Ind. Eng. Chem. Res. 1993,32,2455-2456
2455
Potential Radioprotective Agents. 3. Ketals, Monothioketals, and Dithioketals Robert T. Blickenstaff,' Stephan M. Brandstadter, Emerson Foster, Shailaga Reddy, and Robert Witt Richard L. Roudebush VA Medical Center, Department of Biochemistry & Molecular Biology, and Department of Radiation Oncology, Indiana University School of Medicine, Indianapolis, Indiana 46202
p-Aminobenzophenone, p-aminophenyl alkyl ketones, and their ethylene ketals, monothioketals, and dithioketals were synthesized and tested for radioprotective activity in mice. Radiation was carried out with a linear accelerator 30 min after compounds in soybean oil were injected intraperitoneally. High activity was found for p-aminobenzophenone (4) and for p-aminopropiophenone (2), and its ethylene ketal (6), but monothio- and dithioketals exhibited little or no activity. The acute toxicity of p-aminopropiophenone (2) was greatly reduced by converting it to its Schiff base (13) with salicylaldehyde. A
Introduction The possibility of nuclear exposures from accidents or warfare points up the necessity of developing effective drugs that protect humans against radiation damage. A further application of such a radioprotective compound could be in conjunction with the treatment of cancer patients by radiation. If the compound selectively protected healthy tissue more than cancerous tissue, heavier doses of radiation could be used, resulting in greater destruction of the tumor. For over 40 years the ability of some compounds to protect against radiation has been known. Classes of compounds include mercaptans, di- and trisulfides, phosphorothioates, alcohols, acid hydrazides, imidazoles, benzofurans, amine oxides, and thiazolidines (Foye, 1981). The need still exists, however, for compounds that are less toxic and are orally active (Walker, 1988). p-Aminopropiophenone (2) is known to protect mice against lethal X-irradiation, presumably by producing tissue hypoxia (Storer and Coon, 1950); it reduces the capacity of the blood to carry oxygen (Salerno and Friedell, 1954). Given to dogs, it produces methemoglobinemia; it was not radioprotective alone, but produced a high degree of protection when given with S-fl-aminoethylisoethiouronium bromide (Blouin and Overman, 1962). In the present work we have compared p-aminopropiophenone with a series of analogs and derivatives for a structureactivity study.
Experimental Section Synthesis. 4-Aminobutyrophenone (3)was synthesized by rearrangement of N-phenylbutyramide in polyphosphoric acid at 190-200 "C (Staskun, 1964). It was chromatographed on A1203, eluted by CH2C12, and recrystallized from CHzCl2-petroleum ether (2:l). The ethylene ketals (6 and 6) were prepared with ethylene glycol, p-toluenesulfonic acid, and benzene with the aid of a Dean-Stark trap, refluxed 48 h. The crude product was purified by flash chromatography on Si02 and eluted with EtOAc in hexane. Thioketals 7-9 were prepared with mercaptoethanol, BF3-etherate, and dioxane, held at room temperature overnight. The crude product was purified by chromatography on A1203 and eluted by benzene-petroleum ether (2:l). Dithioketals 10-12 were prepared with ethanedithiol, BF3-etherate, and dioxane, helt at temperature 30-40 min.
n
7,8,Q
Figure 1. Synthesis of ketals, mono- and dithioketals, and Schiff base.
The crude product was purified by chromatography on A1203 and eluted by benzene-petroleum ether (2:l). Structures of all new products were verified with IR and NMR spectra, in addition to elemental analyses. Compounds 1, 2, and 4 in Table I were obtained from Aldrich Chemical Company. Radiation-Protective Evaluation. Male, Swiss ND4 mice were obtained from Harlan Industries and housed five to a cage. Thirty minutes before irradiation with 950 cGy of 6-mV photons produced by a linear accelerator, they were injected with a solution or suspension of the test compound in soybean oil a t a dose level of 0.78 mequiv/ kg. The control group was given soybean oil; most of them died 5-15 days postinjection. The values reported in Table I represent 30-day survival. A linear accelerator was used because this is the most commonly used equipment for therapeutic radiation, and one of our future research projects is to use a selective radioprotective agent in a clinical setting. 4-Amino-N-salicylidenepropiophenone (13). A warm solution of 320 mg (2.15 mmol) of p-aminopropiophenone in 3.56 mL of ethanol containing 0.10 mL of H2O and 15.4 mg of KOH was stirred while salicylaldehyde (262 mg, 2.15 mmol) was added dropwise. The solution was stirred at room temperature for 1h, refrigerated overnight, and filtered to give 367 mg of crude Schiff base in two crops. Recrystallization in ethanol gave the analytical sample: mp 120-122 "C. Anal. Calcd for CleH15N02: 75.87% C; 5.97% H; 5.53% N. Found 75.65% C; 6.02% H; 5.41% N. Toxicity Comparison. p-Aminopropiophenone (2) and its Schiff base (13)with salicylaldehyde were dissolved
QSSS-5SS5/93/2632-2455$04.Q~JQ 0 1993 American Chemical Society
2456 Ind. Eng. Chem. Res., Vol. 32, No. 11, 1993 Table I. Ketones, Ketals, Monothioketals, and Dithioketals R
R 1 0 2 0
3 0 4 0 5 OCH2CH20 6 OCHzCH20
7 OCH2CH2S 8 OCHzCHfi 9 OCH2CH2S 10 SCHzCHzS 11 SCH2CH2S 12 SCHzCHzS a
R' CH3 CZH6 CaH7 CsH6 CHs CzH5 CH3 CzH6 C& CH3 C2H5 C&
mp, OC 85-86
analyses0 radiation survivorsb H,Nc
C,H,N H,Nd H,N,SC oil C, H, N, S 115-116 H,N,Sf 83-84 C,H,Ng oil C, H, Nh 93-94 C,H,N,S 91-93 97-99 89-90
15/21,71% 29/30,96 % 6/8,75% 39/40,97% 0/20,0% 19/20,95% 1/21,5% 3/19,16% 8/29,42% 2/20,10% 1/20,5% 5/20,25%
Analyses for elements shown agree with calculated values within
0.4%;analyses for other elements are shown below. Male, Swiss
mice were injected intraperitoneally with suspensions or solutions of the test compound in soybean oil at a dose level of 0.78 mequiv/kg 30 min before irradiation with 950 cGy of 6-mV photons. Seven out of 105 mice in the control group (soybean oil only) survived 7%. Calculated: 73.59% C. Found: 73.16% C. dCalcd: 68.37% C. Found 67.75% C. e Calcd: 61.50% C. Found 60.92% C. f Calcd 70.00% C. Found 69.43% C. 8 Calcd: 30.34% S. Found: 30.92% S.hCalcd 28.45% S. Found 29.23% S. Table 11. Toxicity of pAminopropiophenone compd 2 13
