DNA repair pathways as targets for cancer therapy

DNA repair pathways can enable tumour cells to survive DNA damage that is induced by chemotherapeutic treatments; therefore, inhibitors of specific DNA repair pathways might prove efficacious when used in combination with DNA-damaging chemotherapeutic drugs. In addition, alterations in DNA repair pathways that arise during tumour development can make some cancer cells reliant on a reduced set of DNA repair pathways for survival. There is evidence that drugs that inhibit one of these pathways in such tumours could prove useful as single-agent therapies, with the potential advantage that this approach could be selective for tumour cells and have fewer side effects.     

Mismatch repair proteins as sensors of alkylation DNA damage

The DNA mismatch repair (MMR) system maintains genome integrity by correcting replication errors. MMR also stimulates checkpoint and cell death responses to DNA damage suggested by the resistance of MMR-defective tumor cells to several chemotherapeutic agents. MMR-dependent cytotoxic response may result from futile repair; however, MMR-mediated apoptosis has been genetically separated from its repair function. In a recent issue of Molecular Cell, Yoshioka and coworkers show that MMR complexes (MutSalpha and MutLalpha) are required for the recruitment of ATR-ATRIP to sites of alkylation damage, demonstrating that MMR complexes can function as sensors in DNA damage signal transduction.     

Translesion DNA replication proteins as molecular targets for cancer prevention

Mutations in DNA are generally considered to have an etiologic role in the development of cancer. If so, it follows that reducing the frequency of such mutations will reduce the incidence of cancer induced by mutagens. Recent advances in elucidating the molecular mechanisms of carcinogen-induced mutagenesis indicate that replication of DNA templates that contain replication-blocking adducts is accomplished with error-prone DNA polymerases. These polymerases have relaxed base-pairing requirements, and can insert bases across from adducted templates, but with potentially mutagenic consequences. In principle, these proteins present new and attractive molecular targets to reduce mutagenesis. If this can be done in vivo without increasing cytotoxic responses to carcinogens, then novel chemopreventive strategies can be designed to reduce the risk of cancer in exposed populations prior to the appearance of disease symptoms.     

DNA and its associated processes as targets for cancer therapy

DNA is the molecular target for many of the drugs that are used in cancer therapeutics, and is viewed as a non-specific target of cytotoxic agents. Although this is true for traditional chemotherapeutics, other agents that were discovered more recently have shown enhanced efficacy. Furthermore, a new generation of agents that target DNA-associated processes are anticipated to be far more specific and effective. How have these agents evolved, and what are their molecular targets?     

DNA polymerases and DNA topoisomerases as targets for the development of anticancer drugs

Studies of a variety of compounds designed as derivatives of prototype active molecules aphidicolin and doxorubicin are reported. So far none of the aphidicolin simpler analogues is as active as the parental molecule. Ten anthracycline analogues, characterized for their cytotoxicity, antitumor activity and inhibition of the relaxing activity of purified human DNA topoisomerase II can be divided into five groups. The majority of the tested compounds shows properties very similar to those of doxorubicin. Epirubicin shows extremely high inhibitory activity toward the relaxing property of topoisomerase II but its antitumor activity and cytotoxicity are similar to those of the former group. The third group includes a compound with extremely high cytotoxicity. The fourth group is represented by a compound which shows a cytotoxicity. The fourth group is represented by a compound which shows a cytotoxicity. The fourth group is represented by a compound which shows a cytotoxicity typical of anthracyclines and good antitumor activity but which has no specific inhibitory activity on topoisomerase II. A fifth group includes a totally inactive compound. Our results suggest that the inhibition of human DNA topoisomerase II is only partially correlated with antitumor activity.     

