Lip symmetry following rotation advancement cleft lip repair in 5-year-old children treated by Ralph Millard and Ron Pigott.

Objective: To compare the symmetry of the lip following Rotation-Advancement cleft lip repair by Millard and Pigott and to investigate the effect on the symmetry of cleft side and gender by using different surgical protocols. Symmetry following cleft surgery was compared to that of non-cleft children. Design: Retrospective study of photographs of children aged 5 years. Setting: Three decades of post-operative photographs of children treated by Millard and Pigott. Patients: Eighty-nine children treated by Millard, 87 by Pigott and 91 non-cleft children. Interventions: Photographs were assessed using the Symnose Computer program, a rapid semi-objective quantitative assessment of lip symmetry. Main Outcome Measures: Asymmetry score for each surgeon, and non-cleft children. Results: There was no significant difference in the median lip % mismatch score of Millard, 36.65% and Pigott, 38.52%. Right-sided clefts showed better symmetry than left-sided clefts for Millard (p<.001). This was reversed for Pigott (P=.0121). There was a difference (P<.001) between the symmetry of the two cleft cohorts and the non-cleft children (asymmetry 19.9%), and between Millard's outcomes following different lip surgical protocols (P < .0001), but no difference between Pigott's outcomes using different palate surgical protocols (P = 0.59). Conclusions: Cleft lip repair by Millard and Pigott resulted in similar lip asymmetry (37% and 39% symmetry mismatch, respectively). Lip surgical protocol and cleft side may affect lip asymmetry. Palate surgery did not affect lip asymmetry. Following cleft surgery, children were more asymmetric than non-cleft children.

Quantitative autoradiographic mapping of herpes simplex virus encephalitis with a radiolabeled antiviral drug.

2'-Fluoro-5-methyl-l-beta-D-arabinosyluracil (FMAU) labeled with carbon-14 was used to image herpes simplex virus type 1-infected regions of rat brain by quantitative autoradiography. FMAU is a potent antiviral pyrimidine nucleoside which is selectively phosphorylated by virus-coded thymidine kinase. When the labeled FMAU was administered 6 hours before the rats were killed, the selective uptake and concentration of the drug and its metabolites by infected cells (defined by immunoperoxidase staining of viral antigens) allowed quantitative definition and mapping of HSV-1-infected structures in autoradiograms of brain sections. These results show that quantitative autoradiography can be used to characterize the local metabolism of antiviral drugs by infected cells in vivo. They also suggest that the selective uptake of drugs that exploit viral thymidine kinase for their antiviral effect can, by appropriate labeling, be used in conjunction with clinical neuroimaging techniques to define infected regions of human brain, thereby providing a new approach to the diagnosis of herpes encephalitis in man.

Characteristics and products of the reductive degradation of 3-nitro-1,2,4-triazol-5-one (NTO) and 2,4-dinitroanisole (DNAN) in a Fe-Cu bimetal system.

It has been shown previously that, under acidic conditions, 3-nitro-1,2,4-triazol-5-one (NTO) and 2,4-dinitroanisole (DNAN) degrade in the presence of iron/copper bimetal particles; the reactions can be modeled by pseudo-first-order kinetics. This study investigates the reaction mechanisms of the degradation processes under different conditions. Batch studies were conducted using laboratory-prepared solutions and an industrial insensitive munition-laden (IMX) wastewater. The influence of parameters such as initial pH of the solution, copper/iron (Fe-Cu) contact, and solid/liquid ratio were systematically investigated to assess their impact on the reaction kinetics. These parameters were subsequently incorporated into pseudo-first-order decomposition models for NTO and DNAN. The activation energies for the degradation reactions were 27.40 and 30.57 kJ mol-1, respectively. Degradation intermediates and products were identified. A nitro-to-amino pathway, which ultimately may lead to partial mineralization, is postulated. The amino intermediate, aminonitroanisole, was detected during DNAN degradation, but for NTO, aminotiazolone is suggested. Additionally, urea was identified as a degradation product of NTO.

