Anti-Cancer Agents in Medicinal Chemistry (Formerly Current Medicinal Chemistry - Anti-Cancer Agents) - Volume 8, Issue 4, 2008

A number of human proteins have been characterized which have important roles in pathways responsible for sensing, responding, and repairing DNA damage. Collectively, these proteins are referred to as DNA repair or DNA damage response proteins, but their functions are specific in terms of pathways that can be potentially manipulated to modulate the biological response to a given cytotoxic agent. Selective inactivation of a DNA repair pathway may enhance existing or developing anticancer therapies. The scope of this Hot Topics series will be to discuss DNA repair proteins that can be targeted to improve cancer therapeutic approaches. A collection of review articles is presented that provides a unique prospective of discussing emerging concepts and strategies to fight cancer through DNA repair inhibition. Treatment with anti-cancer drugs such as small molecule compounds that modulate the expression or activity of a DNA repair protein in specific DNA damage response pathways is proposed to represent a viable approach to selectively kill cancer cells exposed to DNA damaging chemotherapy or radiation. DNA repair proteins that may be suitable targets for fighting cancer are diverse and involve steps of pathways from a variety of DNA repair processes. An overview of DNA repair proteins as molecular targets for cancer therapeutics and discussion of results from experimental studies providing proof-of-concept is addressed by Drs. M. Kelley and M. Fishel. Dr. M. Bignami and colleagues discuss the role of mismatch repair and O6-methylguanine-DNA-methyltransferase in the response to anticancer therapies. Inhibition of specific base excision repair proteins to improve the efficacy of current cheomotherapy strategies is presented in the review by Dr. R. Roy and co-workers. Drs. G. Maga and U. Hübscher present translesion DNA polymerases as a novel target for anti-cancer drugs. Rationale for the development of Tyrosyl-DNA phophosphodiesterase 1 inhibitors is offered by Dr. Y. Pommier and colleagues. Cancer therapy mediated by polynucleotide kinase inhibitors is discussed by Drs. J.N.M. Glover, M. Weinfeld and colleagues. Drs. A.S. Balajee and E.T. Sakamoto Hojo review the prospect of targeting Poly(ADP) ribose polymerase and interacting proteins for cancer treatment. Dr. S. Cantor and colleagues evaluate the evidence that the BRCA-FA pathway is a promising target for anti-tumor drugs. Drs. S. Powell and L. Kachnic elaborate on the therapeutic exploitation of tumor cell defects in homologous recombination. Drs. R. Gupta and R. Brosh propose that helicasedependent DNA repair pathways represent a viable approach to kill cancer cells. Collectively, this Hot Topics review series provides a timely discussion of how DNA repair proteins engaged in distinct pathways of DNA maintenance represent viable candidates for improving the efficacy of anti-cancer therapies.

In both bacteria and eukaryotes the alkylated, oxidized, and deaminated bases and depurinated lesions are primarily repaired via an endogenous preventive pathway, i.e. base excision repair (BER). Radiation therapy and chemotherapy are two important modes of cancer treatment. Many of those therapeutic agents used in the clinic have the ability to induce the DNA damage; however, they may also be highly cytotoxic, causing peripheral toxicity and secondary cancer as adverse side effects. In addition, the damage produced by the therapeutic agents can often be repaired by the BER proteins, which in effect confers therapeutic resistance. Efficient inhibition of a particular BER protein(s) may increase the efficacy of current chemotherapeutic regimes, which minimizes resistance and ultimately decreases the possibility of the aforementioned negative side effects. Therefore, pharmacological inhibition of DNA damage repair pathways may be explored as a useful strategy to enhance chemosensitivity. Various agents have shown excellent results in preclinical studies in combination chemotherapy. Early phase clinical trials are now being carried out using DNA repair inhibitors targeting enzymes such as PARP, DNA-PK or MGMT. In the case of BER proteins, elimination of N-Methylpurine DNA glycosylase (MPG) or inhibition of AP-endonuclease (APE) increased sensitivity of cancer cells to alkylating chemotherapeutics. MPG-/- embryonic stem cells and cells having MPG knock-down by siRNA are hypersensitive to alkylating agents, whereas inhibition of APE by small molecule inhibitors sensitized cancer cells to alkylating chemotherapeutics. Thus, MPG and other BER proteins could be potential targets for chemosensitization.