Beta-oxidized N-nitrosoalkylcarbamates as models for DNA alkylation by N-nitrosobis(2-oxopropyl)amine in Syrian hamsters

A single dose of N-nitrosobis(2-oxopropyl)amine (NDOPA) can selectively induce pancreatic-duct adenocarcinomas in Syrian hamsters. Multiple doses or a higher single dose can induce tumours of the liver and other organs. Our earlier studies employing NDOPA systematically labelled with 14C in the three-carbon chain showed that hamster pancreatic DNA is almost exclusively methylated and that the sole source of the methyl group is the alpha carbon of NDOPA. Hamster liver DNA was equally methylated and alkylated by a three-carbon chain. Current studies using generally labelled tritiated NDOPA with a very high specific activity have shown that the three-carbon alkylation is 2-hydroxypropylation. We have identified two adducts isolated from hamster liver DNA, N7-(2-hydroxypropyl)-guanine and O6-(2-hydroxypropyl)guanine, which contain this group, and we have also isolated and identified N7-methylguanine and O6-methylguanine in DNA from hamster liver and pancreas. beta-Oxidized N-nitrosocarbamates, ethyl N-nitroso-2-oxopropylcarbamate (NOPC) and ethyl N-nitroso-2-hydroxypropylcarbamate (NHPC), are useful models for predicting the DNA adducts observed in vivo following NDOPA treatment. Base-catalysed decomposition of NOPC in the presence of exogenous DNA yields five methylated purines (N3-, N7- and O6-methylguanines and N1- and N3-methyladenines). NHPC, a model for N-nitrosamines containing the 2-hydroxypropyl group, reacts with guanosine to yield N7- and O6-(2-hydroxypropyl)guanines.(ABSTRACT TRUNCATED AT 250 WORDS)     

DNA topoisomerase I and II as targets for rational design of new anticancer drugs

BACKGROUND:: Topoisomerase I and II (topo I and II) are enzymes which alterthe topological state of DNA through DNA strand cleavage, strandpassage and religation. They participate in most aspects ofDNA metabolism and are therefore vital to the cell undergoingdivision. Only one form of topo I has been identified whereastwo isoenzymes of topo II have been described: the     

DNA sequence specificity of antitumor agents. Oncogenes as possible targets for cancer therapy

An examination of the DNA sequence specificity of guanine-N7 alkylation for nitrogen mustards and chlorethylnitrosoureas revealed that large variations in alkylation intensities existed among different guanines in the DNA sequence. The most striking finding was that most agents reacted preferentially at runs of G's, the degree of preference being much greater than would be expected from the number of G's alone. This correlated with the molecular electrostatic potential induced at the guanine-N7 position by the nearest neighbor base pairs. Uracil and quinacrine mustards, however, showed distinctly different reaction patterns from other mustards and a detailed examination has led to structural hypotheses to account for the differences. Certain regions of the genome including regions in some oncogenes and the Epstein-Barr virus have unusually high GC contents (greater than 80% GC) which suggests that the antitumor effectiveness of alkylating agents may in part be due to selective reaction at certain regions in the genome. In fact certain mustards have been shown to exhibit enhanced reactivities with such regions in DNA fragments derived from the c-H-ras oncogene. The above findings point to the possibility of design of alkylating agents to optimise the selectivity of reaction with critical DNA regions. An alternative approach presently under investigation has emerged from an understanding of the characteristics of the sequence specific interaction of the natural oligopeptide antibiotics netropsin and distamycin in the minor groove of DNA. This has led to the synthesis of novel agents (lexitropsins) in which the binding specificity can be shifted from (AT)n in (GC)n in a predictable fashion. Thus the rational design of DNA sequence specific vectors linked to DNA reactive groups, such as alkylating or cleaving agents, could enable DNA damage to be delivered selectively to predetermined critical sites on the genome.     

DNA repair proteins as the targets for paroxetine to induce cytotoxicity in gastric cancer cell AGS

To evaluate the potential anticancer effects of 1175 FDA-approved drugs, cell viability screening was performed using 25 human cancer cell lines covering 14 human cancer types. Here, we focus on the action of paroxetine, which demonstrated greater toxicity toward human gastric adenocarcinoma cell-line AGS cells compared with the other FDA-approved drugs, exhibiting an IC50 value lower than 10 μM. Evaluation of the underlying novel mechanisms revealed that paroxetine can enhance DNA damage in gastric cancer cells and involves downregulation of Rad51, HR23B and ERCC1 expression and function, as well as nucleotide shortage. Enhancement of autophagy counteracted paroxetine-induced apoptosis but did not affect paroxetine-induced DNA damage. Paroxetine also enhanced ROS generation in AGS cells, but a ROS scavenger did not improve paroxetine-mediated DNA damage, apoptosis, or autophagy, suggesting ROS might play a minor role in paroxetine-induced cell toxicity. In contrast, paroxetine did not enhance DNA damage, apoptosis, or autophagy in another insensitive gastric adenocarcinoma cell-line MKN-45 cells. Interestingly, co-administration of paroxetine with conventional anticancer agents sensitized MKN-45 cells to these agents: co-treated cells showed increased apoptosis relative to MKN-45 cells treated with the anticancer agent alone. Unequivocally, these data suggest that for the first time that paroxetine triggers cytotoxicity and DNA damage in AGS cells at least partly by reducing the gene expression of Rad51, HR23B, and ERCC1. Our findings also suggest that paroxetine is a promising candidate anticancer agent and/or chemosensitizing agent for use in combination with other anticancer drugs in cancer therapy. The molecular mechanisms underlying the anticancer activity of co-treatment with paroxetine and chemotherapy appear to be complex and are worthy of further investigation.     

