DNA threading bis(9-aminoacridine-4-carboxamides): effects of piperidine sidechains on DNA binding, cytotoxicity and cell cycle arrest.

We describe the synthesis of a series of DNA-threading bis(9-aminoacridine-4-carboxamides) comprising ethylpiperidino and N-methylpiperidin-4-yl sidechains, joined via neutral flexible alkyl chains, charged flexible polyamine chains and a semi-rigid charged piperazine linker. Their cytotoxicity towards human leukaemic cells gives IC(50) values ranging from 99 to 1100 nM, with the ethylpiperidino series generally being more cytotoxic than the N-methylpiperidin-4-yl series. Measurements with supercoiled DNA indicate that they bisintercalate.

Comparative analysis of mutagenic potency of 1-nitro-acridine derivatives.

The anticancer effect of 1-nitro-9-hydroxyethylamino acridine (C-857), a compound belonging to the 1-nitroacridine class, has been well documented. Despite its therapeutic efficacy, the clinical development of C-857 has been impeded partly due to its high systemic toxicity. In an effort to enhance antitumor efficacy and lower toxicity, derivatives of C-857 have been synthesized with substitutions made at position C-4 and/or an esterified hydroxyl group in side chain at the C-9 position. The introduction of a methyl group at C-4 resulted in C-1748, which has a significantly higher therapeutic efficacy and is being clinically developed as an anticancer agent for solid tumors. The present study was undertaken to correlate the mutagenicity of C-857, C-1748, C-1790, C-1872 and C-1873 with their cytotoxicity and their anti-tumor efficacy. The mutagenicity of these drugs was determined using three Ames Salmonella typhimurium strains TA1537, TA98 and TA102. The bacteria were treated with different molar concentrations, ranging from 10(-3) to 10(-12) M, of the drugs and drug-induced histidine revertants were then counted after a 48 h incubation. C-1748 did not induce any revertants in both TA1537 and TA98 at a dose of 10(-6) M, whereas, C-857 at the same dose induced approximately 842 and approximately 1034 revertants respectively. In TA102, mutagenicity was lower than observed with TA98 and TA1537 with highest revertants observed at 10(-5) M with C-857 (approximately 606) and C-1748 (approximately 108). Higher mutagenicity was observed in the derivatives C-1790, C-1872 and C-1873 compared to C-1748, but lower than C-857. These studies demonstrate that C-1748 has the least mutagenic potential, with a much higher antitumor effect in prostate cancer and is a promising chemotherapeutic agent for clinical development.

Alleviation of mutagenic effects of polycyclic aromatic agents (quinacrine mustard, ICR-191 and ICR-170) by caffeine and pentoxifylline.

Previous studies performed by others indicated that apart from its other biological effects, caffeine (CAF) may have a role in protection of organisms against cancer. However, biological mechanism of this phenomenon remained unknown. Recent studies suggested that caffeine can form stacking (pi-pi) complexes with polycyclic aromatic chemicals. Therefore, one might speculate that effective concentrations of polycyclic aromatic mutagens could be reduced in the presence of caffeine. Here we demonstrate that caffeine and another xanthine, pentoxifylline (PTX), effectively alleviate mutagenic action of polycyclic aromatic agents (exemplified by quinacrine mustard (QM), 2-methoxy-6-chloro-9-(3-(2-chloroethyl)aminopropylamino)acridine.2HCl (ICR-191) and 1,3,7-propanediamine-N-(2-chloroethyl)-N'-(6-chloro-2-methoxy-9-acridinyl)-N-ethyl.2HCl (ICR-170)), but not of aliphatic mutagens (exemplified by mechlorethamine), in the recently developed mutagenicity test based on bacterium Vibrio harveyi. Biophysical studies indicated that caffeine and pentoxifylline can form stacking complexes with the aromatic agents mentioned above. Molecular modeling also confirmed a possibility of stacking interactions between examined molecules.

Frameshift mutations induced by three classes of acridines in the lacZ reversion assay in Escherichia coli: potency of responses and relationship to slipped mispairing models.