Longitudinal Study of Caries Development from Childhood to Adolescence.

The World Health Organization (WHO) stated that globally, dental caries is the most important oral condition. To develop effective prevention strategies requires an understanding of how this condition develops and progresses over time, but there are few longitudinal studies of caries onset and progression in children. The aim of the study was to establish the pattern of caries development from childhood into adolescence and to explore the role of potential risk factors (age, sex, ethnicity, and social deprivation). Of particular interest was the disease trajectory of dentinal caries in the permanent teeth in groups defined by the presence or absence of dentinal caries in the primary teeth. Intraoral examinations to assess oral health were performed at 4 time points by trained and calibrated dentist examiners using a standardized, national diagnostic protocol. Clinical data were available from 6,651 children. Mean caries prevalence (% D3MFT > 0) was 16.7% at the first clinical examination (ages 7-9 y), increasing to 31.0%, 42.2%, and 45.7% at subsequent examinations. A population-averaged model (generalized estimating equations) was used to model the longitudinal data. Estimated mean values indicated a rising D3MFT count as pupils aged (consistent with new teeth emerging), which was significantly higher (4.49 times; 95% confidence interval, 3.90-5.16) in those pupils with caries in their primary dentition than in those without. This study is one of the few large longitudinal studies to report the development of dental caries from childhood into adolescence. Children who developed caries in their primary dentition had a very different caries trajectory in their permanent dentition compared to their caries-free contemporaries. In light of these results, caries-free and caries-active children should be considered as 2 separate populations, suggesting different prevention strategies are required to address their different risk profiles.

Activity of 2-fluoro-5-methylarabinofuranosyluracil against mouse leukemias sensitive to and resistant to 1-beta-D-arabinofuranosylcytosine.

A new pyrimidine nucleoside, 2'-fluoro-5-methyl-1-beta-D-arabinofuranosyluracil, previously has been shown to be active against the herpes group of viruses in vitro and in vivo. It is also active against mouse and human leukemic cells in culture and against mouse leukemias L1210, P388, and P815 in vivo. In contrast to other 1-beta-D-arabinofuranosylcytosine (ara-C) derivatives, 2'-fluoro-5-methyl-1-beta-D-arabinofuranosyluracil, when given either i.p. or p.o., is highly active against lines of leukemias P815 and L1210 made resistant to ara-C. Against P815/ara-C and L1210/araC, it is more effective than is 5-azacytidine, a drug which has shown definite effectiveness in patients with acute leukemia whose disease has become resistant to ara-C. For these reasons, 2'-fluoro-5-methyl-1-beta-D-arabinofuranosyluracil would seem to merit clinical trial in patients with acute nonlymphocytic leukemia whose disease has become resistant to ara-C.

Facial aesthetic outcome analysis in unilateral cleft lip and palate surgery using web-based extended panel assessment.

BACKGROUND AND AIM: The reproducible measurement of aesthetic outcomes after cleft lip and palate (CLP) surgery remains elusive, and there is no internationally recognised system. The aim of this pilot study was to better understand how humans rate post-operative aesthetic outcome after unilateral cleft lip and palate (UCLP) repair using a novel web-based rating platform with an extended panel of surgeon raters. METHODS: Cropped images of 5-year-old UCLP patients were arranged in a randomly generated sequence within a web-based aesthetic scoring tool as part of an agreement/reliability study. Assessors rated the appearances of patients using a five-point Likert-type scale on two occasions. A mixed-effect statistical model was adopted to analyse the effects of rater, image and timing. RESULTS: Images of 76 patients were scored by 29 UK-based cleft surgeons. Intra-rater variability was found, and the linear weighted kappa was 0.56. This allowed identification of the most and least consistent raters. The random image effect (p < 0.001) suggested that a broad range of aesthetic outcomes were included in the current study. Surgeon raters in this study were likely to score the images more preferably at the second assessment. CONCLUSIONS: A web-based scoring system provides extended data capture, and mixed-effect statistical modelling reveals the effect that time, image and rater have on the scorings. The selection and training of raters, in combination with an exemplary yardstick, might improve inter- and intra-rater agreement. The development of objective measures based upon digital facial recognition can replace the highly variable subjective human influence on rating the aesthetic outcome.

