SCIENTIFIC FOCUS AND GOALS
The overall scientific goal within the DNA Damage and Cellular Defense (DDCD) Program is to understand the mechanisms and processes whereby cells sustain – or alternatively, mitigate by prevention or repair – damage to DNA or other critical macromolecular structures. The Program is an interactive group of 23 investigators that brings together cancer researchers with diverse approaches to two complementary themes that are illustrated in the diagram below: 
DNA Damage and Cellular Defense Program | Cell Growth and Survival Program |
Figure 1. Schematic overview of the research focus in the basic science Programs.
Theme 1: Mechanisms of damage to DNA and other macromolecules: detrimental role in carcinogenesis versus beneficial role in chemotherapy.
The detrimental role of somatic and germline mutations in the DNA sequence of key growth regulatory genes is generally accepted to be of paramount importance in the initiation and stepwise progression to malignancy (Figure 1). Ironically, DNA damage is also employed for beneficial therapeutic advantage as a means to selectively kill cancer cells. This occurs via apoptosis that is induced by cellular signaling of DNA damage caused by chemotherapeutic drugs or radiation. The mechanisms whereby these mutations orcytotoxic lesions are induced by different reactive electrophiles of either endobiotic (e.g. reactive oxygen species) or xenobiotic origin (e.g. carcinogens or chemotherapeutic drugs), or by errors of DNA replication that are not properly repaired, are the research focus of about half of the members of the DDCD Program.
Research in Theme 1 revolves around two complementary goals:
1a) To investigate the mechanisms and consequences of DNA damage caused by endogenous or exogenous electrophiles or radiation, and to determine their contributions to carcinogenesis;
1b) To learn how DNA damage can be exploited to improve existing chemo– and radiation therapies, and to develop new cytotoxic and radiosensitizing agents.
Theme 2: Mechanisms and significance of cellular defense: beneficial role in cancer prevention versus detrimental role in chemotherapeutic drug resistance.
Conversely, evolution has endowed mammalian cells with a remarkable array of defensive mechanisms that serve to protect against mutagenic and cytotoxic effects of DNA damage, or to repair DNA damage if it is not prevented (Figure 1). These include a range of mechanisms such as detoxification enzymes, membrane efflux transport proteins, alternative nucleophilic targets for alkylation, cell cycle regulatory changes, DNA damage tolerance, and DNA repair pathways. Genetic and experimental evidence supports the significance of these pathways in human cancer. The challenge for the future is to develop a mechanistic understanding of protection against specific DNA-damaging agents and translate this information into improvements in cancer therapy. In particular, there is a need to learn how these mechanisms may be enhanced or inhibited as a step toward better strategies in chemoprevention, or for enhancement of chemotherapeutic drug efficacy.
Research in Theme 2 is similarly focused around two interrelated goals:
2a) To identify cellular defense mechanisms that protect genomic integrity by either prevention or repair of DNA damage, and to examine the consequences of their efficacy or failure;
2b) To determine how cellular defense factors modulate the susceptibility to chemotherapeutic orradiation treatment and to develop novel chemopreventive agents.
The knowledge gained through basic science investigations into these DNA damage and cellular defense mechanisms provides the foundation for development of strategies to: a) reduce or delay the genetic damage that leads to the development of cancer, and b) enhance the selective killing of tumor cells by chemotherapy and radiation. For example, mutations in, or altered expression of, key genes involved in detoxification or repair may confer upon certain individuals a cancer-prone phenotype. These individuals might benefit from genetic counseling to encourage exposure and lifestyle modifications to reduce cancer risk. Similarly, if these same mechanisms in tumor cells give rise to resistance to drugs or radiation, then they could become targets for modulation of chemo- or radio-therapy. These basic science themes encourage a broad perspective and an integrated and collaborative approach to these interrelated topics. An interwoven translational theme of the Program is to initiate projects leading to the development and implementation of novel prevention and treatment strategies to reduce the risk of cancer in human populations, and to more effectively diagnose and treat malignancies.
Several areas of emphasis in the Program comprise a major part of the DDCD Program emphasis. In each of these subtopics there is a focus on both the detrimental (carcinogenic) and beneficial (anticancer) facets of the research:
1) biology of reactive oxygen species (ROS): damage enhancement by radiosensitizers, and functions of antioxidant defenses in protection from oxidative damage;
2) DNA damaging agents as mutagenic carcinogens or cytotoxic chemotherapeutic agents;
3) Role of DNA replication and repair pathways in facilitation vs. prevention of mutations or apoptosis;
4) Enzymatic activation versus enzymatic detoxification of DNA-damaging electrophiles; and
5) Chemistry–biology interface: synthesis / testing of chemotherapeutic and chemopreventive agents.
