The major theme of our work relates to how DNA repair processes are disrupted in human cancers and how this can be exploited therapeutically. In particular, we have a long-standing focus on homologous recombination, a pathway often dysregulated in human cancers. Our research centers on improving our understanding of how homologous recombination executes its biological functions, including its role in replication-coupled repair. Furthermore, we have interest in the backup pathways needed to maintain cellular viability in the absence of recombination, with the ultimate goal of laying a foundation for the development of mechanism-inspired cancer therapeutics.
In addition to serving as Chair of the Department of Radiation Oncology I serve as the clinical science leader on the MSK SPORE in Genomic Instability in Breast Cancer. As part of the programmatic research efforts in this SPORE our laboratory is involved in efforts to define DNA repair defects in breast cancer with greater reliability and resolution, and to personalize the treatment of breast cancer patients whose tumors display evidence of impaired homologous recombination. We have a wide range of funded projects that can be broadly grouped in the following areas:
Backup DNA repair pathways in HR-deficient human cancers
Defining new targets in HR-deficient cancers. BRCA-deficient cells lacking homologous recombination (HR) are dependent on alternative, often error prone mechanisms, to sustain replication and prevent the formation of DNA intermediates that require HR. Given the incomplete understanding of compensatory mechanisms that allow for HR-deficient cancer cells to sustain replication, we recently conducted a genome-wide CRISPR inactivation screen in unperturbed BRCA2-/- vs BRCA2+/+ cells to identify genes that are synthetically lethal with BRCA2 loss of function. In addition to genes known to be synthetically lethal with BRCA2 loss of function (e.g. POLQ and RAD52), this screen revealed several novel leads that have opened significant new areas of research for the laboratory.
Mechanisms underlying genomic signatures of HR-deficiency. Cancer genomes harbor mutational and structural rearrangements that are jointly shaped by DNA damage and repair mechanisms. Accumulating evidence suggests that genetic alterations in DNA repair-defective tumors reflect the scars of backup repair pathways needed to maintain cellular viability. Detailed analysis of the patterns of mutations and structural rearrangements present in BRCA1/2-deficient tumors has allowed for the delineation of genomic signatures that reflect deficient repair by homologous recombination (HR). In collaboration with Marcin Imielinski and Jorge Reis-Filho, we are investigating the backup repair mechanisms responsible for genomic scarring in HR-deficient tumors.
Replication-coupled DNA damage signaling and repair
The genome is particularly susceptible to DNA damage during replication. Replication forks must deal with a wide variety of potential obstacles to completion of accurate DNA replication, including structured DNA, compact chromatin, and actively transcribed regions. During transcription nascent RNA can base pair with template DNA, displacing the non-template strand as ssDNA, forming a structure known as an R-loop. Depending on the nature of the obstacle or lesion multiple pathways that operate in overlapping layers of repair are known to be engaged, including pathways to manage replication-transcription collisions, or helicase-blocking obstacles such as inter-strand crosslinks. Some types of DNA damage are in fact bypassed, with repriming thought to facilitate subsequent post-replicative repair, while others can lead to replication fork stalling, remodelling, and/or breakage. Many unanswered questions remain including the role of BRCA1, BRCA2, and RAD52 in the cellular response to replication-coupled damage, and the resulting implications for genomic instability in cancer. We have developed a number of approaches to address these questions, including methods to create site-specific replication fork barriers, study post-replicative daughter strand gaps and examine replication-transcription collisions, among others.
Mechanisms of recombination
Different stages in the engagement of homologous recombination can be identified. Key steps include i) DNA end resection, ii) presynaptic filament assembly, iii) strand invasion/D-loop formation, iv) repair synthesis/D-loop extension and v) resolution of recombination intermediates with or without second-end capture. Our lab has an interest in the molecular mechanisms underlying each of these steps, with particular focus on proteins and mechanisms involved in end-resection, D-loop formation/extension, and second-end capture. For example, recently published work demonstrates a previously unappreciated annealing function of the HELQ helicase, including a potential role in second-end capture (https://doi.org/10.1038/s41586-021-04261-0).