Synthesis of the acridone alkaloids glyfoline and congeners. Structure-activity relationship studies of cytotoxic acridones.

Glyfoline (4, 1,6-dihydroxy-10-methyl-2,3,4,5-tetramethoxyacridin-9-one) and its congeners were synthesized for evaluation of their cytotoxicity. A detailed structure-activity relationships (SAR) of these acridone derivatives were also studied. To study the SAR of glyfoline analogues, substituent(s) at C-1 and C-6 and at the heterocyclic nitrogen of glyfoline nucleus were modified. Nitro- and amino-substituted glyfoline analogues were also synthesized to study the effects of substituent(s) (electron-withdrawing vs electron-donating) on their cytotoxicity. These compounds were synthesized via the Ullmann condensation of anthranilic acids with iodobenzenes or 2-chlorobenzoic acids with aniline derivatives. The SAR studies showed that 1-hydroxy-9-acridones were more active than their 1-OMe derivatives against cell growth of human leukemic HL-60 cells in culture. Replacement of NMe of glyfoline with NH or N(CH2)2NEt2 resulted in either total loss or dramatic reduction of cytotoxicity. Glyfoline congeners with nitro function at the A-ring were inactive, while compounds with amino substituent were shown to be cytotoxic in vitro.

Nucleosides. 136. Synthesis and antiviral effects of several 1-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)-5-alkyluracils. Some structure-activity relationships.

In order to study structure-activity relationships between antiherpetic activity and the size of the C-5 alkyl substituents of 2'-fluoro-ara-U derivatives, six new nucleosides (1c-h) were synthesized. The 5-allyl analogue 1c was prepared by a Pd(II)-catalyzed reaction of 5-(chloromercuri)-1-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)uracil with allyl chloride. Partial hydrogenation of 1c afforded the 5-n-propyl derivative 1d (FPAU). Nucleosides 1e-h were obtained by condensation of 3-O-acetyl-5-O-benzoyl-2-deoxy-2-fluoro-D-arabinosyl bromide with the corresponding 5-substituted uracils. Preliminary in vitro data show that, as the alkyl side chain is increased by one carbon unit, the antiherpetic potency is decreased by approximately 1 log order. The cytotoxicity also diminishes as the size of the 5-substituent is increased. FPAU exerts good activity against HSV-1 and HSV-2. FiPAU still shows good therapeutic indices, whereas the higher alkyl analogues are essentially inactive.

Nucleosides. 129. Synthesis of antiviral nucleosides: 5-alkenyl-1-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)uracils.

Synthesis of 1-(2-deoxy-2-fluoro-beta-D-arabinofuranosyl)uracils containing a vinyl (4a), 2-halovinyl (4b-d), or ethyl substituent at C-5 was achieved. These nucleosides were found to be about a log order less active than 2'-fluoro-5-iodo-ara-C (FIAC) against HSV-1, but they are much less cytotoxic against normal human lymphocytes than FIAC. Nucleosides 4a and 4e showed good activity against HSV-1 (ED50 = 0.16 and 0.24 microM, respectively) and HSV-2 (ED50 = 0.69 and 0.65 microM) with very little cytotoxicity (ID50 greater than 100 microM).

Synthesis and biological activities of 5-deaza analogues of aminopterin and folic acid.

