₂₄ H _a2 Cl ₂ M ₄ : C, 65.91; B, 5.07; , 12.*1. round: C, 65.71, B, S.16; U, 12.69.

<4) Calcd. for C- _j B^Cl.: M. i C, 63.48; B, 4.67; If, 14.10.

C, 63.25; B, 4.65; H, 14.15.

C, 64.24; B, 4.90; H, 13.62.

C, 64.07; B, 4.S3; B, 13.56.

C, 64.94, H, 5.21; If, 13.17.

C, 65.15; B, 5.21; H, 13.31.

C, 65.61; B, 5.51; II, 12.75.

C, 65.43; B, 5.5S; R, 12.59.

C, 69.61; M, 5.81; M, 12.75.

C, 65.34; B, 5.69; B, 12.71.

C, 66.22; B, S.7a; t, 12.36.

C, 66.39; B, B.«2; St, 12.33.

C, 66.*l; B, 6.04; ST, 11.99.

C, 66.96; B, 6.2S; K, 11.99.

C, 67.35; B, 6.2*/ St, 11.64.

C, 67.43; B, 6.36, M, 11.75.

C, 67.*7; B, 6.SI; M, 11.31.

C, 6«.02; B, 6.63; M, 11.35.

C, 66.B3; B, 7.07; N, 10.35. C, 66.SO; B, 7.32; N, 9.93.

Pharmacological Methods

In vitro activity against P. falciparum was determined using a modification of the semiautomated microdilution techniqu of Desjardins et al. (1979) and Milhous et al. (1985). Two P. falciparum malaria parasite clones, designated as Sierra Leone (D-6) and Indochina (W-2), are used in susceptibility testing. The former is resistant to mefloquine, and the latter to CQ, pyrimethamine, sulfadoxine, and quinine. Test compounds are dissolved in dimethylsulfoxide, and solutions serially diluted with culture media. Erythrocytes with 0.25 to 0.5% parasitemia are added to each well of a 96-well microdilution plate to give final hematocrit of 1.5%. Inhibition of uptake of tritiated hypoxanthine is used as an index of antimalarial activity. Results are reported as IC ₉₀ (ng/mL) values. For a complete description of this assay, see Milhous et al. (1985) and Lin et al. (1987).

In vivo activity against P. Jbergrhei was obtained against a drug-sensitive strain of P. berghei (strain KBG 173) (Osdenβ et al., 1967). Each test compound is administered subcutaneous to five male mice per dilution in a single subcutaneous dose 3 days after infection. Results are expressed in T - C values which indicate the mean survival time of the treated mice beyond that of the control animals; untreated mice survive on average 6.2 days. Compounds are classified as active (A) when the mean survival time of the treated mice is twice that of the controls (>6.2 days), and curative (C) when one or more test animals live

60 days post-infection. Deaths from 0-2 days post-treatment ar attributed to toxicity (T) .

The compounds set forth in the following Table I have the formula:

wherein R is as defined in the Table.

TABLE I

Antimalarial Activity of 1-13 against P. falciparum in vitro and P. berghei in vivo

P. barohal, Ϊ-C Idayal"

40 160 640β<J Tcg

O .O 0.4 1.4

3.4 5 .7 C-3

KA ^e

0.2 0.4 1.2

0.5 2.3 5.6 l.O s.ii C-ld

C-l

2.6 7.2 C-3

-0.2 l. O C-l

3.1 7.4 ^d 15. Q *

0.2 0.2 O.C

O.O -0.2 O.O • -7 C-l C-l,T-3*

PQ *IC _βo (W-2)/IC _βo (D-6) ratio. ^-C is the mean survival time of the treated mice beyond that of the control animals (single does administered s. 3 days post infection, n=5). This value must be ≥ twice the mean survival time (6.2 days) of the control animals to be considered active (A). Survival beyond 60 days is considered curative (C), and deaths from 0-2 days post-treatment are attributed to toxicity (T). ^c C-5 at 320 mg/kg. ^d Skin lesions observed at site of injection. *T-C values for CQ . represent averages of ten best data sets from WRAIR.

Therapeutic doses and formulations

The compounds of this invention can be administered to th host or patient as an active ingredient in a variety of dosage forms. In addition to the active ingredient, which may be in t form of a pharmaceutically-acceptable derivative, such as a pharmaceutically-acceptable salt, any of a number of pharmaceutically-acceptable excipients which facilitate processing of the active compound into suitable pharmaceutical preparations can be used to formulate these compositions. Thes are well known and need not be detailed here (e.g., see Remington ' s Pharmaceutical Sciences, 1985). Because the bisquinolines of this invention are active orally, dosage forms designed for oral administration are preferred. Exemplary are tablets, capsules, and dragees. In some cases, for example, where the host is seriously ill and time is of the essence, it may be necessary to administer the compounds of this invention parenterally. In such cases intravenous administration is usually preferred. However, other dosage forms designed for parenteral administration can also be employed, e.g., subcutaneous or rectal (usually suppositories).

