Abstract SY28-01: Targeting DNA repair pathways in cancer therapy
Authors: D'Andrea Alan D. 
Published: May 2019

Cancer cells have genomic instability resulting from acquired defects in DNA repair. One DNA repair pathway_the Fanconi Anemia/BRCA homologous recombination pathway (Kennedy and D'Andrea, Genes&Development 19:2925, 2005) _ is defective in many human cancers, including breast, ovarian, pancreatic, and lung neoplasms. Disruption of the FA/BRCA Pathway results in the characteristic chromosome instability and radiation/crosslinker hypersensitivity of these tumors. In general, loss of one DNA repair pathway often leads to hyperdependence on another pathway for tumor cell survival. This hyperdependence offers a unique opportunity for the development of anticancer therapeutics. For instance, FA/BRCA pathway deficient tumors are hyperdependent on Base Excision Repair (BER) and, accordingly, these tumors are hypersensitive to single agent treatment with PARP1 inhibitors which block BER. Our laboratory efforts are focused on profiling the FA/BRCA pathway and the other five major DNA repair pathways in tumor cells (Kennedy and D'Andrea, JCO 24: 3799, 2006). Each DNA repair pathway has a characteristic protein biomarker and repairs a specific type of DNA lesion. By profiling these pathways in primary tumor samples with activation-specific antibodies to DNA repair proteins, we are developing methods (1) to predict the sensitivity of tumors to conventional chemotherapy and radiation (Personalized Medicine) (2) to subset tumors for their sensitivity to novel classes of DNA repair inhibitors, (i.e., PARP1, Chk1, and ATM inhibitors) and (3) to screen for new small molecule inhibitors of other DNA repair pathways. This combination of novel DNA repair inhibitors, conventional DNA damaging agents, and DNA repair biomarkers offers new opportunities for developing more effective anticancer therapy.

The FA/BRCA pathway was elucidated through the systematic study of the rare inherited chromosome breakage disorder, Fanconi Anemia (Moldovan and D'Andrea, Ann Rev Genet 43:223, 2009). FA is an autosomal recessive disease characterized by bone marrow failure, congenital malformations, and cellular sensitivity to Cisplatin, Mitomycin C, and other DNA interstrand crosslinking agents. Patients with FA develop hematopoietic malignancies and squamous cell carcinomas. Based on somatic cell fusion studies, there are thirteen FA complementation groups (A, B, C, D1, D2, E, F, G, I, J, L, M, N), and the corresponding gene for each of these complementation groups has been identified. Interestingly, the thirteen FA proteins cooperate in a common cellular pathway in normal human cells, referred to as the Fannoni Anemia/BRCA pathway. Eight of the FA proteins (A, B, C, E, F, G, L, M) are assembled in a core complex (the FA core complex) which is an active ubiquitin E3 ligase. In response to DNA damage, the FA core complex modifies (monoubiquitinates) the downstream FANCD2 protein. Monoubiquitinated FANCD2 translocates to nuclear foci where it interacts with the FANCD1/BRCA2 protein and participates in the process of homologous recombination DNA repair. Additional FA proteins (namely, FANCJ/BRIP1 and FANCN/PALB2) function downstream of FANCD2 monoubiquitination. At least three of the FA genes (FANCD1, FANCJ, and FANCN) are inherited breast cancer susceptibility genes. Disruption of any step in the FA/BRCA pathway results in the common clinical and cellular phenotype of FA patients.

Human tumor cells, derived from cancer patients from the general (non-FA) population, often exhibit genomic instability. Genomic instability of tumors has important clinical consequences. On the one hand, genomic instability gives the tumor the ability to break and fuse chromosomes, inactivate tumor suppressor genes, form novel oncogene fusions, and amplify drug resistance genes. Thus, the tumor with genomic instability may become more malignant and drug resistant over time. On the other hand, in order to achieve a state of genomic instability, a tumor cell must inactivate one of its major DNA repair pathways. This inactivation appears to account, at least in part, for the selective hypersensitivity of cancer cells to the cytotoxic effects of conventional anti-cancer radiation and chemotherapy.

Recent studies indicate that some human tumors inactivate the FA/BRCA pathway. Pathway inactivation may result from somatic mutation of genes in the FA/BRCA pathway or by epigenetic silencing. For instance, methylation of one of the FA genes, FANCF, has been implicated as a mechanism for genomic instability in a wide variety of cancers, including ovarian, breast, lung, cervical, and head and neck squamous cell carcinomas. Somatic inactivation of the FA/BRCA pathway appears to account for the genomic instability and the cisplatin hypersensitivity of many of these cancers.

A wide array of biomarkers is available for measuring the activity of the FA/BRCA pathway in human tumors, or for measuring the activity of the other human DNA repair pathways. Since many of the genes in the FA/BRCA pathway (i.e., the thirteen cloned FA genes) have been identified, tumors can be screened for germline or somatic mutations in these genes. Furthermore, tumors can be analyzed for function of the pathway by following various biochemical events in the pathway. For instance, the full function of the pathway requires monoubiquitination of the FANCD2 protein, ATR and CHK1 dependent phosphorylation of subunits of the pathway, and assembly of FANCD2 and FANCE nuclear foci. A tumor which has defects in any of these genetic or biochemical biomarkers has a defect in the FA/BRCA pathway and may therefore have a hypersensitivity to crosslinking drugs such as cisplatin. Thus, biochemical monitoring of DNA repair pathways may be a means for predicting drug sensitivity of individual tumors.

Loss of DNA repair pathways can lead to hyperdependence on other survival pathways. Inhibition of these alternative pathways may represent a therapeutic approach which is selectively toxic to repair pathway deficient tumor cells. For instance, breast and ovarian tumor cells which are deficient in the homologous recombination (HR) pathway are hypersensitive to drugs (PARP1 inhibitors) which selectively inactivate a compensatory DNA Repair pathway, the Base Excision Repair (BER) pathway. We recently used a high throughput siRNA screening approach to identify DNA damage response genes that were critical for the survival of FA/BRCA pathway deficient tumor cells (Kennedy et al, J. Clin Invest 117:1440, 2007). Using this approach we identified the DNA damage response kinase, ATM, as being important for the survival of cells deficient in the FA pathway. Accordingly, we found that the Atm -/- Fancg -/- mouse genotype was deleterious when Fancg +/- Atm +/- mice were interbred. We also demonstrated constitutive activation of ATM in FA pathway deficient cells, which was abrogated by reconstitution of the pathway. Furthermore, inhibition of ATM using siRNA oligonucleotides or the specific ATM inhibitor KU55933 resulted DNA breakage and cell death specifically in cells with a non functioning FA pathway.

These data suggest that ATM and the FA pathway function in parallel and compensatory roles following endogenous DNA Damage. Moreover, pharmaceutical inhibition of ATM may be selectively toxic to cancer cells that have lost function of the FA pathway. We are currently exploring other therapeutic strategies to selectively target tumors with a defect in the FA/BRCA pathway or with a defect in one of the other human DNA repair pathways. For instance, we recently determined that tumors which are deficient in the FA/BRCA pathway are hypersensitive to CHK1 inhibitors (Chen et al, Mol Cancer 8: 24, 2009). In my presentation, I will review the features of the six major DNA repair pathways in human cells, and how knowledge of the disruption of these pathways in cancers can lead to the prediction of drug response.

Citation Format: Alan D. D'Andrea. Targeting DNA repair pathways in cancer therapy [abstract]. In: Proceedings of the AACR 101st Annual Meeting 2010; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr SY28-01