The interrelated nature of the DDCD Program research efforts in these areas will be evident as a recurrent theme, and will be discussed in greater detail within this context in the following sections on our Research and Translational Accomplishments.
The following are brief descriptions of the major areas of research focus within the DDCD Program that span both of the major research themes. Key findings selected from these areas will be highlighted in more detail later under Research Accomplishments.
1) Induction of Damage to DNA or Other Macromolecules by Reactive Oxygen Species, and the Role of Antioxidant Defenses
Ionizing radiation induces DNA single-strand and double-strand breaks through a mechanism involving the radiolytic scission of water to form extremely reactive hydroxyl radicals and other reactive oxygen species (ROS), which then attack DNA or other macromolecules such as proteins or lipids. This process, though mutagenic and detrimental under normal circumstances, is used to advantage in radiation therapy to preferentially destroy cancer cells. Research on the biology of ROS is a major focus of the DDCD program, and has led to new discoveries with potential for translation into cancer therapy. For example, DDCD investigators have discovered radiosensitizing activity of agents previously characterized primarily as either chemotherapeutic or chemopreventive agents (Drs. Suzy Torti, William Gmeiner, and Costas Koumenis). Another major area of emphasis of research on ROS is the role of sulfur chemistry, and specifically of protein cysteine sulfenic acid intermediates as both markers of oxidative damage and as sensors and/or regulators of oxidant potential in peroxiredoxins, proteins that likely function in both antioxidant and also redox signaling roles (Drs. Leslie Poole, Jacqueline Fetrow, Bruce King, and Todd Lowther). This is an entirely new area of cellular biochemistry that has major implications for cellular sensitivity to oxidative stress as well as regulation of intracellular oxidant signaling, which is a highly active area of collaboration with investigators in the Cell Growth and Survival Program. The role of NADPH oxidase in oxidative stress and in brain radiation injury is a third area of interest (Drs. Linda McPhail and Mike Robbins), which has led to collaborations with neuroscientists and clinicians in the Clinical Research Program.
2) Oncogenic Mutagenicity or Therapeutic Cytotoxicity of DNA–Damaging Agents
Recognition of the potent mutagenesis of chemical carcinogens that could induce covalent DNA adducts resulted in a paradigm shift in our understanding of the essential role of genetic damage in cancer etiology. The process of discovering the environmental sources of exposure to these alkylating species, identifying their metabolic activation and disposition pathways, defining the DNA lesions that are actually mutagenic and/or carcinogenic, and elucidating their precise mechanisms of mutagenicity remains a major unfinished challenge in cancer research. Several labs focus on mutagenesis by alkylating agents (Drs. Steve Akman, Fred Perrino) or mutagen activation pathways (Drs. Mark Miller and Townsend) and their role in prenatal susceptibility to carcinogens that require metabolic activation (Miller). In addition to the environmental carcinogenesis component of chemical DNA damage, other DDCD labs are engaged in experimental therapeutics projects to develop and test novel DNA targeted chemotherapeutic agents (Drs. Ulrich Bierbach, Gmeiner, Greg Kucera, and S. Torti). These efforts have resulted in three drug development grant awards, including two Rapid Access to Intervention Development (RAID) grants (Gmeiner and S. Torti) and a subsequent Rapid Access to New Drugs (RAND) award (S. Torti).
3) DNA Replication and Repair as Promutagenic or Antimutagenic Mechanisms
Carcinogenic mutations may be induced by unrepaired replication errors, by error-prone replication of template DNA strands damaged by DNA adducts, or by other errors introduced by faulty repair of DNA lesions. This category of mutagenic mechanisms may account for much of the disparity in potency among seemingly similar alkylating agents. Several DDCD investigators (Akman, Drotschmann, and Perrino) focus their research on the role of pathways of DNA replication and repair in cellular sensitivity to mutagenesis or lethality by DNA damaging mutagens. Both promutagenic and antimutagenic effects of so-called “bypass” DNA polymerases (Perrino) with different DNA adducts are a focus of the Perrino-Akman collaboration. These studies are predicated on the notion that a better understanding of DNA repair or maintenance pathways will help to provide a rational basis for the cancer susceptibility profiles that occur in individuals with specific defects in these pathways. Conversely, in cancers that are linked to DNA repair defects, these studies may identify vulnerabilities that may be exploited therapeutically.