Appropriate formulations for parenteral administration include aqueous solutions of the active compound prepared in a water-soluble or water-dispersible form. Alternatively, the active compounds may be administered as suspensions in appropriate oily injection carriers, i.e., in suitable lipophil

carriers, such as fatty oils (sesame oil being an example) , or synthetic fatty acid esters (ethyl oleate or triglycerides being examples). Pharmaceutical formulations prepared for aqueous injection may contain substances which increase the viscosity or the suspension such as, for example, sodium carboxymethyl cellulose, sorbitol, and/or dextran.

The therapeutic bisquinolines of the present invention may also be administered encapsulated in liposomes. In such pharmaceutical preparations, the active compound is contained in corpuscles which consist of concentric aqueous layers interspersed between hydrophobic lipidic layers. The bisquinolines, depending upon their solubility, may be present both in the aqueous layer and in the lipidic layer, or in what is generally termed a liposomic suspension. The hydrophobic layer, generally but not exclusively, comprises phospholipids such as lecithin and sphingomyelin, steroids such as cholesterol, more or less ionic surfactants such as a diacetylphosphate, stearylamine, or phosphatidic acid, and/or other materials of a hydrophobic nature which are generally well known in the art.

To be available for use in systemic administration, the therapeutic bisquinolines must be formulated into suitable pharmaceutical compositions; the protocol for systemic administration would use a therapeutic approach compatible with the particular formulation selected. Pharmaceutical compositions

within the scope of the present invention include those compositions where the bisquinoline is contained in an effectiv amount sufficient to kill the malaria-inducing parasite without causing unacceptable toxicity for the host or patient. The therapeutic amount which represents an effective anti-malaria dose sufficient for treatment of each of the various types of malaria remains to be determined empirically by those skilled i the art of designing and administering anti-malarialε. However it has been determined that the bisquinolines of this invention appear to have high therapeutic indices, thus presenting a wide range of effective dosage options and strategies. A preferred dosage range is from about 5 to about 100 milligrams of bisquinoline per milligram of host body weight, given three tim a day. Doses as high as 500 mg/kg, or even higher, thrice dail can be given, but are not economically practical in the usual case of malaria encountered. As a practical matter, any dose which is sufficient to achieve an effective blood concentration of from about 0.05 to about 0.2 μg/mL can be employed.

References

Benazet, F. Plasmodium berqhei et antimalariques a action de longue duree. Ann. Soc. Beige. Med. Trop. 1965, 45, 459-466.

Benazet, F. Activite D'un Nouvel Antimalarique , Le 16.126 R. P. Sur Le Paludisme Experimental Des Animaux De Laboratoire. Bull . Soc. Pathol. Exot. 1967, 60 , 221-228.

Chen. L., Qu, F.-Y.; Zhou, Y.-C. Field Observations on the Antimalarial Piperaquine. Chin. Med. J. 1982, 95, 281-286.

Cowman, A. F.; Foote, S. J. Chemotherapy and Drug Resistance in Malaria. Jnt. J. Parasitol. 1990, 20, 503-513.

Desjardins, R. E. , Canfield, C. J. , Haynes, J. D. , and Chulay, J D. Quantitative assessment of antimalarial activity in vitro by semiautomated microdilution technique. AntimicroJb. Agents Chemother. 1979, 26, 710-718.

Geary, T. G.; Divo, A. A.; Jensen, J. B. Activity of quinoline-containing antimalarials against chloroquine-sensitive and -resistant strains of Plasmodium falciparum in vitro. Trans. R. Soc. Trop. tied. Hyg. 1987, 82, 499-503.

Geary, T. G.; Jensen, J. B. Lack of Cross-Resistance to 4-Aminoquinolines in Chloroquine-Resistant Plasmodium Falciparum In Vitro. J. Parasltol . 1983, 69 , 97-105.

Knowles, G.; Davidson, W. L.; Jolley, D.; Alpes, M. P. The relationship between the in vitro response of Plasmodium falciparum to chloroquine, quinine and mefloquine. Trans. Roy. Soc. Trop. Med. Hyg. 1984, 78 , 146-150.

LeBras, J. , Deioron, P., and Charmot, G. Dichloroquinazine (A 4-Aminoquinoline) Effective In Vitro Against Chloroquine-Resistant Plasmodium Falciparum. Lancet 1983, 2, 73-74.

Lafaix, C.; Rey, M.; Diop Mar, I.; Nouhouayi, A. Esεai de traitement curatif du paludisme pour un nouvel antipaludique de synthese, le 16,126 RP. Bull . Soc . Med. Afr. Noire Langue Fr. 1967, 22, 546-551.

