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1.
ACS Appl Mater Interfaces ; 12(11): 12609-12617, 2020 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-32073826

RESUMO

Glioblastoma multiforme (GBM) is a grade IV astrocytoma, which is the most aggressive form of brain tumor. The standard of care for this disease includes surgery, radiotherapy and temozolomide (TMZ) chemotherapy. Poor accumulation of TMZ at the tumor site, tumor resistance to drug, and dose-limiting bone marrow toxicity eventually reduce the success of this treatment. Herein, we have encapsulated >500 drug molecules of TMZ into the biocompatible protein nanocage, apoferritin (AFt), using a "nanoreactor" method (AFt-TMZ). AFt is internalized by transferrin receptor 1-mediated endocytosis and is therefore able to facilitate cancer cell uptake and enhance drug efficacy. Following encapsulation, the protein cage retained its morphological integrity and surface charge; hence, its cellular recognition and uptake are not affected by the presence of this cargo. Additional benefits of AFt include maintenance of TMZ stability at pH 5.5 and drug release under acidic pH conditions, encountered in lysosomal compartments. MTT assays revealed that the encapsulated agents displayed significantly increased antitumor activity in U373V (vector control) and, remarkably, the isogenic U373M (MGMT expressing TMZ-resistant) GBM cell lines, with GI50 values <1.5 µM for AFt-TMZ, compared to 35 and 376 µM for unencapsulated TMZ against U373V and U373M, respectively. The enhanced potency of AFt-TMZ was further substantiated by clonogenic assays. Potentiated G2/M cell cycle arrest following exposure of cells to AFt-TMZ indicated an enhanced DNA damage burden. Indeed, increased O6-methylguanine (O6-MeG) adducts in cells exposed to AFt-TMZ and subsequent generation of γH2AX foci support the hypothesis that AFt significantly enhances the delivery of TMZ to cancer cells in vitro, overwhelming the direct O6-MeG repair conferred by MGMT. We have additionally encapsulated >500 molecules of the N3-propargyl imidazotetrazine analog (N3P), developed to combat TMZ resistance, and demonstrated significantly enhanced activity of AFt-N3P against GBM and colorectal carcinoma cell lines. These studies support the use of AFt as a promising nanodelivery system for targeted delivery, lysosomal drug release, and enhanced imidazotetrazine potency for treatment of GBM and wider-spectrum malignancies.

2.
J Cell Mol Med ; 24(2): 1750-1759, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31876059

RESUMO

Stomach cancer is the fourth most common cancer worldwide. Identification of novel molecular therapeutic targets and development of novel treatments are critical. Against a panel of gastric carcinoma cell lines, the activity of 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203) was investigated. Adopting RT-PCR, Western blot and immunohistochemical techniques, we sought to determine molecular pharmacodynamic (PD) markers of sensitivity and investigate arylhydrocarbon (AhR) receptor-mediated signal transduction activation by 5F 203. Potent (IC50  ≤ 0.09 µmol/L), selective (>250-fold) in vitro antitumour activity was observed in MKN-45 and AGS carcinoma cells. Exposure of MKN-45 cells to 5F 203 triggered cytosolic AhR translocation to nuclei, inducing CYP1A1 (>50-fold) and CYP2W1 (~20-fold) transcription and protein (CYP1A1 and CYP2W1) expression. G2/M arrest and γH2AX expression preceded apoptosis, evidenced by PARP cleavage. In vivo, significant (P < .01) 5F 203 efficacy was observed against MKN-45 and AGS xenografts. In mice-bearing 5F 203-sensitive MKN-45 and 5F 203-insensitive BGC-823 tumours in opposite flanks, CYP1A1, CYP2W1 and γH2AX protein in MKN-45 tumours only following treatment of mice with 5F 203 (5 mg/kg) revealed PD biomarkers of sensitivity. 5F 203 evokes potent, selective antitumour activity in vitro and in vivo in human gastric cancer models. It triggers AhR signal transduction, CYP-catalysed bioactivation to electrophilic species causing lethal DNA double-strand breaks exclusively in sensitive cells. 5F 203 represents a novel therapeutic agent with a mechanism of action distinct from current clinical drugs, exploiting novel molecular targets pertinent to gastric tumourigenesis: AhR, CYP1A1 and CYP2W1. PD markers of 5F 203 sensitivity that could guide patient selection have been identified.

