the Roy lab is working towards discovering new drug-like
compounds that could be used
to treat human disease
we are developing C. elegans as a
model for human diseases related to
vascular malfunctions, and
we are also using the non-parasitic
C. elegans as a model of parasitic nematodes, which infect over
one third of humans
we use these models to identify drug-like compounds that ameliorate the defects, and partner with other labs to test our compounds in corresponding disease models in fish and mice
In 1974, Sydney Brenner published his seminal description of C. elegans as a genetic model system. In it, he described his preliminary investigation into C. elegans’ sensitivity to small molecules. Only 2 of the ~100 bioactive molecules he tested exhibited activity. Consequently, no one has championed the worm as a model system for the discovery of novel bioactive molecules that could be developed as biological reagents or potential drug leads. My group’s work over the past 10 years, however, has challenged this general view by demonstrating that the worm has heretofore untapped potential to serve as a powerful platform with which to identify and characterize new small molecule tools.
|To identify new small molecule tools for the biological analysis of C. elegans, my lab has screened over 70,000 small molecules for the induction of a variety of phenotypes, including those that resemble the disruption of conserved components and pathways. We have found over 600 potent bioactive molecules. Our first detailed characterization was of one that we call nemadipine (pronounced ne-ma-dee-peen) and discovered that it is a new L-type calcium channel antagonist that is active in both worms and vertebrates. Nemadipine has unique properties compared to other FDA-approved calcium channel antagonists in that it is effective in whole living worms. Thus, nemadipine allows us to investigate the genetics of how animals interact with this important class of calcium channel antagonists for this first time. We published this work in Nature (Kwok et al., 2006), and have two follow-up papers on the screening method (Burns et al., 2006. Nature Protocols) and the use of nemadipine to study calcium channels (Kwok et al., 2008. PLoS Genetics).|
|The second molecule that we characterized, called dafadine (pronounced daf-a-deen) extends lifespan. Dafadine induces a highly penetrant dauer phenotype in C. elegans. Dauer is a stress-resistant alternative third larval stage that is engaged in response to stress. The genetic analysis of dauer engagement has led to many fundamental insights, including the discovery that the insulin pathway, which is one of two key pathways that controls dauer formation, negatively regulates lifespan. Through a series of phenotypic, chemical-genetic, cell-based, and biochemical experiments, we demonstrated that the DAF-9 cytochrome P450, which functions within the insulin pathway, is the direct and physiological target of dafadine.|
We also found that dafadine can also antagonize DAF-9’s human ortholog, called CYP27A1. Because of the insulin pathway’s role in regulating lifespan, we tested whether dafadine could extend the life span of C. elegans, and found that it did so by inhibiting DAF-9 activity (Luciani et al., 2011. Nature Chemical Biology). Since publication, both nemadipine and dafadine have been commercially available and are being used by several others in the community as research tools. We are in the process of characterizing several other molecules of interest, including pipersma, HBAC, migrazole, and mezole.
|Nematode parasites exist for nearly every agriculturally important animal and plant on earth, and currently infect over 2 billion people. Resistance to existing anthelmintics is rapidly growing and the need to develop novel anthelmintics is pressing. To identify potential anthelmintic leads, we screened the 600+ bioactive molecules that we have identified using C. elegans in the bovine parasitic nematode Cooperia oncophora in collaboration with John Gilleard (Calgary). C. oncophora also serves as a model for several nematode parasites of humans. We counter-screened our bioactive molecules in preliminary models of parasitic hosts, including zebrafish and a human cell line.|
We discovered 73 compounds that: i) kill both C. elegans and C. oncophora; ii) fail to induce obvious phenotypes in fish or HEK293 cells; and iii) have no obvious structural relationship to any characterized anthelmintic. Our work now focuses on exploiting C. elegans genetics and whole-genome sequencing to identify the target of ~45 of these molecules. With Prof. Gilleard, we will further investigate the anthelmintic potential of the most potent molecules that we find to have distinct mechanisms of action. At the very least, this project will yield over 20 new small molecule tools with defined targets that will enable future discoveries. We are optimistic, however, that we can develop many of these compounds into new anthelmintic drug leads for the benefit of human health.