DNA repair pathways may also modify the anticancer efficacy of DNA-damaging chemotherapeutic drugs. This can result in resistance via reversal of the DNA damage. Moreover, recent evidence suggests that mismatch repair proteins mediate the cell death response to DNA damage. Thus, one DDCD project (Drs. Karin Drotschmann and Thomas Hollis) aims to understand how DNA repair proteins participate in the apoptotic response to chemotherapeutic agents, and how repair defects may serve as prognostic markers to predict the efficacy of chemotherapy. Another novel DDCD project has resulted in the first purification, characterization, cloning, expression and solution of the 3-dimensional structure of novel human 3’-exonucleases that may play a role in DNA repair (Perrino and Hollis). A third project has discovered a unique quadruplex DNA resolvase activity, which was first purified, cloned and expressed at this Cancer Center (Akman).
4) Enzymatic Activation Versus Cellular Defenses Against DNA Damage by Electrophiles
Several decades have elapsed since the importance of metabolic activation (“phase I”) and detoxification (“phase II”) pathways in the intracellular formation and disposition of reactive electrophilic carcinogens was first appreciated. The role of phase II enzymes and other proteins in protection against cellular damage via detoxification of electrophiles has long been an area of research focus within the DDCD Program. This area includes mechanisms of enzymatic activation versus detoxification of electrophilic chemical carcinogens (Townsend) or cytotoxic drugs (Morrow and Townsend), and also protection via other mechanisms such as by chelation of iron by ferritin, thus preventing production of hydroxyl radicals via Fenton chemistry. Two of the most prolific collaborations within the DDCD Program are focused on the detoxification functions of glutathione transferases (GST) together with glutathione conjugate transporters (Morrow and Townsend), and on prevention of iron toxicity by ferritin (F. Torti and S. Torti). The goal of these studies is to define the mechanistic basis and specificity of the roles of these proteins in prevention of DNA damage and resulting mutagenicity or cytotoxicity of carcinogens or chemotherapeutic drugs. This research has a major translational emphasis in the area of chemoprevention. Indeed, this work laid the foundation for the collaboration with Dr. Mark Welker, a Chemistry Dept. faculty member in the DDCD Program, who has synthesized novel chemopreventive compounds that were tested in the Torti and Townsend labs and shown to induce expression of ferritin, GST and other protective proteins.
5) Chemistry–biology interface at CCCWFU: synthesis and testing of novel chemotherapeutic and chemopreventive agents
The cancer demographics of recent decades, and expected trends for the next several decades portend a significant increase in cancer diagnoses due to an aging population, lifestyle / exposures, and more effective early detection. Thus, new therapeutic breakthroughs are urgently needed. The DDCD Program Leader, Dr. Townsend, recruited members of the Chemistry Department to become members of CCCWFU beginning nearly a decade ago. Currently, 6 Chemistry Department faculty are members of the DDCD Program. Research at the chemistry-biology interface of CCCWFU has resulted in an innovative new class of acridine-conjugated platinum compounds (Bierbach) that show promising activity. Another urgently needed approach is the development of effective strategies to prevent or at least delay the progression of abnormal (i.e. damaged) cells to frank malignancy.
Research to test for chemopreventive activity by dietary or pharmacologic agents is now a major emphasis of research in the DDCD Program. This is approached by a variety of strategies, from synthesis and testing of new compounds to animal and human studies with nutriceuticals. The strong interactions between the Cancer Center and the WFU Chemistry Department have resulted in a fruitful collaboration involving the synthesis (Welker, Chemistry) and testing of novel dithiolthione analogs related to the chemopreventive agent oltipraz, using a cell culture-based phase II enzyme induction assay (S. Torti and Townsend, Biochemistry). Compounds implicated as so-called complementary or alternative medicine nutriceuticals, such as caffeic acid phenethyl ester (Koumenis), sulindac (Miller) or curcumin (S. Torti) are tested in animal models of cancer. The studies with curcumin have led to a funded proposal to conduct a clinical trial for prevention of cervical intraepithelial neoplasia by topical application (S. Torti; C. Koumenis).