The frameshift mutagenicity of 9-aminoacridine (9AA) was compared with that of quinacrine, the acridine mustards ICR-191 and quinacrine mustard (QM), and the nitroacridine Entozon in the lacZ reversion assay in Escherichia coli. As intercalating agents, 9AA and quinacrine cause mutations through noncovalent associations with DNA. Mustards and nitroacridines form covalent adducts in DNA and give rise to different spectra of mutations. Quinacrine and 9AA most effectively induced -1 frameshifts in a run of guanine residues, with 9AA being the more potent mutagen. They also induced +G frameshifts. The acridine mustard ICR-191 was a stronger mutagen than 9AA, owing largely to its potent induction of +G frameshifts. QM induced +G frameshifts more strongly than did its nonreactive counterpart quinacrine. The nitroacridine Entozon differed from the other acridines in being a potent inducer of -2 frameshifts, but it was less effective in inducing +/-1 frameshifts. Quinacrine, although a simple intercalator, induced all five kinds of frameshift mutations detected in the assay, as did the acridine mustards. Although +A and -A frameshifts were induced, adenine runs were less susceptible to acridine mutagenesis than guanine runs. The patterns of frameshift mutagenicity in the lacZ assay are similar to those in an assay based on the reversion of mutations in the tetracycline-resistance gene of the plasmid pBR322. The similarity suggests that the responses reflect the inherent bacterial mutagenicity of the compounds in the local sequence context and are not highly dependent on the broader sequence context. The results are interpreted with respect to slipped mispairing models of frameshift mutagenesis.

Identification of ICR170-induced XPD mutations in UV-sensitive CHO cells.

We highlight selected contributions of Dr. Richard Setlow that contributed to our earlier understanding of excision repair processes and set the stage for dissecting nucleotide excision repair (NER) in mammalian cells through molecular genetics. More than 20 years ago, large-scale screens for UV-sensitive mutants of hamster CHO cells isolated approximately 200 mutants, many of which were assigned to the XPD/ERCC2 complementation group, but the nature of the mutations was not determined. The XPD protein performs not only an essential viability function as a structural component of transcription initiation factor TFIIH, but also an NER function as a 5' to 3' DNA helicase within TFIIH that unwinds DNA on the 3' side of bulky lesions. Alterations in these XPD functions are responsible for three UV-sensitivity genetic disorders that have distinguishable clinical features. In this study, we sequenced six UV-sensitive ICR170-induced Chinese hamster ovary (CHO) cell mutants that previously were assigned to the XPD complementation group to determine whether they carry frameshift mutations. All six mutants show 3- to 5-fold increased hypersensitivity to UV irradiation, similar to the XPD mutant prototype UV5. Even though ICR170 is a strong frameshift mutagen, all six cell lines contain base substitution mutations, five of which are unique among all mutations identified so far in human and rodent cells. The sixth mutation was identical to the R75W mutation previously found in CHO UVL-1. The results presented here contribute to a mutation database that should prove useful in structure-function studies of this unique DNA-structure-specific helicase and its complex mutant phenotypes.

Lack of modification by environmental estrogenic compounds of thyroid carcinogenesis in ovariectomized rats pretreated with N-bis(2-hydroxypropyl)nitrosamine (DHPN).

The effects of environmental estrogenic compounds, soy isoflavone mixture (SI), genistein (GEN), and nonylphenol (NP), and the possible goitrogen 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), on thyroid carcinogenesis were investigated in ovariectomized (OVX) female rats. Five-week-old OVX F344 rats were given a single subcutaneous injection of N-bis(2-hydroxypropyl)nitrosamine (DHPN; 2400 mg / kg, body weight) or vehicle alone. Starting 1 week later, GEN (250 or 25 ppm in diet), SI (400 ppm in diet), NP (250 or 25 ppm in diet), MX (30 ppm, in drinking water), sulfadimethoxine (SDM), a known thyroid tumor-promoter (1000 ppm in drinking water), or beta-estradiol 3-benzoate (EB), a synthetic estrogen (0.5 mg in cholesterol pellet, s.c.) were administered for 12 weeks. SDM and EB were included as positive controls. At sacrifice the major organs including the thyroid, pituitary, liver, kidney, uterus, vagina, brain and pancreas were collected and histopathological observation was performed. Thyroid weights were significantly increased (P < 0. 001) only in the SDM treatment group and pituitary weights were elevated with SDM (P < 0.05) and EB (P < 0.001). Kidney and uterus weights were also significantly increased (P < 0.05) by EB. Histopathologically, proliferative lesions of the thyroid were only observed in the SDM treatment group and of the pituitary in the SDM or EB treatment groups. Renal tubule lesions, uterine squamous metaplasia, vaginal keratinization and telangiectasia of pancreatic islets were also observed with EB. There were no organ weight changes or histopathological lesions in the major organs, including the thyroid, in the GEN, SI, MX or NP treatment groups. Our results thus indicated a lack of modifying effects on thyroid carcinogenesis in female OVX rats, in agreement with our previous finding in males.