Interrelationships between cellular proliferation,DNA alkylation and age as determinants of ethylnitrosourea-induced neoplasia

Thirty-day-old Sprague-Dawley rats were used to study the persistence of DNA lesions (e.g., O⁶-alkylguanine) induced by various doses of ethylnitrosourea (ENU). Cellular proliferation was measured as an increment of DNA content per organ at 7 days post-treatment. We observed that the persistence of O⁶-EtGua was not affected by the various dose levels. Comparing the 3 organs, the persistence of O⁶-EtGua ranked in the order of brain > kidney > liver, while the percent increase in DNA content was measured as liver > kidney > brain. When the target specificity of ENU carcinogenesis in 30-day-old rats was compared to that following transplacental exposure in terms of its relationship to the persistence of DNA lesions and the rate of tartet cellular proliferation, it permitted the conclusion that induction of neoplasia in target cells is not only determined by persistent DNA lesions but also by the rate of proliferation of target cells at the time of exposure.     

DNA alkylation and interstrand cross-linking by treosulfan

The anti-tumour drug treosulfan (L-threitol 1,4-bismethanesulphonate, Ovastat) is a prodrug for epoxy compounds by converting non-enzymatically to L-diepoxybutane via the corresponding monoepoxide under physiological conditions. The present study supports the hypothesis that this conversion of treosulfan is required for cytotoxicity in vitro. DNA alkylation and interstrand cross-linking of plasmid DNA is observed after treosulfan treatment, but this is again produced via the epoxide species. Alkylation occurs at guanine bases with a sequence selectivity similar to other alkylating agents such as the nitrogen mustards. In treosulfan-treated K562 cells, cross-links form slowly, reaching a peak at approximately 24 h. Incubation of K562 cells with preformed epoxides shows faster and more efficient DNA cross-linking. © 1999 Cancer Research Campaign     

Lysosomes as targets for cancer therapy

Tumor invasion and metastasis are associated with altered lysosomal trafficking and increased expression of the lysosomal proteases termed cathepsins. Emerging experimental evidence suggests that such alterations in lysosomes may form an "Achilles heel" for cancer cells by sensitizing them to death pathways involving lysosomal membrane permeabilization and the release of cathepsins into the cytosol. Here, we highlight recent results on cancer-related changes in the composition and function of lysosomes, focusing on possible implications for the development of novel cancer therapeutics that target tumor cell lysosomes.     

Mitochondria as targets for cancer chemotherapy

Heterogeneity of tumors dictates an individual approach to anticancer treatment. Despite their variability, almost all cancer cells demonstrate enhanced uptake and utilization of glucose, a phenomenon known as the Warburg effect, whereas mitochondrial activity in tumor cells is suppressed. Considering the key role of mitochondria in cell death, it appears that resistance of most tumors towards treatment can be, at least in part, explained by mitochondrial silencing in cancer cells. This review is devoted to the role of mitochondria in cell death, and describes how targeting of mitochondria can make tumor cells more susceptible to anticancer treatment.     

