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Research Background

Conventional cancer therapies target the characteristics of uncontrolled growth of cancer cells. Due to high level of DNA replication DNA damage compromising or inhibiting DNA replication can trigger cell death. However, cell death is also triggered in healthy tissues, causing dose-limiting side effects. Thus, many efforts are focused on developing cancer therapies that selectively kill cancer cells without adversely affecting healthy tissues. The concept of ‘synthetic lethality’ refers to the cell-lethal effects of the combined inactivation of two genetically distinct pathways. In a cancer setting, synthetic lethality provides a conceptual framework for the development of drugs that are selectively toxic in specific genetic backgrounds associated with tumors. One example is the synthetic lethality of PARP inhibition in tumors carrying mutations in brca1 or brca2. It is likely that other examples of DNA repair-associated synthetic lethal relationships relevant to cancer exist.

We are focusing on small molecules and genome editing strategies that target cancer cells more specifically based on unique genomic features. We will use them as probes to study DNA repair mechanisms and to evaluate their therapeutic potential for cancer treatment.

We previously isolated ~300 small molecules affecting DNA replication and repair by high throughput screening of a ~300,000 compound collection at the United States National Institutes of Health. We found and characterized two small molecules that showed synthetic lethal phenotypes with DNA mismatch repair (MMR) deficient and PARP1-inhibitor resistant brca1 tumor cells, respectively (Figure 1).

Resistance to PARP1-inhibitor in brca defective tumors is typically caused by the activation of HR. We found that a compound, 418 inhibits the autophagy pathway at the stage of lysosome activation. Inhibition of autophagy by 418 results in the selective proteasomal degradation of proteins involved in HR including RAD51 and CHK1. We identified a potential target protein of 418 using the CETSA (Cellular Thermal Shift Assay) method which we recently established.


Figure 1. Small molecules, baicalein and 418 selectively kill mismatch repair-deficient (A, B)

and parp1-deficient tumors (C), respectively

Our research on a small molecule, baicalein, revealed that there is cross-talk between MMR and homologous recombination (HR). We found that the MMR proteins, MSH2-MSH3 have a role in HR mediating the proper recruitment of the EXO1 nuclease to DNA damage sites thus providing us with a promising lead to reveal the complex crosstalk between three different repair pathways (Figure 2). Further analysis of MMR using baicalein led us to the discovery that MSH2-MSH3 directly interacts with the chromatin remodeler SMARCAD1 and EXO1, thereby contributing to the regulation of the DNA end resection step of HR


Figure 2. EXO1 recruitment to UV-laser irradiation depends on SMARCAD1 and MSH2



Recently, we developed an approach that tumors can be targeted by inducing double strand breaks in tumor-specific In/DEL (insertion/deletion) sequences by CRISPR-Cas9 (Figure 3). The selective killing of cancer cells without affecting normal cells is a desirable and often elusive goal in cancer therapy. We developed a potential therapeutic method to achieve selective cancer cell death by targeting a unique feature of cancer cells, cancer-specific small Insertions and deletions (InDels). The CINDELA (Cancer specific INsertions-DELetions (InDels) Attacker) approach targets these Indels with specific guide RNAs and Cas9 leading to cell-type specific DNA double strand breaks, and could provide a broadly applicable approach for targeting a variety of tumors based on tumor specific DNA sequences. We are currently improving CINDELA. In addition, we use CINDELA as a tool to understand various DNA repair mechanisms in cells.


Figure 3. CINDELA strategy selectively kill tumor cells

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