Inhibition of mammalian topoisomerase I by 1-nitro-9-aminoacridines. Dependence on thiol activation.

The cytotoxic activity, susceptibility to thiol activation and ability of eight 1-nitroacridine derivatives to stabilize the topoisomerase I-DNA cleavable complex, were compared. Among the acridines tested three compounds exhibited high ability to stabilize the cleavable complex. This ability was correlated with susceptibility to thiol activation as well as with cytotoxic activity. Our results suggest that 1-nitroacridine-DNA adducts interfering with topoisomerase I action may contribute to the lethal effects of some 1-nitroacridine derivatives.

Induction of frameshift mutations in cultured mammalian cells within a transfected sequence containing a poly(dC-dA).poly(dT-dG) microsatellite.

A cultured mouse cell line with an integrated copy of a plasmid that contains a short dinucleotide repeat sequence (microsatellite) has been used to determine the frequencies and types of mutation induced by two frameshift mutagens. The presence of the microsatellite, which consists of 17 repeats of a poly(dC-dA).poly(dT-dG) sequence, disrupts the reading frame of a gene coding for neomycin resistance. Revertants were selected in G418, and mutations were analyzed by PCR. ICR-170 was found to increase the reversion frequency by ten- to 15-fold at its LD50, although most of the frameshifts that it induced were single-base insertions outside the microsatellite sequence. NA-AAF brought about a more modest increase in mutation frequency, but nearly all of the revertants in the NA-AAF-treated cultures had insertions or deletions of multiples of two base pairs within the DNA segment that included the microsatellite. This system can be modified to include different short tandem repeat sequences as targets for testing of compounds that are suspected of having frameshift-inducing activities.

Imidazoacridinones arrest cell-cycle progression in the G2 phase of L1210 cells.

Imidazoacridinones are a new class of highly potent antineoplastic agents synthesised at the Technical University of Gdansk. The pharmacophoric alkyldiamine group, which is also present in anthracenediones (e.g. ametantrone, mitoxantrone), has been shown to be responsible for their antineoplastic activity. In view of their chemical similarity to anthracenediones, we anticipated that the imidazoacridinones would have a mechanism of action similar to that of these agents and that this would be reflected by a similar influence on cell-cycle progression. Flow cytometry was used to monitor the effect of three derivatives of imidazoacridinone (C-1263, C-1310 and C-1311) on L1210 cell cycle traverse at concentrations ranging from 0.01 to 0.9 microgram/ml, corresponding to their 50% and 90% effective concentrations (EC50 and EC90 values), over times of drug treatment ranging from 1 to 48 h. The results demonstrate that all of the compounds produced a similar effect, inducing preferential and complete arrest (accumulation) of cells in the G2 phase of the cell cycle (i.e. G2 block). The kinetics of the induction of G2 arrest were dependent on both the dose and the duration of treatment. Cell-cycle arrest was reversible for up to about 3 h of treatment, being quite irreversible at longer incubation times. Microscopic inspection of cells performed in parallel with flow cytometry confirmed that imidazoacridinones induced a G2, not a G2/M, block.

Potential neurotoxicity of a novel aminoacridine analogue.