3.
Front Oncol ; 9: 485, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31263673

RESUMO

Temozolomide (TMZ) is the standard of care chemotherapeutic agent used in the treatment of glioblastoma multiforme. Cytotoxic O6-methylguaine lesions formed by TMZ are repaired by O6-methyl-guanine DNA methyltransferase (MGMT), a DNA repair protein that removes alkyl groups located at the O6-position of guanine. Response to TMZ requires low MGMT expression and functional mismatch repair. Resistance to TMZ conferred by MGMT, and tolerance to O6-methylguanine lesions conferred by deficient MMR severely limit TMZ clinical applications. Therefore, development of new TMZ derivatives that can overcome TMZ-resistance is urgent. In this study, we investigated the anti-tumor mechanism of action of two novel TMZ analogs: C8-imidazolyl (377) and C8-methylimidazole (465) tetrazines. We found that analogs 377 and 465 display good anticancer activity against MGMT-overexpressing glioma T98G and MMR deficient colorectal carcinoma HCT116 cell lines with IC50 value of 62.50, 44.23, 33.09, and 25.37 µM, respectively. Analogs induce cell cycle arrest at G2/M, DNA double strand break damage and apoptosis irrespective of MGMT and MMR status. It was established that analog 377, similar to TMZ, is able to ring-open and hydrolyze under physiological conditions, and its intermediate product is more stable than MTIC. Moreover, DNA adducts of 377 with calf thymus DNA were identified: N7-methylguanine, O6-methylguanine, N3-methyladenine, N3-methylthymine, and N3-methylcytidine deoxynucleotides. We conclude that C8 analogs of TMZ share a mechanism of action similar to TMZ and are able to methylate DNA generating O6-methylguanine adducts, but unlike TMZ are able at least in part to thwart MGMT- and MMR-mediated resistance.

4.
Medchemcomm ; 9(3): 545-553, 2018 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-30108945

RESUMO

A series of 3-(benzyl-substituted)-imidazo[5,1-d]-1,2,3,5-tetrazines (13) and related derivatives with 3-heteromethyl groups has been synthesised and screened for growth-inhibitory activity in vitro against two pairs of glioma cell lines with temozolomide-sensitive and -resistant phenotypes dependent on the absence/presence of the DNA repair protein O6-methylguanine-DNA methyltransferase (MGMT). In general the compounds had low inhibitory activity with GI50 values >50 µM against both sets of cell lines. Two silicon-containing derivatives, the TMS-methylimidazotetrazine (9) and the SEM-analogue (10), showed interesting differences: compound (9) had a profile very similar to that of temozolomide with the MGMT+ cell lines being 5 to 10-fold more resistant than MGMT- isogenic partners; the SEM-substituted compound (10) showed potency across all cell lines irrespective of their MGMT status.

5.
J Cell Biochem ; 119(7): 5350-5358, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29331023

RESUMO

The efficacy of temozolomide (TMZ) treatment for cancers is currently limited by inherent or the development of resistance, particularly, but not exclusively, due to the expression of the DNA repair enzyme O6-methylguanine-DNA methyltransferase (MGMT) in a significant proportion of tumors. We have found that TMZ analog C8-methyl imidazole tetrazine (PMX 465) displayed good anticancer activity against the colorectal carcinoma HCT116 cells which are MGMT-overexpressing and mismatch repair (MMR)-deficient. In this study, we found that PMX 465 could downregulate the expression of MGMT in HCT116 cells at the protein and mRNA levels. We found that PMX 465 could reduce MGMT expression by increasing the binding of wild-type p53 to the MGMT promoter and reducing the binding of Sp1 to the MGMT promoter.


Assuntos
Antineoplásicos Alquilantes/farmacologia , Neoplasias Colorretais/metabolismo , Metilases de Modificação do DNA/metabolismo , Enzimas Reparadoras do DNA/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Temozolomida/análogos & derivados , Temozolomida/farmacologia , Proteínas Supressoras de Tumor/metabolismo , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Metilases de Modificação do DNA/genética , Enzimas Reparadoras do DNA/genética , Regulação para Baixo , Células HCT116 , Humanos , Regiões Promotoras Genéticas , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/genética
6.
Org Biomol Chem ; 14(16): 3889-905, 2016 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-27035072

RESUMO

The imidazole ring is widespread in biologically active compounds, and hence imidazole-containing scaffolds are useful starting points for drug discovery programmes. We report the synthesis of a series of novel imidazole-containing compounds fused with either phenanthrene or phenanthroline, which show enhanced growth inhibitory potency against human colon, breast and melanoma cancer cell lines, as well as evidence of inhibition of the molecular chaperone heat shock protein 70 (Hsp70) pathway in cells, as shown by depletion of downstream oncogenic client proteins of the Hsp90 chaperone pathway, and induction of apoptosis.


