Stephen S. Sternberg

THE TOXIC EFFECTS OF 6‐MERCAPTOPURINE AND RELATED COMPOUNDS

The present review of the toxic effects of 6-mercaptopurine (6-MP) in experimental mammals has two primary aims. The first and more practical purpose concerns risks to be anticipated during therapeutic use. The second aim concerns the specific cytocidal effects of 6-MP as a possible contribution to the elucidation of its mechanism of action. As such, the study is a continuation of work of this laboratory by which i t has been possible to differentiate the cytotoxic effects of folic acid antagonists such as aminopterinl and certain diaminopyrimidines2s2a from those of the nitrogen mustards and mustardlike compounds, such as HN2 and TEM; and from those of 2,6-diaminopuIn the present instance, tissue changes induced by 6-MP are compared with those caused by 2,6-diaminopurine (2,6-DAP) and by other purine analogs. Certainly if the action of 6-MP is to be explained in terms of a specific metabolite antagonism, then such an exposition must draw sustenance not only from differences in toxic properties between 6-MP and, for example, 2,6-DAP1 but must also reconcile similarities in the effects of these compounds. Toxicity studies. FIGURE 1 provides a list of the purines employed and presents graphically their relative potency in terms of toxicity in Swiss male mice. In most instances, two schedules of dosage have been used, i.e., single and five successive daily injections. The agents are listed in an order largely based on their relative potency by chronic administration. The bars topped by arrows refer to the fact that the highest dose employed, the quantity indicated by the height of the serrated column, is less than LDso. Data are taken from previous reports45 , 6. and from unpublished observations of the author^.^ The graph illustrates that there is more than a hundredfold difference in potency between the two extreme members of the series. Moreover, 6-MP is neither the most nor least toxic of its close structural analogs. The extent by which the toxicity of each compound is enhanced by cumulative action can be appreciated by comparing the LDE0 of single doses with that of multiple doses. It may be seen that most of the compounds, including 6-MP, exhibit relatively minor cumulative activity, i.e., the LDso in mgm./kg./day for five daily doses, when expressed as a fraction of the single median lethal dose, ranges between one fourth in the case of adenine to more than one half in the case of 6-chloro-

THE TOXICITY OF ACTINOMYCIN D

The high toxicity of the actinomycins in mammals was first described after the isolation of the A complex from Streptomyces antibioticus by Waksman and Woodruff,’ whose early work directed attention to what appeared a t first a selective atrophy of the mammalian spleet1.2,~ These investigators also showed that doses not greatly exceeding the LDSo killed, within less than 24 hours. Somewhat more delayed fatalities were caused by lower single doses or by repeated daily injections of small fractions of the acute LDbo. Anorexia, diarrhea, disturbances of locomotion, and weakness were features of chronic poisoning. Hackmann’s later studies of the C complex showed that the spleen was not the only susceptible lymphoid tissue: thymus and lymph nodes were equally or, possibly, more sensitive sites of damage.4t6 He also reported mitotic inhibition in the crypts of Lieberkuhn, as well as degeneration and desquamation of the intestinal epithelium. Untoward clinical effects such as stomatitis, nausea, and diarrhea were considered by him to be related manifestations of gastrointestinal damage. Our present interest in these compounds stemmed primarily from the description given by Hackmann and by Schmidt-Kastner of the biosynthetic actinomycin F1 (KSJ . 6 J Since F1 had antitumor activity but was significantly less toxic than earlier actinomycins, it seemed worthwhile to learn whether or not it might be distinguished by other qualitatively different pharmacological effects. For a more meaningful comparison, we decided to become familiar first with the toxic properties of actinomycin D8t9 in different laboratory animal species. This is the subject of the present communication, which extends the earlier observations of other

Effects of N-Methylformamide and Related Compounds in Mouse Sarcoma 180.

Summary and Conclusions 1. Formamide and its more potent N-methyl derivative have been described as transient inhibitors of the growth of mouse sarcoma 180. This effect is not a property of formamides in general since other compounds containing the formamide moiety have failed to affect the growth of the tumor. 2. N-methylformamide has been shown to exert its effects when therapy is started either 24 or 96 hours after implantation of the tumor or when the agent is given in a single, large dose. Further, the compound has been found equally effective whether given by the oral or intraperitoneal route. 3. Histologic changes of a non-specific nature have been described in tumors from animals treated with N-methylformamide. 4. In view of the structural resemblance between formamide and urethane, the actions of both agents have been compared. Urethane, like formamide, inhibits the growth of S-180; however, treatment with the former substance causes toxic manifestations not encountered in mice given formamide. 5. Several conceivable mechanisms have been proposed to account for the inhibitory effects of the formamides. Further studies of these suggested modes of action may provide useful information concerning biochemical mechanisms involved in the growth of tumors. 6. In view of the hepatotoxicity of formamide and its N-methyl derivative it is not expected that these agents will prove therapeutically useful.