N-[p-[[(2,4-Diaminopyrido[2,3-d]pyrimidin-6-yl)methyl] amino]benzoyl]-L-glutamic acid (1a, 5-deazaaminopterin) and the 5-methyl analogue (1b) were synthesized in 14 steps from 5-cyanouracil (4a) and 5-cyano-6-methyluracil (4b), respectively, by exploitation of the novel pyrimidine to pyrido[2,3-d]pyrimidine ring transformation reaction. The 5-cyanouracils 4 were treated with chloromethyl methyl ether to the 1,3-bis(methoxymethyl)uracils (5, which were treated with malononitrile in NaOEt/EtOH to give the pyrido[2,3-d]pyrimidines 6. Diazotization of 6 in concentrated HCl afforded the 7-chloro derivatives 8 in high yield. After reduction of 8, the 7-unsubstituted products 9 were reduced in the presence of Ac2O and the products, 6-(acetamidomethyl)pyridopyrimidines 10, were converted into the 6-acetoxymethyl derivatives 12 via nitrosation. After removal of the N-methoxymethyl groups from 12, the 6-(acetoxymethyl)pyrido[2,3-d]pyrimidine-2,4(1H,3H)-diones 14 were converted into 2,4-diamino-6-(hydroxymethyl)pyrido[2,3-d]pyrimidine (15a) and its 5-methyl analogue 15b by the silylation-amination procedure. Compounds 15 were brominated to the 6-bromomethyl derivatives 16, which were treated with diethyl (p-aminobenzoyl)-L-glutamate, and the products 17 were saponified to afford 5-deazaaminopterin (1a) and its 5-methyl analogue 1b. Compound 1b was also prepared by an alternative procedure in 10 steps from cyanothioacetamide and ethyl beta-(ethoxymethylene)acetoacetate via 2,4-diamino-6-(hydroxymethyl)-5-methylpyrido[2,3-d]pyrimidine (15b). 5-Deaza-5-methylfolic acid (2) was also prepared in four steps from 15b. The aminopterine analogues 1 showed significant anticancer activity in vitro and in vivo, whereas the folic acid analogue 2 did not exhibit any significant toxicity.

Chemical synthesis and biological activities of 5-deazaaminopterin analogues bearing substituent(s) at the 5- and/or 7-position(s).

Condensation of cyanothioacetamide (4) with ethyl alpha-(ethoxymethylene)acetoacetate (5b), ethyl 4-ethoxy-2-(ethoxymethylene)-3-oxobutanoate (5c), ethyl 2-(ethoxymethylene)-3-oxo-4-phenylpropanoate (5d) afforded exclusively the corresponding 6-substituted pyridines (6b-d). Cyclization of 4 with 3-carbethoxybutane-2,4-dione (5e) gave 3-cyano-5-(ethoxycarbonyl)-4,6-dimethylpyridine-2(1H)-thione (6e), whereas reaction of 4 with 3-carbethoxy-1-phenylpropane-1,3-dione (5f) yielded two products, 3-cyano-5-(ethoxycarbonyl)-4-methyl-6-phenylpyridine-2(1H)-thione (6f) and the 6-methyl-4-phenyl isomer 6g. The structural assignments for 6f and 6g are made on the basis of 1H and 13C NMR spectral analyses of the 2-(methylthio)nicotinates (7f,g) prepared from 6f and 6g by treatment with MeI/K2CO3. Nicotinates 7b,d-g were converted into their corresponding 2,4-diaminopyrido[2,3-d]pyrimidines 12b,d-g in five steps, via reduction, protection, oxidation, condensation with guanidine, and deprotection. The 7-mono- and 5,7-disubstituted-5-deazaaminopterins (1b,d-g) were prepared from the respective pyrido[2,3-d]pyrimidines 12b,d-g. Preliminary biological studies showed that 7-methyl and 5,7-dimethyl analogues (1b and 1e) were less active than methotrexate against human leukemic HL-60 and murine L-1210 cells in tissue culture. Compound 1e produced an ILS of 71% at 100 mg/kg per day X 5 (ip) in BDF mice inoculated ip with 10(6) L-1210 cells.