1. A class of compounds, 9-aminoacridines, have long been known to be reversible inhibitors of acetylcholinesterase (AChE-EC 3.1.1.7), the most familiar of which is 9-amino-1,2,3,4-tetrahydroacridine (Tacrine). 2. A novel aminoacridine was synthesised: -2-tertiary-butyl-9-amino-1,2,3,4- tetrahydroacridine (2tBuTHA). 3. In vitro comparisons of the acetylcholinesterase inhibitory potential and neurotoxicity compared to Tacrine were performed using a chemically differentiated neuroblastoma cell line (Neuro 2A). 2tBuTHA, but not Tacrine, was cytotoxic to the neural cell following 20 h exposure, despite being the least potent AChE inhibitor (IC80 AChE 12.53 microM +/- 1.14 s.e.m., Neutral Red Uptake IC50 9.53 microM +/- 0.98 s.e.m., MTT Reduction IC80 14.6 microM +/- 1.43 s.e.m.). 4. In vivo studies used a novel application of a five arm radial maze to assess neuropharmacological effects on working memory in control and Scopolamine (1 mg kg-1 i.p.) treated mice. There was an impairment of short term cognitive function with 2tBuTHA (15 mg kg-1 i.p.), but not Tacrine (10 mg kg-1 i.p.) which improved the Scopolamine deficit as expected. 5. This combined in vitro and in vivo data infers a neurotoxic property for the novel compound 2tBuTHA, a close structural analogue of Tacrine.

Activity and structure relationship of acridine derivatives against African trypanosomes.

48 newly synthesized acridine derivatives of different classes were screened for antitrypanosomal activity. They showed a dose dependent effect on Trypanosoma rhodesiense and T. brucei bloodstream forms measured by the inhibition of esterase activity in a fluorescence based in vitro assay. After analysis of the IC50 and MIC values of the investigated acridines it was obvious that no new compound reached the level of the trypanocidal drugs in use (50 ng/ml). Most of the derivatives had IC50 values in the range of 1 to 10 micrograms/ml. 9 derivatives from different classes of acridines were in vitro active below 1 microgram/ml. Correlations between structure and effect on trypanosomes have been elucidated by comparing the IC50 and MIC values of these compounds, in the course of which no significant differences in the drug susceptibility between T. brucei und T. rhodesiense was noticed. The dialkylaminoalkyl derivatives among the group of the 9-thioacridines were slightly more potent than the mono-alkylated ones. 1,2,3,4-tetrahydro-9-thioacridines showed the influence of higher substituted side chains on the trypanocidal activity in the same way as 9-thioacridines. The corresponding ketones of 9-thioacridines confirmed the tendency of increasing toxicity due to the derivatisation of the dialkylaminoalkyl side chain. Within the series of the 9-aminoacridines the elongation of the side chain did not markedly change the activity, however the IC50 values are generally low between 0.13 and 1.2 micrograms/ml.(ABSTRACT TRUNCATED AT 250 WORDS)

9-[3-(2-Nitro-1-imidazolyl)propylamino]-1,2,3,4-tetrahydroacridine hydrochloride. A novel DNA-affinic hypoxic cell cytotoxin and radiosensitizer. Comparison with NLA-1.

Electron-affinic compounds with strong DNA intercalating properties have demonstrated less than the expected radiosensitization due to restriction of their mobility along the DNA backbone and their lower extravascular diffusion in tumors. A 2-nitroimidazole linked 1,2,3,4-tetrahydroacridine derivative (THNLA-1) has been synthesized as a hypoxia-selective cytotoxin and radiosensitizer with presumably lower DNA-binding affinity due to the perturbation of the planarity in the acridine ring. THNLA-1 is a good hypoxia-selective cytotoxin with a differential toxicity of approximately equal to 11 in V79 cells, but it is approximately equal to 2 times less potent on a concentration basis than NLA-1 (the 2-nitroimidazole linked acridine analog). However, THNLA-1 is a very efficient radiosensitizer, showing a sensitization enhancement ratio (SER) of 3.04 +/- 0.05 at 100 microM at 25 degrees C, and the concentration giving an SER of 1.6(C1.6) is 19.0 +/- 0.5 microM. The therapeutic index, defined as the ratio of the clonogenic IC50 under aerobic conditions for 1-h exposure (IC50A,1h) to the C1.6 value, is 20 for THNLA-1 vs. 11 for NLA-1. THNLA-1's partition coefficient in octanol/water is 0.14 +/- 0.02. Topoisomerase I and II interaction studies with THNLA-1 showed that topoisomerase I-mediated relaxation of supercoiled DNA was inhibited at relatively high THNLA-1 concentrations (> or = 1000 microM), while topoisomerase II-mediated decatenation of kinetoplast DNA remained unaffected even in concentrations toxic in vitro under aerobic conditions. Uptake studies under aerobic conditions showed high intracellular drug concentrations, compatible with the required ones for topoisomerase I inhibition.