The cytotoxicity of many antineoplastic agents is due to their capacity to damage DNA and there is evidence indicating that DNA repair contributes to the cellular resistance to such agents. DNA strand breaks constitute a significant proportion of the lesions generated by a broad range of genotoxic agents, either directly, or during the course of DNA repair. Strand breaks that are caused by many agents including ionizing radiation, topoisomerase I inhibitors, and DNA repair glycosylases such as NEIL1 and NEIL2, often contain 5'- hydroxyl and/or 3'-phosphate termini. These ends must be converted to 5'-phosphate and 3'-hydroxyl termini in order to allow DNA polymerases and ligases to catalyze repair synthesis and strand rejoining. A key enzyme involved in this end-processing is polynucleotide kinase (PNK), which possesses two enzyme activities, a DNA 5'-kinase activity and a 3'-phosphatase activity. PNK participates in the single-strand break repair pathway and the non-homologous end joining pathway for double-strand break repair. RNAi-mediated down-regulation of PNK renders cells more sensitive to ionizing radiation and camptothecin, a topoisomerase I inhibitor. Structural analysis of PNK revealed the protein is composed of three domains, the kinase domain at the C-terminus, the phosphatase domain in the centre and a forkhead associated (FHA) domain at the N-terminus. The FHA domain plays a critical role in the binding of PNK to other DNA repair proteins. Thus each PNK domain may be a suitable target for small molecule inhibition to effectively reduce resistance to ionizing radiation and topoisomerase I inhibitors.

Tumor resistance to cytotoxic chemotherapy drugs and their toxicity to normal cells are major clinical obstacles to anticancer therapy effectiveness. Alterations in various DNA repair pathways play a key role in the development of both mechanisms of drug resistance and toxicity. Since deregulation of the DNA damage response and alterations in DNA repair pathways are relatively common in human cancer, the knowledge of these alterations in cancer cells would be an important predictive factor for the clinical response to chemotherapy and a useful guide in designing an appropriate therapeutic strategy. This review is focused on the mismatch repair (MMR) pathway and the O6-methylguanine-DNA-methyltransferase (MGMT) repair protein. In particular, we examine how inactivation of these DNA repair mechanisms might affect the response of tumor cells to chemotherapy, with a special emphasis on agents inducing methylation and oxidative DNA damage and interstrand DNA cross-links (ICLs). In addition, we provide novel experimental evidence indicating that MMR is required for efficient repair of ICLs via stabilization of RAD51 containing repair intermediates. Finally, we discuss possible emerging therapeutical strategies for treating MMR-defective tumors.

Tyrosyl-DNA phosphodiesterase 1 (Tdp1) is a recently discovered enzyme that catalyzes the hydrolysis of 3'-phosphotyrosyl bonds. Such linkages form in vivo following the DNA processing activity of topoisomerase I (Top1). For this reason, Tdp1 has been implicated in the repair of irreversible Top1-DNA covalent complexes, which can be generated by either exogenous or endogenous factors. Tdp1 has been regarded as a potential therapeutic co-target of Top1 in that it seemingly counteracts the effects of Top1 inhibitors, such as camptothecin and its clinically used derivatives. Thus, by reducing the repair of Top1-DNA lesions, Tdp1 inhibitors have the potential to augment the anticancer activity of Top1 inhibitors provided there is a presence of genetic abnormalities related to DNA checkpoint and repair pathways. Human Tdp1 can also hydrolyze other 3'-end DNA alterations including 3'-phosphoglycolates and 3'-abasic sites indicating it may function as a general 3'-DNA phosphodiesterase and repair enzyme. The importance of Tdp1 in humans is highlighted by the observation that a recessive mutation in the human TDP1 gene is responsible for the inherited disorder, spinocerebellar ataxia with axonal neuropathy (SCAN1). This review provides a summary of the biochemical and cellular processes performed by Tdp1 as well as the rationale behind the development of Tdp1 inhibitors for anticancer therapy.