Synthesis and structure-activity relationships of potential anticancer agents: alkylcarbamates of 3-(9-acridinylamino)-5-hydroxymethylaniline.

A series of potential 9-anilinoacridine antitumor agents, 3-(9-acridinylamino)-5-hydroxymethylaniline (AHMA) derivatives with monosubstituent at C4' and disubstituents at C4' and C5' of the acridine ring and their alkylcarbamates, were synthesized for evaluation of their antitumor activity. A structure-activity relationship (SAR) study showed that the AHMA-alkylcarbamates were more potent than their corresponding parent AHMA compounds. In addition, the cytotoxicity of the AHMA-alkylcarbamate decreased with increasing length and size of the alkyl function. Among these compounds, AHMA-ethylcarbamate (18) and 4'-methyl-5'-dimethylaminoethylcarboxamido-AHMA-ethylcarb amate (34) possess potent cytotoxicity on the inhibition of human leukemic HL-60 cell growth in culture. Further in vivo studies of these compounds displayed significant anticancer therapeutic effects in mice bearing sarcoma 180, Lewis lung carcinoma, and P388 leukemia.

Cyclopent[a]anthraquinones as DNA intercalating agents with covalent bond formation potential: synthesis and biological activity.

A series of mitomycin C (MMC) analogues, namely cyclopentanthraquinone derivatives, were synthesized via Diels-Alder cyclization of naphthoquinone with 1-vinylcyclopent-1-enes. These new compounds are planar structures, like MMC, and bear an aziridine ring and a methyl carbamate side chain. After bioreduction, they are anticipated to be capable of intercalating into double-stranded DNA and bind covalently. Structure-activity relationships were studied. Of these compounds, 2,3-aziridino-4-[[(methylamino)carbonyl]methyl] cyclopent[alpha]anthracene-6,11-dione (4) was shown to have inhibitory activity against several leukemic and solid tumor cell lines. Mice (BDF1) bearing Lewis lung adenocarcinoma were treated with 4 and MMC (i.p., QD x 5). At a dose of 30.0 mg/kg, compound 4 was as effective as MMC (0.8 mg/kg). Compound 4 appears to be less toxic than MMC. DNA unwinding assay indicated that 4 is able to intercalate into DNA double strands and is also a topoisomerase II inhibitor.

Nucleosides. 123. Synthesis of antiviral nucleosides: 5-substituted 1-(2-deoxy-2-halogeno-beta-D-arabinofuranosyl)cytosines and -uracils. Some structure-activity relationships.

The syntheses of several 2'-halogeno-5-substituted-arabinofuranosylcytosines and -uracils are described, and relationships of structure to anti herpes virus activity in vitro were examined. Those arabinonucleosides containing the 2'-fluoro function exhibit, generally, more potent anti herpes virus (HSV) activity than do their 2'-chloro of 2'-bromo analogues. The importance of the fluorine in the 2'-"up" (arabino) configuration for enhancement of antiviral effectiveness is demonstrated by the superior activity of 2'-fluoro-5-iodo-ara-C [3a, FIAC] to that of 2'-fluoro-5-iodo-ribo-C. Of all the nucleosides tested herein, FIAC exhibited the most potent in vitro activity against HSV. 2'-Chloro-5-iodo- and -5-methyl-ara-C (3b and 4b) were 37 to greater than 500 times more effective in vitro against HSV type 2 than against type 1, suggesting that these latter derivatives might serve clinically as useful probes to distinguish between HSV types 1 and 2 in the diagnosis of HSV infections in man.

9-substituted acridine derivatives with long half-life and potent antitumor activity: synthesis and structure-activity relationships.