Induction of mitotic crossing-over by the topoisomerase II poison DACA (N-[2-dimethylamino)ethyl]acridine-4-carboxamide) in Saccharomyces cerevisiae.

The antitumor agent DACA (N-[2-dimethylamino)ethyl]acridine-4-carboxamide) a new DNA intercalating topoisomerase II poison, was distinguishable from clinical topoisomerase poisons (amsacrine, daunorubicin, doxorubicin and etoposide) in its induction of aberrant colonies in the yeast Saccharomyces cerevisiae D5. It was not only more recombinogenic, but was recombinogenic at non-toxic drug concentrations. DACA at 680 microM (2-h exposure time), induced 1.2% aberrant colonies of which 0.32% were mitotic crossing-over events. The presence of the rad52 mutation abolished mitotic crossing-over and greatly increased drug toxicity. The concentration for 50% inhibition of survival of the rad52 mutant was 100 microM, as compared with 4900 microM for the wild-type. Drug toxicity was marginally increased by the presence of rad3 and rad18 mutations. Rad3 mutations increased the incidence of crossing-over events but had little effect on other mutagenic or recombinogenic events. In contrast, the rad18 mutation increased the incidence of all types of aberrant colonies. The inclusion of hydroxyurea and caffeine, as non-specific repair inhibitors, caused weak and strong inhibition, respectively, of all types of aberrant colonies. Inclusion of the protein-synthesis inhibitor cycloheximide reduced mitotic cross-over but had little effect on the incidence of other aberrations. It is concluded that DACA induces lesions which are repaired by a recombinational repair pathway involving the RAD52 product, and that RAD3 and RAD18 products are each involved in the generation of recombinational events.

Synthesis and antiparasitic activity of new 1-nitro, 1-amino and 1-acetamido 9-acridinones.

The synthesis of 1-amino-10-alkyl-9-acridinones 4 and 5 and of homologous 9-thio-acridinones 7 and 8 are described. Similarly, the synthesis of 1-nitro-4-(2'-aminoethylamino)-9-acridinones 13 and 15, is described. Compound 4 was used as starting material for the preparation of 10-alkyl-9-acridinone dimers 6, bridged with an alpha,omega-diamido chain. Compound 15 was selected for biological investigations on pathogenic parasite strains, and a promising antiamoebic activity could be shown.

Potentiation of 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea's toxicity in vitro by two new bioreductive agents.

Two new bioreductive compounds, 9-[3-(2-nitro-1-imidazolyl)propylamino]acridine hydrochloride (NLA-1) and 9-[2-(2-nitro-1-imidazolyl)ethylamino]acridine hydrochloride (NLA-2), which behave as hypoxic cytotoxins and radiosensitizers, have been investigated for potentiation of 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea's (CCNU) cytotoxic activity in vitro using V-79 cells. The preincubation effect as well as conditions of coadministration of CCNU with each sensitizer have been examined. In this latter case, the median-effect analysis was applied to evaluate whether the phenomenon was additive or synergistic. A clonogenic assay was used to score survival. Both bioreductive compounds, even at very low concentrations, significantly enhance the cytotoxic activity of CCNU under conditions of hypoxic preincubation. The enhancement of CCNU cytotoxicity is dependent upon preincubation time and the concentrations of both CCNU and the specific bioreductive agent. Coincubation of cells under hypoxia with CCNU and each bioreductive agent led to some potentiation, but only at lower survival levels. No chemosensitization was observed under aerobic conditions with either sensitizer.

Photo-enhancement of the mutagenicity of 9-anilinoacridine derivatives related to the antitumour agent amsacrine.

