Columbia Scientists Discover New Way of Selectively Killing Cancer Cells [link]Columbia Scientist Wins Beckman Young Investigator Award for Research into Novel Methods of Cell Death [link]
Read an article called "Microarrays for Drug Screening" by Linda Sage in the American Chemical Society's Analytical Chemistry that features Bailey et al.'s novel microarray technique [Download PDF] [Download Bailey et al.]
Research:
Diagramming Disease Networks with Chemical and Biological ToolsSummary
We are diagramming the interconnected signaling networks underlying cancer and neurodegenerative diseases using chemical and biological tools. Our approach is to design high-throughput screens in mammalian cells that allow us to test tens of thousands of small organic molecules and small interfering RNAs for their ability to affect cellular phenotypes associated with oncogenic signaling or neurodegeneration. These screens reveal reagents that are used to identify specific proteins and genes that act as the critical regulators of cellular disease pathologies. We define the molecular function of these critical regulators using protein biochemistry, molecular cell biology and chemical synthesis.
Potential research projects include:
• identifying the protein targets and signaling pathways affected by novel compounds we have discovered in screens related to cancer and neurodegeneration
• creating and executing high-throughput screens related to oncogenic signaling and neurodegeneration to identify novel compounds and genes of interest
• defining protein ligation events (ubiquitylation, sumoylation, etc) associated with specific disease proteins
•creating novel photoaffinity reagents, fluorescent sensors and chemical libraries
Probing Oncogenic Signaling with the Tools of Chemical Biology
Discovery of Genotype-Selective Anti-Tumor Agents
Recent advances in characterizing genetic aberrations present in human tumor cells have created the opportunity to design molecularly targeted therapeutic agents. Such oncoprotein-directed reagents may ultimately allow for tailoring of treatment regimens to the specific genetic makeup of each tumor. We are using synthetic lethal screens to identify reagents that selectively kill tumor cells expressing specific oncoproteins. Such selectively lethal compounds may target the designated oncoprotein itself, or they may target other proteins in an oncoprotein-linked signaling network. These reagents allow us to identify signaling pathways that are altered in cells due to expression of these oncoproteins; furthermore, such compounds may be developed into clinically effective therapeutic agents.
We recently discovered a novel compound, derived from a combinatorial library, that selectively kills tumorigenic cells expressing both the RASV12 and Small T (ST) oncoproteins. We observed that this new compound, which we named erastin, kills tumorigenic cells without activating apoptosis, which is surprising given the fact that most antitumor agents act by initiating an apoptotic cell death program. We seek to define the mode of cell death initiated by erastin and the specific proteins that erastin interacts with. Current projects include (i) using cDNA and siRNA libraries to identify specific genes that mediate erastin’s selective lethality, (ii) using expression profiling to define the global response of sensitive and resistant cells to erastin and (iii) performing synthetic lethal screens with other oncogenes and tumor suppressors to identify new genotype-selective compounds.
Defining the Cellular Functions of E3 Ubiquitin Ligases
Synthesis of a Library of E3 Ubiquitin Ligase Inhibitors
Tumor cells exhibit numerous molecular defects compared to normal cells, including changes in the expression and stability of critical regulatory proteins. For example, the human papilloma virus (HPV) E6 oncoprotein appropriates the ubiquitin-proteasome machinery in order to induce degradation of the tumor suppressor protein p53, which is required for DNA damage-induced apoptosis. Such dysregulated protein degradation occurs in a variety of cancers, including HPV-induced cervical cancer, and contributes to the transformation of tumor cells and their insensitivity to anti-tumor agents. Disrupting the activity of E6 and other proteins within the ubiquitin-degradation pathway would restore the level and function of tumor suppressors, such as p53, that would otherwise be degraded. We are developing small-molecule inhibitors of E3 ubiquitin ligases, which are components of the ubiquitin-degradation pathway, and determining whether such inhibitors restore p53 levels and function in tumor cells. This strategy will provide a framework for developing small molecule inhibitors of other E3 ubiquitin ligases with relevance to cancer and neurodegeneration. Current projects include synthesizing and screening biased chemical libraries of potential E3 ubiquitin ligase inhibitors.
Probing Neurodegeneration with the Tools of Chemical Biology
Huntington’s Disease
Huntington’s Disease is a neurodegenerative disorder caused by a trinucleotide (CAG) repeat expansion in the huntingtin gene. We have developed two high-throughput screens for small molecule suppressors of the toxicity of mutant huntingtin in neuronal cell lines. We tested 46,500 compounds in these assays and identified ~10 compounds that prevent selectively the death of neuronal cells expressing mutant huntingtin. These compounds will reveal new molecular mechanisms underlying huntingtin-induced cellular toxicity. Current projects include (i) creating affinity reagents based on these compounds to identify their cellular binding proteins, (ii) using microarray-based expression profiling to define the global response of neuronal cells to these compounds and (iii) testing the activity of these compounds in organismal models of triplet repeat diseases.
Spinal Muscular Atrophy
Spinal muscular atrophy (SMA) is an autosomal recessive disease caused by degeneration of a-motor neurons in the spinal cord anterior horn, leading to progressive muscular atrophy, paralysis, respiratory failure and infant death. SMA is caused by deletion of the survival motor neuron 1 (SMN1) gene, which encodes the SMN protein. Humans have two copies of the SMN gene (SMN1 and SMN2) that are located in a 500-kilobase (kb) inverted repeat on chromosome 5q13. In SMA patients, the SMN1 gene is deleted entirely and SMN2 is spliced such that only ~30% of mRNAs are full length, and ~70% of spliced transcripts exclude exon 7. The SMN2 gene product is sufficient to maintain fetal development, but affected individuals manifest the symptoms of SMA early in life and typically die as infants. We are developing screens for compounds that increase the amount of SMN protein in cells. Current projects include (i) developing a high-throughput assay for SMN protein level in cells to screen for compounds that directly stabilize this protein and (ii) determining the mechanisms regulating SMN turnover in cells.
Developing New Technologies and Informatic Systems
Creating Fluorescent Sensors to Monitor Protein Ligation Events in Vivo
Protein ligation involves the covalent coupling of small ubiquitin-like proteins (UBLs) to larger target proteins via the enzymatic action of E3 protein ligases. Such covalent modifications have been shown to regulate the stability and activity of target proteins, most notably by regulating proteasome-mediated protein degradation of polyubiquitylated proteins. There is a need to develop fluorescent sensors that indicate the extent to which specific proteins have been modified by ligation to UBLs. There is a further need to determine global protein ligation events that occur in response to introducing to or removing from cells these critical disease-related proteins. Current projects include creating fluorescent sensors that reveal the extent and identity of protein ligation events in vivo.
Creating an Informatics Management System for Chemical Biology
A critical emerging issue for academic laboratories is information management. This is especially true for laboratories that perform high-throughput experiments by testing large numbers of compounds or other reagents in microarrays or microtiter plates. Our incipient software suite, which we call Stand-alone Laboratory Information Management System (SLIMS), is being developed to facilitate data management and analysis and to create a publishable repository for such varied data sources as compound-based biological assays, gene expression analysis, protein-small molecule interactions and images. Current projects include the creation of software tools that will be integrated into SLIMS, an informatics system that we will make freely available once completed.
Research Resources
Two non-technical summaries of research in the laboratory:
Industrialized Screening for Cancer
Unlocking the Mysteries of Protein Function