Assuntos
Apoptose/efeitos dos fármacos , Proteínas de Choque Térmico HSP70/antagonistas & inibidores , Imidazóis/química , Imidazóis/farmacologia , Fenantrolinas/química , Linhagem Celular Tumoral , Cristalografia por Raios X , Humanos , Fenantrolinas/farmacologia
7.
Bioorg Med Chem ; 23(21): 6891-9, 2015 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-26474663

RESUMO

Potent, selective antitumour AhR ligands 5F 203 and GW 610 are bioactivated by CYPs 1A1 and 2W1. Herein we reason that DNA adducts' generation resulting in lethal DNA double strand breaks (DSBs) underlies benzothiazoles' activity. Treatment of sensitive carcinoma cell lines with GW 610 generated co-eluting DNA adducts (R(2)>0.7). Time-dependent appearance of γ-H2AX foci revealed subsequent DNA double strand breaks. Propensity for systemic toxicity of benzothiazoles steered development of prodrugs' hydrogels for localised delivery. Clinical applications of targeted therapies include prevention or treatment of recurrent disease after surgical resection of solid tumours. In vitro evaluation of 5F 203 prodrugs' activity demonstrated nanomolar potency against MCF-7 breast and IGROV-1 ovarian carcinoma cell lines.


Assuntos
Antineoplásicos/síntese química , Adutos de DNA/análise , Hidrogéis/química , Pró-Fármacos/síntese química , Tiazóis/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Benzotiazóis/síntese química , Benzotiazóis/química , Benzotiazóis/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Adutos de DNA/metabolismo , Resistencia a Medicamentos Antineoplásicos , Regulação da Expressão Gênica/efeitos dos fármacos , Histonas/metabolismo , Humanos , Microscopia Confocal , Pró-Fármacos/química , Pró-Fármacos/farmacologia , Resveratrol , Estilbenos/química , Tiazóis/síntese química , Tiazóis/farmacologia
8.
Oncology ; 88(1): 28-48, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25277441

RESUMO

Glioblastoma multiforme (GBM) treatment includes temozolomide (TMZ) chemotherapy. O6-Methylguanine lesions are repaired by methylguanine-DNA methyltransferase (MGMT). Response to TMZ requires low MGMT and functional mismatch repair (MMR); resistance, conferred by MGMT or MMR deficiency, represents a barrier to successful treatment. TMZ analogs were synthesized, substituting N3-methyl with propargyl (1) or sulfoxide (2). MTT assays were conducted in SNB19 and U373 isogenic glioma cell lines (V = vector control; M = MGMT-transfected). TMZ potency was reduced >5-fold in SNB19M and U373M cells; in contrast, MGMT-expressing cells were equisensitive as vector controls to analogs 1 and 2 . GI50 values <50 µM of analogs 1 or 2 were detected in V cells possessing acquired TMZ resistance: SNB19VR (hMSH6 loss) and U373VR (MGMT upregulation). Analogs 1 and 2 inhibited MMR-deficient colorectal carcinoma cell growth (irrespective of p53); G2/M cell cycle arrest preceded apoptosis. γH2AX foci inferred the generation of DNA double-strand breaks by analogs 1 and 2 . Acridine orange-stained vesicles, intracellular punctate GFP-LC3 protein and double-membraned autophagosomes indicate that TMZ, 1 and 2 induce autophagy in apoptotis-resistant GBM cells. Analogs 1 and 2 elicit in vitro antitumor activity irrespective of MGMT, MMR and p53. Such imidazotetrazines may treat MGMT+ GBM and possess broader spectrum activity causing apoptosis and autophagy in malignancies which evade apoptosis.


Assuntos
Antineoplásicos Alquilantes/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Reparo de Erro de Pareamento de DNA/efeitos dos fármacos , Metilases de Modificação do DNA/análise , Enzimas Reparadoras do DNA/análise , Dacarbazina/análogos & derivados , Glioblastoma/tratamento farmacológico , Proteínas Supressoras de Tumor/análise , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Quebras de DNA de Cadeia Dupla , Dacarbazina/farmacologia , Glioblastoma/patologia , Histonas/análise , Humanos , Temozolomida , Proteína Supressora de Tumor p53/análise , Proteína Supressora de Tumor p53/fisiologia
9.
Chem Commun (Camb) ; 50(96): 15202-5, 2014 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-25338751

RESUMO

Biophysical studies of ligand interactions with three human telomeric repeat sequences (d(AGGG(TTAGGG)n, n = 3, 7 and 11)) show that an oxazole-based 'click' ligand, which induces parallel folded quadruplexes, preferentially stabilises longer telomeric repeats providing evidence for selectivity in binding at the interface between tandem quadruplex motifs.