Retinoblastoma family proteins as key targets of the small DNA virus oncoproteins

RB, the most investigated tumor suppressor gene, is the founder of the RB family of growth/tumor suppressors, which comprises also p107 (RBL1) and Rb2/p130 (RBL2). The protein products of these genes, pRb, p107 and pRb2/p130, respectively, are also known as 'pocket proteins', because they share a 'pocket' domain responsible for most of the functional interactions characterizing the activity of this family of cellular factors. The interest in these genes and proteins springs essentially from their ability to regulate negatively cell cycle processes and for their ability to slow down or abrogate neoplastic growth. The pocket domain of the RB family proteins is dramatically hampered in its functions by the interference of a number of proteins produced by the small DNA viruses. In the last two decades, the 'viral hypothesis' of cancer has received a considerable renewed impulse from the notion that small DNA viruses, such as Adenovirus, Human papillomavirus (HPV) and Polyomavirus, produce factors that can physically interact with major cellular regulators and alter their function. These viral proteins (oncoproteins) act as multifaceted molecular devices that have evolved to perform very specific tasks. Owing to these features, viral oncoproteins have been widely employed as invaluable experimental tools for the identification of several key families of regulators, particularly of the cell cycle homeostasis. Adenovirus early-region 1A (E1A) is the most widely investigated small DNA tumor virus oncoprotein, but relevant interest in human oncology is raised by the E1A-related E7 protein from transforming HPV strains and by Polyomavirus oncoproteins, particularly large and small T antigens from Simian virus 40, JC virus and BK virus.     

DNA alkylation in the hamster induced by two pancreatic carcinogens

N-Nitrosobis(2-oxopropyl)amine (BOP) and N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine (HPOP) alkylate DNA and other macromolecules in the liver, kidney, pancreas, and lungs when injected s.c. in the Syrian hamster. Two of the most abundant DNA adducts found were the N7-methylguanine and O6-methylguanine, which in the liver accounted for about 60% of total DNA alkylation. A third adduct which was invariably found in liver and kidneys, but could not always be detected in pancreas and lungs, was identified as N7-(2-hydroxypropyl)guanine. Quantitation of N7-methylguanine by its UV spectrum and radioactivity, following administration of single-labeled [1-14C]BOP or HPOP, showed that the specific activity of this adduct was one half that of the nitrosamine. This excludes participation of the gamma carbons of these nitrosamines in methylation reactions and indicates that intermediates in which scrambling of the alpha and gamma carbons is possible are not involved in yielding the ultimate methylating agent. Finally, a comparison of the alkylation levels caused by equivalent doses of BOP and HPOP showed that BOP targeted DNA and other cytoplasmic components of kidney, lungs, and pancreas more extensively than HPOP. Ratios of N7-methylguanine in BOP versus HPOP treated hamsters, at doses less than 40 mg/kg body weight, were 3.1 in the kidney, 7.0 in the pancreas, 3.9 in the lung, and only 3.5 in the liver. These ratios are in accordance with the greater carcinogenic potency of BOP compared to HPOP and also the different organotropic properties of the two carcinogens.     

Gangliosides as targets for immunotherapy for pancreatic adenocarcinoma

BACKGROUND: Pancreatic adenocarcinoma cells express gangliosides and sialyl Lewis (sLe) antigens. It is not known whether these carbohydrate antigens can be targeted by immunotherapy. The authors measured the expression of GM(2) and sLe antigens on the surface of pancreatic carcinoma cells and the serum levels of total gangliosides, GM(2), and antiganglioside antibodies in patients with pancreatic carcinoma. METHODS: Cell surface GM(2) and sLe antigens were measured by cell suspension enzyme linked immunoadsorbent assay (ELISA) in four pancreatic carcinoma cell lines. Sera from 20 pancreatic carcinoma patients and 20 age- and gender-matched healthy volunteers were analyzed for antiganglioside and anti-sLe immunoglobulin (Ig) M titers by ELISA. Serum levels of total gangliosides and GM(2) also were measured. RESULTS: All cell lines expressed GM(2) and sLe antigens. When compared with age- and gender-matched volunteers, patients had significantly higher serum levels of total gangliosides (25.6 +/- 9.0 mg/dL vs. 15.6 +/- 2.7 mg/dL; P < 0.001), GM(2) (0.278 +/- 0.415 mg/dL vs. 0.013 +/- 0.018 mg/dL; P = 0.02), ELISA units of anti-GM(2) IgM antibody (368 +/- 95 vs. 155 +/- 25; P = 0.04) and anti-GD(1b) IgM antibody (351 +/- 91 vs. 138 +/- 26; P = 0.03), but not anti-sLe(x) IgM (1389 +/- 345 vs. 1081 +/- 224; P = 0.46) or anti-sLe(a) IgM antibody (1097 +/- 253 vs. 1200 +/- 315; P = 0.80). Patients with unresectable tumors had higher serum levels of total gangliosides compared with patients with resectable tumors, and a serum level > 25 mg/dL was found to correlate significantly with poor overall survival (P < 0.02). CONCLUSIONS: Increased serum levels of total gangliosides and GM(2) may reflect shedding or release of gangliosides from the surface of tumor cells. Production of IgM antibody against GM(2) and GD(1b) indicates that these gangliosides are immunogenic antigens that may be potential targets for effective active immunotherapy.     