The Roy lab is currently developing & screening models for diseases related to:
defects in CNS vasculature
the guided migration of cells and axons is essential for the proper development of all animals, including humans
the Roy Lab identifies new components
required for the guided migration of
cells and axons by studying membrane
extensions from both neurons and muscles
of the C. elegans nematode
through the genetic analyses of these
processes, the Roy lab has discovered a
novel guidance cue called MADD-4 that
signals through an UNC-40/DCC-EVA-1
the Roy lab has also discovered a novel
adaptor protein of the UNC-40/DCC
receptor called MADD-2, which is also
conserved with humans
Guided cell migration is an essential process that is used repeatedly throughout animal development. For example, without the ability to find their respective targets, neural crest cells could not travel from the dorsal midline to generate specific cranial structures near the ventral midline, and axons could not make stereotypical connections to other neurons and muscles. In addition, cancer depends on cell migration for the vascularization and metastasis of tumors.
By the time Peter started his own lab in 2002, neurobiologists had already made leaps and bounds in our understanding of guided cell migration in the previous decade. However, he hypothesized that there had to be more ‘global’ conserved cues and pathways that guide the migration of cells and cell extensions during embryonic development than just the netrins, semaphorins, slits and ephrins. This idea was based on the fact that for several migrations, the four major cues were unlikely players in the process.
To identify new genes required for guided cell migration, the Roy lab pioneered the study of specialized extensions from muscles, called muscle arms, which are guided to the motor axons of the model nematode Caenorhabditis elegans. Muscle arm termini harbor the post-synaptic machinery of the neuromuscular junction and are therefore essential for proper neuromuscular development. Our paper, “The Characterization of Muscle Arm Development in Caenorhabditis elegans” (Dixon & Roy, Development, 2005), was the first detailed description of muscle arm development and is recognized as a seminal piece of work.
the discovery of MADD-4
In the hopes of identifying novel and conserved guidance components, we screened for mutants that have disrupted muscle arm extension to their neuronal targets and identified 18 different complementation groups (i.e., genes) that are necessary for this process. An important initial finding was that the conserved guidance receptor UNC-40 is a key element in directing muscle arm extension to the motor neurons. We found that UNC-40 functions in muscles and likely signals through the UNC-73 RhoGEF and members of the WAVE complex of actin modulators to direct arm extension. To our surprise, the canonical ligand of UNC-40, called UNC-6/netrin was dispensable for muscle arm extension, which implied the existence of a novel guidance cue.
As part of our forward genetic screen, we isolated three alleles of a gene we call madd-4. MADD-4 has all the hallmarks of a muscle arm chemoattractant. It is likely secreted from the motor neurons to elicit muscle arm extension, and can redirect muscle arms and sensory axons when ectopically expressed. Furthermore, the ability to redirect muscle arms is dependent on UNC-40, suggesting that MADD-4 likely signals through UNC-40 to attract cell extensions. MADD-4 is orthologous to the human non-enzymatic proteins ADAMTSL-1 and 3. Despite its conservation among animals, the biological role any MADD-4 ortholog beyond C. elegans remains a mystery.
More recently, we have developed a deeper understanding of the mechanism of action of MADD-4 in that it physically interacts with not only UNC-40, but to a greater extent, a co-receptor called EVA-1. Our genetic analyses indicate that EVA-1 functions as an UNC-40 co-receptor for the MADD-4 cue. EVA-1 allows MADD-4 to outcompete UNC-6 for UNC-40.
the discovery of MADD-2
In our screen for genes required for muscle arm guidance, we also recovered 12 alleles of gene that we call madd-2. MADD-2 is not only required for muscle arm extension, but for the guidance of many cell extensions towards the ventral midline. We discovered that madd-2 functions cell-autonomously at the leading edge of the cell, and likely functions as a scaffold to facilitate a physical interaction between the UNC-40 receptor and the UNC-73 RhoGEF. MADD-2 is a homolog of a human gene called MID1. The disruption of MID1 causes Opitz Syndrome, which is characterized by numerous congenital defects along the ventral midline. Despite being causative in the disease, it is currently unclear how MID1 mutations lead to Opitz Syndrome. However, given the analogous biological roles of MADD-2 and MID1, we speculate that MID1 may regulate an UNC-40-like pathway in cells that extend towards the ventral midline in humans, and that mis-regulation of this pathway may be at the root of Opitz Syndrome pathology. We published this story in Developmental Cell (Alexander et al., 2010).
The Roy lab is interested in:
better understanding the guidance capabilities of MADD-4
better understanding the mechanism of action of MADD-4
understanding the role of several new genes required for guided membrane extension that we uncovered in forward genetic screens
better understanding the guidance of the mechanosensory axons