Assuntos
Quadruplex G , Ligantes , Telômero/química , Sequência de Bases , Sítios de Ligação , Dicroísmo Circular , Química Click , Humanos , Simulação de Dinâmica Molecular , Conformação de Ácido Nucleico , Oxazóis/química
10.
PLoS One ; 9(1): e86187, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24454961

RESUMO

BACKGROUND: Telomeric 3' overhangs can fold into a four-stranded DNA structure termed G-quadruplex (G4), a formation which inhibits telomerase. As telomerase activation is crucial for telomere maintenance in most cancer cells, several classes of G4 ligands have been designed to directly disrupt telomeric structure. METHODS: We exposed brain tumor cells to the G4 ligand 3,11-difluoro-6,8,13-trimethyl-8H-quino[4,3,2-kl]acridinium methosulfate (RHPS4) and investigated proliferation, cell cycle dynamics, telomere length, telomerase activity and activated c-Myc levels. RESULTS: Although all cell lines tested were sensitive to RHPS4, PFSK-1 central nervous system primitive neuroectodermal cells, DAOY medulloblastoma cells and U87 glioblastoma cells exhibited up to 30-fold increased sensitivity compared to KNS42 glioblastoma, C6 glioma and Res196 ependymoma cells. An increased proportion of S-phase cells were observed in medulloblastoma and high grade glioma cells whilst CNS PNET cells showed an increased proportion of G1-phase cells. RHPS4-induced phenotypes were concomitant with telomerase inhibition, manifested in a telomere length-independent manner and not associated with activated c-Myc levels. However, anti-proliferative effects were also observed in normal neural/endothelial cells in vitro and ex vivo. CONCLUSION: This study warrants in vivo validation of RHPS4 and alternative G4 ligands as potential anti-cancer agents for brain tumors but highlights the consideration of dose-limiting tissue toxicities.


Assuntos
Acridinas/farmacologia , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Acridinas/química , Animais , Antineoplásicos/química , Neoplasias Encefálicas , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cílios/efeitos dos fármacos , Cílios/fisiologia , Ensaios de Seleção de Medicamentos Antitumorais , Células Endoteliais/efeitos dos fármacos , Epêndima/patologia , Quadruplex G , Humanos , Concentração Inibidora 50 , Proteínas Proto-Oncogênicas c-myc/metabolismo , Ratos , Taq Polimerase/antagonistas & inibidores , Telomerase/antagonistas & inibidores , Telomerase/química , Homeostase do Telômero/efeitos dos fármacos
11.
Bioorg Med Chem ; 20(4): 1607-15, 2012 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-22264753

RESUMO

Quinols have been developed as a class of potential anti-cancer compounds. They are thought to act as double Michael acceptors, forming two covalent bonds to their target protein(s). Quinols have also been shown to have activity against the parasite Trypanosoma brucei, the causative organism of human African trypanosomiasis, but they demonstrated little selectivity over mammalian MRC5 cells in a counter-screen. In this paper, we report screening of further examples of quinols against T. brucei. We were able to derive an SAR, but the compounds demonstrated little selectivity over MRC5 cells. In an approach to increase selectivity, we attached melamine and benzamidine motifs to the quinols, because these moieties are known to be selectively concentrated in the parasite by transporter proteins. In general these transporter motif-containing analogues showed increased selectivity; however they also showed reduced levels of potency against T. brucei.


Assuntos
Sistemas de Liberação de Medicamentos , Hidroquinonas/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma brucei brucei/efeitos dos fármacos , Benzamidinas/síntese química , Benzamidinas/química , Benzamidinas/farmacologia , Linhagem Celular , Humanos , Hidroquinonas/síntese química , Hidroquinonas/química , Concentração Inibidora 50 , Modelos Moleculares , Estrutura Molecular , Triazinas/síntese química , Triazinas/química , Triazinas/farmacologia , Tripanossomicidas/síntese química , Tripanossomicidas/química
12.
Curr Mol Pharmacol ; 5(1): 102-14, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22122467