A series of DNA-intercalating 9-anilinoacridines, namely 9-phenoxyacridines, 9-(phenylthio)acridines, and 9-(3',5'-disubstituted anilino)acridines, were synthesized as potential antitumor agents with inhibitory effects on DNA topoisomerase II. Unlike amsacrine (m-AMSA), these agents were designed to avoid the oxidative metabolic pathway. These acridine derivatives were, therefore, expected to have long half-life in plasma. Both 9-phenoxyacridines and 9-(phenylthio)acridines were found to have moderate cytotoxicity against mouse leukemia L1210 and human leukemic HL-60 cell growth in culture. Among 9-(3',5'-disubstituted anilino)acridines, 3-(9-acridinylamino)-5-(hydroxymethyl)aniline (AHMA) was found to be a potent topoisomerase II inhibitor and exhibited significant antitumor efficacy both in vitro and in vivo. Chemotherapy of solid-tumor-bearing mice with 10, 10, and 5 mg/kg (QD x 4, ip) AHMA, VP-16, and m-AMSA, respectively, resulted in more tumor volume reduction by AHMA than by VP-16 or m-AMSA for E0771 mammary adenocarcinoma and B-16 melanoma. For Lewis lung carcinoma, AHMA was as potent as VP-16 but more active than m-AMSA. Structure-activity relationships of AHMA derivatives are discussed.

Serum hepatocyte growth factor level associate with gastric cancer progression.

We have measured serum HGF levels from 80 gastric cancer patients and 51 normal subjects by enzyme-linked immunosorbent assay. The results showed that the mean value of serum HGF level in gastric cancer patients was significantly higher than in normal subjects (0.30 +/- 0.02 vs 0.22 +/- 0.05 ng/ml; p = 0.005). The increase was stage related. Patients with serum HGF < or = 0.30 ng/ml survived longer than those with serum HGF > 0.30 ng/ml (p = 0.02). These data suggest that HGF involve in progression of gastric cancer.

Differential paclitaxel-induced cytotoxicity in rodent and human hepatoma cell lines.

Hepatoma is the leading cause of death in male cancer patients in Taiwan. In this study, we examined the effect of Paclitaxel on the in vitro growth of 2 rodent and 4 human hepatoma cell lines. Differential Paclitaxel-induced cytotoxicity was observed among hepatoma cell lines. In Paclitaxel-sensitive Hep3B and N1S1 cells, Paclitaxel-induced cytotoxicity was dose- and time-dependent. The effective doses of Paclitaxel were in the range 0.1-1.0 microM. Flow cytometric analysis showed that Paclitaxel-treated hepatoma cells were arrested in G2-M phases prior to apoptosis. In addition, growth inhibition by Paclitaxel was accompanied by an increase in the expression of proliferating cell nuclear antigen (PCNA) in hepatoma cells. For Paclitaxel-resistant hepatoma cells, cytostatic response and/or polyploidization was observed. Our results indicated that two thirds of the hepatoma cell lines examined showed some degree of resistance to Paclitaxel treatment in vitro. The expression of p53 gene had no direct effect on Paclitaxel-induced cytotoxicity. The expression of PCNA and the development of polyploidization appear to be good markers for measuring Paclitaxel response. These findings suggest that Paclitaxel alone appears to by cytostatic to hepatoma cells, combination of Paclitaxel with other chemotherapeutic agents may show better cytotoxic effects.

Anti-fertility and other actions of gossypol analogues.

From a series of gossypol derivatives studied, we conclude that the carbonyl groups of gossypol are needed for inhibition of erythrocyte anion transport and the hydroxy groups affect but are not essential to that inhibition. In an in vitro mouse erythroleukemia cytocidal assay, the most active compounds were gossypol and apogossypol. The latter was not active in the inhibition of erythrocyte anion transport or in a spermicidal assay. Of the more simple structures related to gossypol, those that were active in the cytocidal and spermicidal assays were bi-aromatic, linked by a 1- and not a 4-carbon chain and had free phenolic hydroxyl groups. These results are included in a discussion of the specificity and mechanism of action of gossypol.