The frameshift mutagenicity of the DNA intercalating drug proflavine is known to be enhanced by photoirradiation of bacterial cultures. To determine whether this phenomenon was also present in acridine-derived antitumour drugs, cultures of Salmonella typhimurium were exposed to the antileukaemia agent amsacrine and the experimental agent N-[2-(dimethylamino)ethyl]acridine-4-carboxamide dihydrochloride (acridine carboxamide) in the presence or absence of visible light. A small increase in mutagenicity was observed with amsacrine but not with acridine carboxamide. A series of analogues of amsacrine were then tested, and a striking relationship was found between the minimum drug concentration for mutagenicity and DNA binding affinity. In each case, photoirradiation was associated with a small increase in mutagenicity. Each of the compounds showing the photo-enhancement effect was capable of reversible one-electron oxidation. It is suggested that this oxidation occurs in bacteria, and that the DNA binding constant of the resulting acridine radical species will increase because of the extra positive charge. This increased DNA binding would be sufficient to explain the photo-enhancement of mutagenicity of these drugs.

Unusual dynamics of killing of cultured Lewis lung cells by the DNA-intercalating antitumour agent N-[2-(dimethylamino)ethyl]acridine-4-carboxamide.

The cytotoxicity of N-[2-(dimethylamino)ethyl]acridine-4-carboxamide (AC; NSC 601,316), a new experimental DNA-intercalating antitumor drug, against a cultured Lewis lung adenocarcinoma cell line was compared with that of the DNA-intercalating antitumor drug amsacrine. In contrast to amsacrine, AC demonstrated self-inhibition of cytotoxicity following short (3-9 h) incubation periods and exponential killing (with a shoulder) after long (24-72 h) periods of incubation. The difference between these drugs was best demonstrated using a constant concentration x time (C x T) exposure (AC, 12 mumol h l-1; amsacrine, 3 mumol h l-1). In contrast to amsacrine, AC was minimally effective over exposure periods of less than or equal to 1 h and maximally effective over intermediate periods (4-6 h). The results suggest the possibility of designing AC administration protocols that maximise the drug's cytotoxicity towards solid tumors, which, because of diffusion barriers, are subjected to longer drug exposures than are well-vascularised tumours.

Pharmacokinetics and toxicity of the antitumour agent N-[2-(dimethylamino)ethyl]acridine-4-carboxamide after i.v. administration in the mouse.

The pharmacokinetics, tissue distribution and toxicity of the antitumour agent N-[2-(dimethylamino)ethyl]acridine-4-carboxamide(AC) were studied after i.v. administration to mice. Over the dose range of 9-121 mumol/kg (3-40 mg/kg), AC displayed linear kinetics with the following model-independent parameters: clearance (C), 21.0 +/- 1.9 1 h-1 kg-1; steady-state volume of distribution (Vss), 11.8 +/- 1.4 l/kg; and mean residence time (MRT), 0.56 +/- 0.02 h. The plasma concentration-time profiles for AC fitted a two-compartment model with the following parameters: Cc, 19.4 +/- 2.3 1 h-1 kg-1; Vc, 7.08 +/- 1.06 l/kg; t1/2 alpha 13.1 +/- 3.5 min; and t1/2Z, 1.60 +/- 0.65 h. AC displayed moderately high binding in healthy mouse plasma, giving a free fraction of 15.9%-25.3% over the drug concentration range of 1-561 microM. After the i.v. administration of 30 mumol/kg [3H]-AC, high radioactivity concentrations were observed in all tissues (especially the brain and kidney), showing a high t1/2c value (37-59 h). At 2 min (first blood collection), the AC concentration as measured by high-performance liquid chromatography (HPLC) comprised 61% of the plasma radioactivity concentration (expressed as AC equivalents/l). By 48 h, 73% of the dose had been eliminated, with 26% and 47% of the delivered drug being excreted by the urinary and faecal routes, respectively; less than 1% of the total dose was excreted as unchanged AC in the urine. At least five distinct radiochemical peaks were distinguishable by HPLC analysis of plasma extracts, with some similar peaks appearing in urine. The 121-mumol/kg dose was well tolerated by mice, with sedation being the only obvious side effect and no significant alterations in blood biochemistry or haematological parameters being recorded. After receiving a dose of 152 mumol/kg, all mice experienced clonic seizures for 2 min (with one death occurring) followed by a period of sedation that lasted for up to 2 h. No leucopenia occurred, but some mild anaemia was noted. There was no significant change in blood biochemistry. A further 20% increase in the i.v. dose (to 182 mumol/kg) resulted in mortality, with death occurring within 2 min of AC administration.