RESUMO

Glioblastoma multiforme is the most common aggressive adult primary tumour of the central nervous system. Treatment includes surgery, radiotherapy and adjuvant temozolomide (TMZ) chemotherapy. TMZ is an alkylating agent prodrug, delivering a methyl group to purine bases of DNA (O6-guanine; N7-guanine and N3-adenine). The primary cytotoxic lesion, O6-methylguanine (O6-MeG) can be removed by methylguanine methyltransferase (MGMT; direct repair) in tumours expressing this protein, or tolerated in mismatch repair-deficient (MMR-) tumours. Thus MGMT or MMR deficiency confers resistance to TMZ. Inherent- and acquired resistance to TMZ present major obstacles to successful treatment. Strategies devised to thwart resistance and enhance response to TMZ, including inhibition of DNA repair mechanisms which contribute to TMZ resistance, are under clinical evaluation. Depletion of MGMT prior to alkylating agent chemotherapy prevents O6-MeG repair; thus, MGMT pseudosubstrates O6-benzylguanine and lomeguatrib are able to sensitise tumours to TMZ. Disruption of base excision repair (BER) results in persistence of potentially lethal N7- and N3- purine lesions contributing significantly to TMZ cytoxicity particularly when O6-MeG adducts are repaired or tolerated. Several small molecule inhibitors of poly(ADP-ribose)polymerase-1 (PARP-1), a critical BER protein are yielding promising results clinically, both in combination with TMZ and as single agent chemotherapy in patients whose tumours possess homologous recombination DNA repair defects. Another validated, but as yet preclinical protein target, mandatory to BER is abasic (AP) endonuclease-1 (APE-1); in preclinical tests, APE-1 inhibition potentiates TMZ activity. An alternative strategy is synthesis of a molecule, evoking an irrepairable cytotoxic O6-G lesion. Preliminary efforts to achieve this goal are described.


Assuntos
Antineoplásicos Alquilantes/farmacologia , Reparo do DNA/efeitos dos fármacos , Dacarbazina/análogos & derivados , Antineoplásicos Alquilantes/uso terapêutico , Linhagem Celular Tumoral , Neoplasias do Sistema Nervoso Central/tratamento farmacológico , DNA Glicosilases/metabolismo , Metilases de Modificação do DNA/química , Metilases de Modificação do DNA/metabolismo , Enzimas Reparadoras do DNA/química , Enzimas Reparadoras do DNA/metabolismo , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/antagonistas & inibidores , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Dacarbazina/farmacologia , Dacarbazina/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Glioblastoma/tratamento farmacológico , Guanina/análogos & derivados , Guanina/química , Humanos , Inibidores de Poli(ADP-Ribose) Polimerases , Poli(ADP-Ribose) Polimerases/metabolismo , Temozolomida , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/metabolismo
13.
Mol Cancer Ther ; 10(10): 1982-92, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-21831963

RESUMO

Both 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole (5F-203; NSC 703786) and 2-(3,4-dimethoxyphenyl)-5-fluorobenzothiazole (GW-610; NSC 721648) are antitumor agents with novel mechanism(s). Previous studies have indicated that cytochrome (CYP) P450 1A1 is crucial for 5F-203 activity. In the present study, we investigated the functional role of 2 newly identified CYP P450 enzymes, CYP2S1 and CYP2W1, in mediating antitumor activity of benzothiazole compounds. We generated isogenic breast cancer (MDA-MB-468, MCF-7) and colorectal cancer (CRC; KM12 and HCC2998) cell lines depleted for CYP1A1, CYP2S1, or CYP2W1. The sensitivity of these cells to 5F-203 and GW-610 was then compared with vector control cells. 5F-203 exhibited potent activity against breast cancer cells, whereas GW-610 was effective against both breast and colorectal cancer cells. CYP1A1 was induced in both breast cancer and CRC cells, while CYP2S1 and CYP2W1 were selectively induced in breast cancer cells only following treatment with 5F-203 or GW-610. Depletion of CYP1A1 abrogated the sensitivity of breast cancer and CRC cells to 5F-203 and GW-610. Although depletion of CYP2S1 sensitized both breast cancer and CRC cells toward 5F-203 and GW-610, CYP2W1 knockdown caused marked resistance to GW-610 in CRC cells. Our results indicate that CYP-P450 isoforms, with the exception of CYP1A1, play an important role in mediating benzothiazole activity. CYP2S1 appears to be involved in deactivation of benzothiazoles, whereas CYP2W1 is important for bioactivation of GW-610 in CRC cells. Because CYP2W1 is highly expressed in colorectal tumors, GW-610 represents a promising agent for CRC therapy.