PIP: The analysis of multiple biological assays of gossypol and its derivatives suggests that the carbonyl groups of gossypol are required for inhibition of erythrocyte anion transport and the hydroxy groups affect but are not essential to that inhibition. In an in vitro mouse erythroleukemia cytocidal assay, the most active compounds were gossypol and apogossypol. The latter was not active in the inhibition of erythrocyte anion transport or in a spermicidal assay. Of the more simple structures related to gossypol, those that were active in the cytocidal and spermicidal assays were biaromatic, linked by a 1- and not a 4-carbon chain, and had free phenolic hydroxyl groups. When gossypol inhibits the anion transporter, the carbonyl group does not seem to form a Schiff base. Gossypol is a unique compound since it alone, but not any of its derivatives, has in vivo as well as in vitro antifertility activity. It remains unknown, however, whether similar mechanisms are involved in gossypol's in vivo and in vitro effects. In whatever manner gossypol exerts its toxic effects, the selectivity for testicular tissue must be explained.

Kinetics of the alkaline hydrolysis of nitrocellulose.

Cellulose nitrate (nitrocellulose) is an explosive solid substance used in large quantities in various formulations of rocket and gun propellants. Safe destruction of nitrocellulose can be achieved by alkaline hydrolysis, which converts it to biodegradable products that can then be treated by conventional biological processes. The kinetics of the alkaline hydrolysis of munitions-grade nitrocellulose in sodium hydroxide solutions were investigated in completely mixed batch reactors. Experiments were conducted using solutions of alkaline strength ranging from 0.1 to 15% by mass and temperatures in the range of 30 to 90 degrees C. Regression analysis of the kinetic data revealed that alkaline hydrolysis of nitrocellulose is of the order 1.0 and 1.5 with respect to nitrocellulose and hydroxide concentration, respectively. The activation energy of the hydrolysis reaction was found to be 100.9 kJ/mol with a preexponential Arrhenius constant of 4.73 x 10(13). Nitrite and nitrate, in a 3:1 ratio, were the primary nitrogen species present in the posthydrolysis solution. The kinetic information is pertinent to the development and optimization of nitrocellulose chemical-biological treatment systems.

Multidrug-resistant cells overexpressing P-glycoprotein are susceptible to DNA crosslinking agents due to attenuated Src/nuclear EGFR cascade-activated DNA repair activity.

We synthesized several novel bifunctional alkylating derivatives of 3a-aza-cyclopenta[a]indene (BO-1012, BO-1005, BO-1099 and BO-1101) that are potent DNA interstrand crosslinking agents. In in vitro cytotoxicity assay, these compounds were more cytotoxic to multidrug-resistant (MDR) cells, such as KBvin10, KBtax50 and CEM/VBL, than their parental cells. Using a xenograft model, BO-1012, at a dose of 5 mg/kg, partially suppressed the growth of parental KB cells but completely suppressed the growth of KBvin10 cells in nude mice. In exploring the possible mechanism, we found that DNA double-strand break (DSB) repair activity in MDR cells, KBvin10 and CEM/VBL, was significantly reduced compared with their parental cells, KB and CEM. Reduced DSB repair activity in KBvin10 cells was likely due to a defect in nuclear translocation of DNA-dependent protein kinase (DNA-PK), a component of the non-homologous end-joining repair machinery. Furthermore, BO-1012-induced DNA-PK translocation from the cytosol into the nucleus in KB cells is associated with the activation of the Src/nuclear epidermal growth factor receptor (EGFR) cascade, which is defective in MDR cells. As knockdown of P-glycoprotein (P-gp) by siRNA reactivated the Src/nuclear EGFR cascade, DNA-PK translocation and DNA repair activity in MDR cells, overexpression of P-gp attenuates the activity of DNA DSB repair through suppression of Src/nuclear EGFR cascade. Therefore, DNA interstrand crosslinking agents may have potential therapeutic use against P-gp-overexpressing MDR cells.