Apoptosis regulators as targets for cancer therapy

Apoptosis serves to remove excess or damaged cells and its dysregulation may lead to a number of pathological disorders including cancer. Studies during the last 20 years have unravelled much of the molecular mechanisms that control apoptosis. Whether a cell dies in response to diverse apoptotic stimuli, including DNA-damaging agents, is determined largely by interactions between proteins of the Bcl-2 family. A death signal is transmitted through the BH3-only proteins to Bax and Bak which in turn permeabilise the outer mitochondrial membrane allowing the release of apoptogenic factors, which triggers activation of cell-deathpromoting caspases. These proteolytic enzymes are tightly controlled by members of the inhibitor of apoptosis (IAP) family. Activation of the caspase cascade via cell death receptors also represents a key apoptotic pathway in both normal and tumour cells. Basic knowledge of these apoptosis regulators provides the basis for novel therapeutic strategies aimed at promoting tumour cell death or enhancing susceptibility to apoptotic inducers. This review focuses on these strategies.     

Aurora kinases as targets for cancer therapy

Aurora kinases represent a family of serine/threonine kinases highly conserved during evolution, whose main function is to promote mitotic spindle assembly by regulating centrosome duplication and separation. Inhibition of Aurora kinase activity may lead to defects in centrosome function, misaligned sister chromatids, mitotic spindle malformation, problematic cytokinesis and eventually mitotic arrest. Aurora kinases are overexpressed in a variety of tumor cell lines, suggesting their potential role in tumorigenesis and indicating that they could represent an appealing target for molecular therapies. Extensive pre-clinical information supports the development of Aurora kinase inhibitors in specific tumor types and a number of these novel agents are currently being extensively studied in phase I and II clinical trials exhibiting an acceptable toxicity profile and promising clinical efficacy. The current study aims to provide a comprehensive overview of the development of Aurora kinases as molecular targets for anticancer therapy by focusing on their physiological role in mitosis, their implication in oncogenesis and the potential ways of inhibiting their activity. The main pre-clinical and clinical studies concerning Aurora kinase inhibitors currently under investigation are reported and important considerations for their future development are discussed.     

Mitochondria as therapeutic targets for cancer chemotherapy

Mitochondria are vital for cellular bioenergetics and play a central role in determining the point-of-no-return of the apoptotic process. As a consequence, mitochondria exert a dual function in carcinogenesis. Cancer-associated changes in cellular metabolism (the Warburg effect) influence mitochondrial function, and the invalidation of apoptosis is linked to an inhibition of mitochondrial outer membrane permeabilization (MOMP). On theoretical grounds, it is tempting to develop specific therapeutic interventions that target the mitochondrial Achilles' heel, rendering cancer cells metabolically unviable or subverting endogenous MOMP inhibitors. A variety of experimental therapeutic agents can directly target mitochondria, causing apoptosis induction. This applies to a heterogeneous collection of chemically unrelated compounds including positively charged alpha-helical peptides, agents designed to mimic the Bcl-2 homology domain 3 of Bcl-2-like proteins, ampholytic cations, metals and steroid-like compounds. Such MOMP inducers or facilitators can induce apoptosis by themselves (monotherapy) or facilitate apoptosis induction in combination therapies, bypassing chemoresistance against DNA-damaging agents. In addition, it is possible to design molecules that neutralize inhibitor of apoptosis proteins (IAPs) or heat shock protein 70 (HSP70). Such IAP or HSP70 inhibitors can mimic the action of mitochondrion-derived mediators (Smac/DIABLO, that is, second mitochondria-derived activator of caspases/direct inhibitor of apoptosis-binding protein with a low isoelectric point, in the case of IAPs; AIF, that is apoptosis-inducing factor, in the case of HSP70) and exert potent chemosensitizing effects.