Assuntos
Benzotiazóis/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/enzimologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/enzimologia , Sistema Enzimático do Citocromo P-450/metabolismo , Antineoplásicos/farmacologia , Neoplasias da Mama/patologia , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias Colorretais/patologia , Citocromo P-450 CYP1A1/biossíntese , Citocromo P-450 CYP1A1/metabolismo , Sistema Enzimático do Citocromo P-450/biossíntese , Família 2 do Citocromo P450 , Indução Enzimática/efeitos dos fármacos , Feminino , Humanos , Masculino , Tiazóis/farmacologia
14.
Oncology ; 80(3-4): 195-207, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21720182

RESUMO

Resistance to temozolomide (TMZ), conferred by O6-methylguanine-DNA methyltransferase (MGMT) or mismatch repair (MMR) deficiency, presents obstacles to successful glioblastoma multiforme (GBM) treatment. Activities of novel TMZ analogs, designed to overcome resistance, were tested against isogenic SNB19 and U373 GBM cell lines (V = vector control, low MGMT; M = MGMT overexpression). TMZ and triazene MTIC demonstrated >9-fold resistance in SNB19M cells (cf SNB19V). N-3 methyl ester analog 11 and corresponding triazene 12 inhibited growth of TMZ-sensitive (V) and TMZ-resistant (M) cells (GI(50) <50 µM). Ethyl ester 13 and triazene 14 gave similar profiles. MMR-deficient colorectal carcinoma cells, resistant to TMZ (GI(50) >500 µM), responded to analog 11 and 13 treatment. Cross-resistance to these agents was not observed in cell lines possessing acquired TMZ resistance (SNB19VR; U373VR). Methyl ester 11 blocked SNB19V, SNB19M and SNB19VR cells in S and G(2)/M, causing dose- and time-dependent apoptosis. DNA damage, recruiting excision repair was detected by alkaline comet assay; H2AX phosphorylation indicated a lethal DNA double-strand break formation following analog 11 exposure. Compounds 11 and 13 demonstrated 3.7- and 5.1-fold enhanced activity in base excision repair-deficient Chinese hamster ovary cells; furthermore, poly (ADP-ribose) polymerase-1 inhibition potentiated HCT-116 cells' sensitivity to analog 11. In conclusion, analogs 11 and 13 exert anticancer activity irrespective of MGMT and MMR.


Assuntos
Antineoplásicos Alquilantes/farmacologia , Ciclo Celular/efeitos dos fármacos , Reparo de Erro de Pareamento de DNA/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , DNA de Neoplasias/efeitos dos fármacos , Dacarbazina/análogos & derivados , Glioma/genética , Imidazóis/farmacologia , O(6)-Metilguanina-DNA Metiltransferase/genética , Linhagem Celular Tumoral , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/genética , Dacarbazina/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Glioma/tratamento farmacológico , Glioma/enzimologia , Humanos , Projetos Piloto , Poli(ADP-Ribose) Polimerase-1 , Poli(ADP-Ribose) Polimerases/efeitos dos fármacos , Regiões Promotoras Genéticas/efeitos dos fármacos , Temozolomida
15.
Clin Cancer Res ; 17(8): 2227-36, 2011 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-21355072

RESUMO

PURPOSE: We previously reported that the G-quadruplex (G4) ligand RHPS4 potentiates the antitumor activity of camptothecins both in vitro and in tumor xenografts. The present study aims at investigating the mechanisms involved in this specific drug interaction. EXPERIMENTAL DESIGN: Combination index test was used to evaluate the interaction between G4 ligands and standard or novel Topo I inhibitors. Chromatin immunoprecipitation was performed to study the presence at telomeres of various types of topisomerase, while immunolabeling experiments were performed to measure the activation of DNA damage both in vitro and in tumor xenografts. RESULTS: We report that integration of the Topo I inhibitor SN-38, but not the Topo II poison doxorubicin with telomere-based therapy is strongly effective and the sequence of drug administration is critical in determining the synergistic interaction, impairing the cell ability to recover from drug-induced cytotoxicity. The synergistic effect of this combination was also observed by using novel camptothecins and, more interestingly, mice treated with ST1481/RHPS4 combination showed an inhibition and delay of tumor growth as well as an increased survival. The study of the mechanism(s) revealed that treatment with G4 ligands increased Topo I at the telomeres and the functional relevance of this observation was directly assessed by showing that standard and novel camptothecins stabilized DNA damage both in vitro and in xenografts. CONCLUSIONS: Our results demonstrate an outstanding efficacy of Topo I inhibitors/G4 ligands combination, which likely reflects an enhanced and persistent activation of DNA damage response as a critical determinant of the therapeutic improvement.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Dano ao DNA , Ensaios Antitumorais Modelo de Xenoenxerto , Acridinas/administração & dosagem , Acridinas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Camptotecina/administração & dosagem , Camptotecina/análogos & derivados , Camptotecina/farmacologia , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Sinergismo Farmacológico , Quadruplex G/efeitos dos fármacos , Células HCT116 , Células HT29 , Humanos , Irinotecano , Masculino , Camundongos , Camundongos Nus , Telômero/efeitos dos fármacos , Telômero/genética , Telômero/metabolismo , Inibidores da Topoisomerase I/administração & dosagem , Inibidores da Topoisomerase I/farmacologia , Resultado do Tratamento
16.
Org Biomol Chem ; 8(9): 2078-84, 2010 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-20401384