It has been proposed that selective inactivation of a DNA repair pathway may enhance anti-cancer therapies that eliminate cancerous cells through the cytotoxic effects of DNA damaging agents or radiation. Given the unique and critically important roles of DNA helicases in the DNA damage response, DNA repair, and maintenance of genomic stability, a number of strategies currently being explored or in use to combat cancer may be either mediated or enhanced through the modulation of helicase function. The focus of this review will be to examine the roles of helicases in DNA repair that might be suitably targeted by cancer therapeutic approaches. Treatment of cancers with anti-cancer drugs such as small molecule compounds that modulate helicase expression or function is a viable approach to selectively kill cancer cells through the inactivation of helicase-dependent DNA repair pathways, particularly those associated with DNA recombination, replication restart, and cell cycle checkpoint.

Cancer is a disease of uncontrolled cellular proliferation. Chemotherapy and radiation therapy are the two main modalities for cancer treatment. However, some cancer types have been found to be refractory to these treatments. Additionally, certain chemicals that are used in clinical trials produce high cytotoxicity as a secondary effect. Hence, current research is focused on finding ways by which cancer cells can be specifically sensitized to apoptotic death with minimal or no secondary effects on normal healthy cells. Since the resistance of cancer cells to DNA damaging agents stems from the modulation of DNA repair pathways, pharmacological inhibition of these pathways has been emerging as an effective tool for cancer treatment. Inhibition of key proteins involved in the molecular cascade of DNA damage detection and repair such as poly (ADP) ribose polymerase I (PARP-1) and its interacting proteins [DNA dependent protein kinase (DNA-PK) and Cockayne syndrome group B (CSB)] has recently proven to be successful for the treatment of various types of cancer cells and tumor xenografts in vitro. This review summarizes some of the recent findings and the potential application of DNA repair inhibitors in cancer treatment.

Cancer therapeutics include an ever-increasing array of tools at the disposal of clinicians in their treatment of this disease. However, cancer is a tough opponent in this battle and current treatments which typically include radiotherapy, chemotherapy and surgery are not often enough to rid the patient of his or her cancer. Cancer cells can become resistant to the treatments directed at them and overcoming this drug resistance is an important research focus. Additionally, increasing discussion and research is centering on targeted and individualized therapy. While a number of approaches have undergone intensive and close scrutiny as potential approaches to treat and kill cancer (signaling pathways, multidrug resistance, cell cycle checkpoints, anti-angiogenesis, etc.), much less work has focused on blocking the ability of a cancer cell to recognize and repair the damaged DNA which primarily results from the front line cancer treatments; chemotherapy and radiation. More recent studies on a number of DNA repair targets have produced proof-of-concept results showing that selective targeting of these DNA repair enzymes has the potential to enhance and augment the currently used chemotherapeutic agents and radiation as well as overcoming drug resistance. Some of the targets identified result in the development of effective single-agent anti-tumor molecules. While it is inherently convoluted to think that inhibiting DNA repair processes would be a likely approach to kill cancer cells, careful identification of specific DNA repair proteins is increasingly appearing to be a viable approach in the cancer therapeutic cache.