Long-term and multigeneration toxicity studies with cyclohexylamine hydrochloride

Cyclohexylamine (CHA), the metabolite of cyclamate produced in varying degree by gastrointestinal microorganisms, was subjected to a 2-year multi-generation feeding study in rats, at dosages of 15, 50, 100, and 150 mg/kg/d. Observations included growth, feed efficiency, clinical and hematological tests, reproduction, teratology, mortality and gross and microscopic pathology. Rats from the first litters of each generation from F0 through F4 were mated to produce the next succeeding generation. Those from the second litters of F1 through F4 were also mated, half the dams being delivered by hysterotomy for teratological examination, while litters from the other half were raised to maturity. Except for some non-progressive growth retardation in the higher dosage groups, due to lower food consumption, the physical and clinical observations in the test groups fell substantially within normal limits and were not significantly different from the untreated controls. Reproduction rates were normal in all groups but at the higher dosages the size of the litters and their weaning weights were slightly reduced. At the 150 mg/kg level, histopathological examination revealed mucosal thickening of the bladder walls and evidence of renal calcification; however, no bladder tumors were seen, such as occurred in the chronic feeding study in which rats received 2500 mg/kg of a cyclamate: saccharin (10 : 1) mixture. A significantly higher incidence of testicular atrophy, characteristic of aged rats, was observed in the F0 group at the highest dosage level; however, these males continued to be fertile, in two cases up to 6 consecutive matings.

Toxicologic studies of petrolatum in mice and rats

Abstract The present studies were undertaken as a basis for determining the status of food grade petrolatum as a food additive under the terms of the Food Additives Amendment of 1958. The investigations of the petrolatum included 2-year rat feeding studies and single subcutaneous injections in mice followed by observation over an 18-month period. Previous work and the considerations leading to regulatory interest in the potential carcinogenicity of paraffin hydrocarbons have been reviewed in the extensive report on the toxicity of petroleum waxes by Shubik et al. (1962). Three specific color grades of petrolatum which met either USP XVI or NF XI (White or Yellow Petrolatum) were selected to represent the range of degrees of decolorization of petrolatum commonly used as direct food additives, or which may be incorporated in foods as an indirect additive, incidental to its use in processing.

Susceptibility of Intestinal Crypt Epithelium to Hydroxyurea and Other Antitumor Agents

The outstanding limitation in the effective use of most antitumor agents is selective damage in normal cell renewal systems in vivo. These systems contain undifferentiated or partially differentiated precursors of various mature cells such as leukocytes, erythrocytes, villar epithelium of intestine, sperm, and non-germinal components of epidermis. Normally the systems operate to maintain a steady balance between the birth of differentiated descendants and their continuous loss from the organism (6). The susceptibility of two of the cell renewal populations, that is, hematopoietic tissues and digestive tract epithelium, commonly determines the upper limit of doses of antitumor drugs. It is likely that lesions in the digestive tract are the major determinant of maximal dosage since these give rise to distressing symptoms such as oropharyngitis, nausea, and abdominal cramps and cause alarming signs of intoxication such as diarrhea and ulceration of pharyngeal and intestinal mucosa.

Hydroxyurea: inhibition of DNA-synthesis and selective susceptibility of S-phase cells in vivo.

The cytotoxic effects of hydroxyurea and of related hydroxamic acid derivatives in vivo are briefly described. They occur selectively in tissues with high rates of cell renewal and they are of brief duration. Tissue concentration of hydroxyurea diminish rapidly as the result of renal excretion and metabolism; there is a close temporal relation between the physiological disposition of the agents and the cytotoxic changes. Hydroxyurea induces an immediate inhibition of DNA synthesis in proliferating tisdsues such as thymus, small intestine, and regenerating liver. Autoradiographic studies of mouse duodenum using tritiated thymidine have shown that the lethal susceptibility to hydroxyurea is restricted to cells in the S-phase of the mitotic cycles. Cells in G 1 , G 2 , and M are not damaged by the agent.