RESUMO

4-Ethynyl-4-hydroxycyclohexa-2,5-dien-1-one 5 undergoes cycloaddition reactions with a range of substituted azides in the presence of copper salts to form 1,4-disubstituted triazoles 8-11 bearing the 4-hydroxycyclohexa-2,5-dien-1-one (quinol) pharmacophore; one example of an isomeric 1,5-disubstituted triazole 12 was formed from 5 and benzyl azide in the presence of a ruthenium catalyst. Compounds were screened for growth-inhibitory activity against five cancer cell lines of colon, breast and lung origin, but were overall less potent than the benzothiazolyl- and indolyl-substituted quinols 2 and 3.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Cobre/química , Cicloexanonas/síntese química , Cicloexanonas/farmacologia , Triazóis/síntese química , Triazóis/farmacologia , Antineoplásicos/química , Catálise , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ciclização , Cicloexanonas/química , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Relação Estrutura-Atividade , Triazóis/química
17.
Oncology ; 78(2): 103-14, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20357518

RESUMO

Treatment for glioblastoma multiforme includes the alkylating agent temozolomide combined with ionizing radiation. Persistent O6-guanine methylation by temozolomide in O6-methylguanine methyl transferase negative tumors causes cytotoxic lesions recognized by DNA mismatch repair, triggering apoptosis. Resistance (intrinsic or acquired) presents obstacles to successful temozolomide treatment, limiting drug efficacy and life expectancy. Two glioma cell lines, SNB19 and U373, sensitive to temozolomide (GI(50) values 36 and 68 microM, respectively) were exposed to increasing temozolomide concentrations (1-100 microM). Variant cell lines (SNB19VR, U373VR) were generated that displayed acquired temozolomide resistance (GI(50) values 280 and 289 microM, respectively). Cross-resistance to mitozolomide was observed in U373VR cells only. In clonogenic and MTT assays, methylguanine methyltransferase (MGMT) depletion using O6-benzylguanine sensitized U373VR cells to temozolomide, indicating the resistance mechanism involves MGMT re-expression. Indeed, Western blot analyses revealed MGMT protein in cell lysates. In SNB19VR cells, down-regulation of MSH6 message and protein expression may confer temozolomide tolerance. Inhibition of poly(ADP-ribose) polymerase-1 (a key base excision repair (BER) enzyme) partially restored sensitivity, and DNA repair gene arrays demonstrated up-regulation (>5-fold) of BER gene NTL1 in SNB19VR cells. In conclusion, we have developed two glioma cell lines whose distinct mechanisms of acquired resistance to temozolomide, involving expression of MGMT, or inactivation of DNA mismatch repair and recruitment of BER enzymes, are consistent with clinical observations. These cell lines provide valuable models for the development of strategies to combat temozolomide resistance.


Assuntos
Antineoplásicos Alquilantes/uso terapêutico , Dacarbazina/análogos & derivados , Resistencia a Medicamentos Antineoplásicos/fisiologia , Glioma/tratamento farmacológico , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Reparo do DNA/efeitos dos fármacos , DNA de Neoplasias/genética , Dacarbazina/uso terapêutico , Relação Dose-Resposta a Droga , Citometria de Fluxo , Glioblastoma/tratamento farmacológico , Glioblastoma/enzimologia , Glioblastoma/genética , Glioma/enzimologia , Glioma/genética , Glioma/patologia , Humanos , Compostos de Mostarda Nitrogenada/uso terapêutico , O(6)-Metilguanina-DNA Metiltransferase/genética , O(6)-Metilguanina-DNA Metiltransferase/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Quinazolinas/farmacologia , Temozolomida , Ensaio Tumoral de Célula-Tronco
18.
J Org Chem ; 74(24): 9372-80, 2009 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-19921793

RESUMO

Carbenes derived from diazoimidazolecarboxylates 4 under thermal or photochemical conditions undergo O-H and N-H insertion reactions with alcohols and amines, respectively, in moderate yield, in competition with reduction in good H-donor solvents. Dichloromethane reacts to give the corresponding 4-chloroimidazole. Aromatic hydrocarbons are excellent traps for the imidazolylidene carbene and lead to a range of arylimidazole derivatives 7. Reaction with pyridine leads to the first example of a pyridinium ylide 8 formed from an imidazolylidene carbene, whereas irradiation in hexafluorobenzene gives the imidazoazocine 11, presumably by way of an initial norcaradiene intermediate.