Promising research on DNA repair signaling pathways predicts a new age of anti-tumor drugs. This research was initiated through the discovery and characterization of proteins that functioned together in signaling pathways to sense, respond, and repair DNA damage. It was realized that tumor cells often lacked distinct DNA repair pathways, but simultaneously relied heavily on compensating pathways. More recently, researchers have begun to manipulate these compensating pathways to reign in and kill tumor cells. In a striking example it was shown that tumors derived from mutations in the DNA repair genes, of BRCA-FA pathway, were selectively sensitive to inhibition of the base excision repair pathway. These findings suggest that tumors derived from defects in DNA repair genes will be easier to treat clinically, providing a streamlined and targeted therapy that spares healthy cells. In the future, identifying patients with susceptible tumors and discovering additional DNA repair targets amenable to anti-tumor drugs will have a major impact on the course of cancer treatment.

We have very recently highlighted possible connections between DNA polymerases, the main enzymes in the DNA metabolism, and human diseases (Ramadan, K., Maga, G. and Hubscher, U.: DNA polymerases and diseases, In: Genome Integrity: Facets and Perspectives ed. Lankenau, D.-H. Springer Verlag, Heidelberg Germany, Vol 1, pp. 69-102, 2007). Beside a role in DNA replication of the genome DNA polymerases have fundamental functions in other aspect of DNA metabolism, such as DNA repair, DNA recombination, translesion DNA synthesis and cell cycle checkpoint. In the last decade many novel DNA polymerases have been identified, but their exact cellular functions still await clarification. We know that many DNA polymerases have redundant functions. It is a fact that specific inhibition of certain DNA polymerases is a promising approach to develop anticancer drugs. In this review we will concentrate on DNA repair proteins and translesion DNA polymerases as possible targets for anti cancer drugs.

In the decade since the BRCA1 and BRCA2 genes were cloned, much has been learned about the function of these two major causes of familial breast cancer. BRCA2 has been shown to play a direct role in the repair of DNA by homologous recombination, by interacting with the Rad51 protein and facilitating the formation of Rad51 aggregates at the site of DNA damage. It likely plays a similar role when double strand breaks are created in the course of normal DNA replication; the absence of BRCA2 results in chromosomal instability, which is likely secondary to the defect in DNA repair. In the absence of BRCA2, the cell is more dependent on residual repair via Rad52, which makes Rad52 a target for therapy in BRCA-deficient tumors. BRCA1 plays a role in sensing DNA damage and replication stress and mediating the signaling responses. Therefore, in addition to its role in mediating DNA repair by homologous recombination via BRCA2, it can also signal cell cycle checkpoints and mediate other transcriptional responses to DNA damage. We have argued that the mechanism of cancer susceptibility from BRCA1 or BRCA2 deficiency is mediated via the defect in homologous recombination, since it is the main feature they share in common. We and others have recently demonstrated that the defect in homologous recombination changes the drug sensitivity profile, rendering the BRCA-deficient breast cancers sensitive to MitomycinC, cisplatin, etoposide and other drugs that produce complex double-stranded lesions in DNA. Furthermore, they show resistance to taxanes and navelbine. Fanconi anemia defective cells also show sensitivity to the same class of drugs, although their defect in homologous recombination in response to strand breaks appears less marked than in BRCA-deficient cells. However, Fanconi anemia cells also show chromosomal fragility, and appear to have defects in maintenance of the replication fork. Therefore, knowledge of whether this specific DNA repair pathway of homologous recombination is defective in breast cancer cells would be valuable information in planning optimized individual therapy. We have developed techniques to measure the functional integrity of homologous recombination in human breast cancers. Core biopsy samples are obtained and immediately irradiated ex vivo, allowing 3-4 hours for the appearance of Rad51, BRCA1 and FancD2 foci. Thin sections are obtained, permeabilized and stained by immunofluorescent techniques. We have identified tumors with defects in the ability to form Rad51 and BRCA1 foci, where there is no known genetic predisposition, implying that this BRCA-dependent repair pathway may be inactivated in sporadic as well as familial breast cancers. Thus, functional assays of homologous recombination could become a useful technique to determine phenotype of human breast cancer, which in turn will influence the choice of therapy.