Li, Y.; Hu, Y.; Huang, H.; Zhu, D.; Huang, W.; Wu, D.; Qian, Y. Hydroxypiperaquine Phosphate in Treatment of Falciparum Malaria. Chin. Med. J. 1981a, 94 , 301-302.

Li, Y.r Qin, Y.; Qu, Y.; Gong, J. Hydroxypiperaquine Phosphate in Treating Chloroquine Resistant Falciparum Malaria. Chin. Med. J. 1981b, 94 , 303-304.

Li, Y.; Chen, L. ; Dai. Z.-R.; Gong, J.-Z. Antimalarial Activiti of Hydroxypiperaquine and its Phosphate against Plasmodium Berghei and P. cynomolgi . Acta Pharmacol . Sinica 1984, 5, 57-6

Li, J. ; Huang, W.-J. Effects of artesunate, pyronaridine and * hydroxypiperaquine on chloroquine-sensitive and chloroquine-resistant isolates of Plasmodium falciparum in vitr Acta Pharmacol . Sinica 1988, 9 , 83-86.

Lin, A. J. ; Klayman, D. L. ; Milhous, W. K. Antimalarial Activit of New Water-Soluble Dihydroartemisinin Derivatives. J ^" . Med . Chem. 1987, 30 , 2147-2150.

Lin, C; Qu, F.-Y.; Zhou, Y.-C. Field Observations on the Antimalarial Piperaquine. Chin. Med. J. 1982, 95 , 281-286.

Milhous, W. K. ; Weatherly, N. F.; Bowdre, J. H.; Desjardins, R. E. In vitro activities of and mechanisms of resistance to antif antimalarial drugs. Antimicrob. Agents Chemother. 1985, 27 , 525-530.

Oduola, A. M. J. ; Weatherly, N. F.; Bowdre, J. H. ; Desjardins, E. Plasmodium falciparum: Cloning by Single-Erythrocyte Micromanipulation and Heterogeneity in vitro. Exp. Paraεitol . 1988, 66 , 86-95.

Osdene, T. S.; Russell, P. B. ; Rane, L.

2,3,7-Triamino-6-εubstituted arylpteridines. A new series of potent antimalarial agents. J. Med. Chem. 1967, 10 , 431-434..

Payne, D. Spread of Chloroquine Resistance in Plasmodium falciparum. Parasitol . Today 1987, 3 , 241-246.

Pearson, D. E.; Jones, W. H. ; Cope, A. C. Synthesis of Monoalkyl-substituted Diamines and their Condensation Products with 4,7-Dichloroquinoline. J. Amer. Chem. Soc. 1946, 68 , 1225-1229.

Remingtons Pharmaceutical Sciences. 17th Ed., 1985, A. R Gennaro, Ed., Mack Publishing Co., Easton, PA.

Schmidt, L. H.; Vaughan, D. ; Mueller, D. ; Crosby, R.; Hamilton, R. Activities of Various 4-Aminoquinolines Against Infections with Chloroquine-Resistant Strains of Plasmodium falciparum. Antimicrob. Agents Chemother. 1977, 22, 826-843.

Singh, T. ; Hoops, J. F.; Biel, J. H. ; Hoya, W. K. ; Stein, R. G. Cruz, D. R. Antimalarials. "Distal" Hydrazine Derivatives of 7-Chloroquinoline. J. Med. Chem. 1971, 24, 532-535.

Sowunmi, A.; Salako, L. A.; Walker, O.; Ogundahunsi, O. A. T. Clinical efficacy of mefloquine in children suffering from chloroquine-resistant Plasmodium falciparum malaria in Nigeria. Trans. R. Soc. Trop. Med. Hyg. 1990, 84 , 761-764.

Sturchler, D. How much Malaria is there Worldwide? Paraεitol . Today 1989, 5, 39-40.

Tyman, J.; Ghorbanian, S.; Muir, M.; Tychopoulous, V.; Bruce, I.; Fisher, I. Improved Nucleophilic Displacements in N-Methylpyrrolidinone as a Solvent. Synthetic Comm. 1989,29, 179-188.

Warhurst, D. C. Chloroquine-Resistant Rodent Malaria and the Long-Acting Antimalarial 12,278 R. P. Trans. R. Soc. Trop. Med. Hyg. 1966, 60 , 565-566.

Watkins, W. M.; Sixsmith, D. G. ; Spencer, H. C. ; Boriga, D. A.; Kariuki, D. M.; Kipingor, T. ; Koech, D. K. Effectiveness of Amodiaquine as Treatment for Chloroquine-Resistant Plasmodium Falciparum Infections in Kenya. Lancet 1984, 357-359.

Zhang, K.; Zhou, J. ; Wu, Z.; Huang, Q. Susceptibility of Plasmodium falciparum to Chloroquine, Piperaquine, Amodiaquine, Mefloquine and Quinine with In Vitro Microtechnique in Hainan Island. Chin. J. Parasitol . Parasitic Diε. 1987, 5, 165-169.