Assuntos
Compostos Azo/química , Hidrocarbonetos Aromáticos/química , Imidazóis/química , Metano/análogos & derivados , Álcoois/química , Aminas/química , Antineoplásicos/química , Ciclização , Fluorcarbonetos/química , Hidrogênio/química , Metano/química , Nitrogênio/química , Oxigênio/química , Fotoquímica , Piridinas/química , Solventes/química , Temperatura Ambiente
19.
Org Biomol Chem ; 7(20): 4194-200, 2009 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-19795057

RESUMO

We report CD, ESI-MS and molecular modelling studies of ligand binding interactions with DNA quadruplex structures derived from the human telomeric repeat sequence (h-Tel) and the proto-oncogenic c-kit promoter sequence. These sequences form anti-parallel (both 2 + 2 and 3 + 1) and parallel conformations, respectively, and demonstrate distinctively different degrees of structural plasticity in binding ligands. With h-Tel, we show that an extended heteroaromatic 1,4-triazole (TRZ), designed to exploit pi-stacking interactions and groove-specific contacts, shows some selectivity for parallel folds, however, the polycyclic fluorinated acridinium cation (RHPS4), which is a similarly potent telomerase inhibitor, shows selectivity for anti-parallel conformations implicating favourable interactions with lateral and diagonal loops. In contrast, the unique c-kit parallel-stranded quadruplex shows none of the structural plasticity of h-Tel with either ligand. We show by quantitative ESI-MS analysis that both sequences are able to bind a ligand on either end of the quadruplex. In the case of h-Tel the two sites have similar affinities, however, in the case of the c-kit quadruplex the affinities of the two sites are different and ligand-dependent. We demonstrate that two different small molecule architectures result in significant differences in selectivity for parallel and anti-parallel quadruplex structures that may guide quadruplex targeted drug-design.


Assuntos
Acridinas/química , Acridinas/farmacologia , DNA/química , Quadruplex G/efeitos dos fármacos , Triazóis/química , Triazóis/farmacologia , Dicroísmo Circular , DNA/metabolismo , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Humanos , Ligantes , Espectrometria de Massas , Simulação de Dinâmica Molecular , Proteínas Proto-Oncogênicas c-kit/genética , Especificidade por Substrato , Telomerase/antagonistas & inibidores , Telômero/genética
20.
Toxicol Appl Pharmacol ; 237(1): 102-10, 2009 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-19265716

RESUMO

2-(4-Amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203) and related compounds are a series of anti-cancer candidate pharmaceuticals, that have been shown to activate the AhR. We show that these compounds are high-affinity ligands for the rat AhR, but a quantitative assay for their ability to induce CYP1A1 RNA in H4IIEC3 cells, a measure of activation of the AhR, showed a poor relationship between affinity for the AhR and ability to induce CYP1A1 RNA. 5F 203, an agonist with low potency, was able to antagonise the induction of CYP1A1 RNA by TCDD, while IH 445, a potent agonist, did not antagonise the induction of CYP1A1 RNA by TCDD, and Schild analysis confirmed 5F 203 to be a potent antagonist of the induction of CYP1A1 RNA by TCDD in H4IIEC3 cells. In contrast, several benzothiazoles show potent induction of CYP1A1 RNA in human MCF-7 cells, and 5F 203 is unable to detectably antagonise the induction of CYP1A1 RNA in MCF-7 cells, showing a species difference in antagonism. Evaluation of the anti-proliferative activity of benzothiazoles showed that the ability to agonise the AhR correlated with growth inhibition both in H4IIEC3 cells for a variety of benzothiazoles, and between H4IIEC3 and MCF-7 cells for 5F 203, suggesting an important role of agonism of the AhR in the anti-proliferative activity of benzothiazoles.


Assuntos
Antineoplásicos/farmacologia , Benzotiazóis/farmacologia , Citocromo P-450 CYP1A1/efeitos dos fármacos , Receptores de Hidrocarboneto Arílico/agonistas , Tiazóis/farmacologia , Actinas/efeitos dos fármacos , Actinas/genética , Actinas/metabolismo , Análise de Variância , Animais , Linhagem Celular Tumoral , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Indução Enzimática/efeitos dos fármacos , Humanos , Masculino , RNA Mensageiro/análise , Ratos , Ratos Wistar , Receptores de Hidrocarboneto Arílico/metabolismo , Especificidade da Espécie , Estatísticas não Paramétricas
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