0.60 8/10b 10/10
0.78 8/10 loll0
1.17 41 10 10/10
1.56 4110 10/10
0 Administered suspended in soybean oil intraperitoneally to male, Swiss mice. b Survivors after 10 days.
in soybean oil and injected intraperitoneally in male. Swiss ND4 mice a t four different dose levels. Ten-day survivals are shown in Table 11.
Results and Discussion Our initial finding of high radioprotective activity of p-aminopropiophenone (2), its ethylene ketal (6), and p-aminobenzophenone (4) (Blickenstaff et al., 1993), prompted the synthesis and testing of additional ketals; in particular, thioketals inasmuch as many other S-containing compounds are radioprotective (Foye, 1981). As shown in Table I, conversion of p-aminopropiophenone (2) to its corresponding ethylene ketal (6) retained the high activity of the former, but conversion of p-aminoacetophenone (1) to its ethylene ketal (5) destroyed activity. The low activity of 5 implies that the ketal ring remains intact a t least throughout the radiation, or else 71% protection would have been observed had it hydrolyzed to the starting ketone. In the homologous series p-aminoacetophenone (l), p-aminopropiophenone (2), and p-aminobutyrophenone (3),radioprotective activity passes through a maximum with 2, but toxicity increases with increasing molecular weight. In 40 mice injected with 2,lO died in the first few days; nearly all deaths from radiation occurred in the control group 5-15 days after exposure. Of 20mice injected
with 3, 12 died in the first few days. This is a seriousproblem for this class of compounds, but fortunately it is easily resolved. Conversion of 2 to its corresponding Schiff base (13) with salicylaldehyde gave a remarkably nontoxic derivative even at twice the dose level used in the radiation experiments (Table 11). Schiff bases of this type are stabilized by hydrogen bonding between the phenolic hydroxyl and the nitrogen atom. Furthermore, compound 13 is highly protective, giving 21 survivors out of 21 tested.
Conclusions Ten new compounds have been synthesized and assayed for radioprotective activity. Mono- and dithioketals are less active than their keto precursors. The toxicity of p-aminopropiophenone was ameliorated by converting it to the corresponding Schiff base with salicylaldehyde, and high radioprotective activity was retained. Acknowledgment We thank Gerry Reed and Jeff Gray for expert technical assistance in the radiation experiments. We are grateful to the Chemistry Department of IUPUI for the use of a Varian 390 NMR spectrophotometer. This work was supported by the Department of Veterans Affairs. Nomenclature CGY centigrays CH& methylene chloride EtOAc ethyl acetate mequiv/kg milliequivalents of compound administered per kilogram of body weight of the test animal Schiff base compound containing an -N==CRz group Literature Cited Blickenstaff, R. T.; Brandstadter, S. M.; Reddy, S.; Witt, R.; Lipkowitz, K. B. Potential Radioprotective Agents. 11. Substituted Anilines. J. Pharm. Sci. 1993, in press. Blouin, L. T.; Overman, R. R. Protection of the Irradiated Dog by Aminoethylisothiouronium (AET)and p-Aminopropiophenone (PAPP). Radiat. Res. 1962, 16,699. Foye, W. 0. Radioprotective Drugs. In Burger's 'Medicinal Chemistry", 4th ed.; Wolff, M. E., Ed.; Wiley: New York, 1981; Part 3, pp 11-45. Salerno, P. R.; Friedell, H. L. Further Studies on the Relationship between Oxygen Tensions and the Protective Actions of Cysteine, Mercaptoethylamine and p-Aminopropiophenone. Radiat. Res. 1954, 1, 559, Abstr. No. 77. Staskun, B. Nuclear Acylation of Arylamines. J. Org. Chem. 1964, 29, 2856.
Storer, J. B.; Coon, J. M. Protective Effect of Para-Aminopropiophenone Against Lethal Doses of X-Radiation. Roc. SOC. Erp. Biol. Med. 1950, 74, 202. Walker, R. I. Requirements of Radioprotectors for Military and Emergency Needs. Pharmacol. Ther. 1988,39, 13. Received for review May 17, 1993 Accepted July 25, 1993. Abstract published in Advance ACS Abstracts, October 1, 1993.