Intraperitoneal administration of the antitumour agent N-[2-(dimethylamino)ethyl]acridine-4-carboxamide in the mouse: bioavailability, pharmacokinetics and toxicity after a single dose.

The pharmacokinetics, tissue distribution and toxicity of the antitumour agent N-[2-(dimethylamino)-ethyl]acridine-4-carboxamide (AC) were studied after i.p. administration of [3H]-AC (410 mumol/kg) to mice. The latter is the optimal single dose for the cure of advanced Lewis lung tumours. AC was rapidly absorbed into the systemic circulation after i.p. administration, with the maximal concentration (Cmax) occurring at the first time point (5 min). There was no reduction in bioavailability as compared with previous i.v. studies, but the shape of the plasma concentration-time profile was considerably different, reflecting a 3-fold lower Cmax value (20.9 +/- 3.6 mumol/l) and a longer t1/2 value (2.7 +/- 0.3 h) as compared with that observed after i.v. administration (1.6 +/- 0.6 h). Model independent pharmacokinetic parameters after i.p. administration were: clearance (C), 17.5 l h-1 kg-1; steady-state volume of distribution (Vss), 14.1 l/kg; and mean residence time (MRT), 1.46 h. High but variable tissue uptake of AC was observed, with tissue/plasma AUC ratios being 5.7 for heart, 8.4 for brain, 18.9 for kidney and 21.0 for liver but with similar elimination t1/2 values ranging from 1.3 to 2.7 h. All radioactivity profiles in plasma and tissues were greater than the respective parent AC profiles and showed prolonged elimination t1/2 values ranging from 21 h in liver to 93 h in brain. However, tissue/plasma radioactivity AUC ratios were near unity, ranging from 0.7 to 1.57, with the exception of the gallbladder (15.6), which contained greater amounts of radioactivity. By 48 h, approximately 70% of the total dose had been eliminated, with the faecal to urinary ratio being approximately 2:1. This i.p. dose was well tolerated by mice, with sedation being the only obvious side effect. No major change was observed in blood biochemistry or haematological parameters. Comparisons of Cmax, tmax and AUC values determined for AC in brain after its i.p. and i.v. administration suggest that the reduction in acute toxicity after i.p. administration is not due to reduced exposure of the brain to AC as measured by AUC but may be associated with the lower Cmax value or the slower rate of entry of AC into the brain after i.p. administration.

The genetic toxicology of acridines.

Acridine and its derivatives are planar polycyclic aromatic molecules which bind tightly but reversibly to DNA by intercalation, but do not usually covalently interact with it. Acridines have a broad spectrum of biological activities, and a number of derivatives are widely used as antibacterial, antiprotozoal and anticancer drugs. Simple acridines show activity as frameshift mutagens, especially in bacteriophage and bacterial assays, by virtue of their intercalative DNA-binding ability. Acridines bearing additional fused aromatic rings (benzacridines) show little activity as frameshift mutagens, but interact covalently with DNA following metabolic activation (forming predominantly base-pair substitution mutations). Compounds where the acridine acts as a carrier to target alkylating agents to DNA (e.g. the ICR compounds) cause predominantly frameshift as well as base-pair substitution mutations in both bacterial and mammalian cells. Nitroacridines may act as simple acridines or (following nitro group reduction) as alkylating agents, depending upon the position of the nitro group. Acridine-based topoisomerase II inhibitors, although frameshift mutagens in bacteria and bacteriophage systems, are primarily chromosomal mutagens in mammalian cells. These mutagenic activities are important, since the compounds have considerable potential as clinical antitumour drugs. Although evidence suggests that simple acridines are not animal or human carcinogens, a number of the derived compounds are highly active in this capacity.