Honors & Awards
Poster Prize, Gordon Research Conference in Glycobiology (2019)
Banting Postdoctoral Fellowship, Canadian Institutes of Health Research (2018-2020)
NIH F32 Ruth L. Kirschstein National Research Service Award (NRSA) (declined), NIGMS (2018)
Sela Cheifetz Centennial Award for Excellence in Graduate Studies, Biochemistry Department, University of Toronto (2017)
First Place Poster Prize, University of Toronto, Biochemistry Department Research Symposium (2015)
First Place Poster Prize, CIHR EIRR21 Training Program Research Symposium (2015)
Vanier Canada Graduate Scholarship, Canadian Institutes of Health Research (2014-2017)
Excellence in Radiation Research for the 21st century (EIRR21) Alumni Award, Canadian Institutes of Health Research/Terry Fox Foundation (2014-2015)
Banting & Best Canada Graduate Scholarship (declined), Canadian Institutes of Health Research (2014)
Excellence in Radiation Research for the 21st century (EIRR21) training award, Canadian Institutes of Health Research/Terry Fox Foundation (2013-2014)
Honour Roll, McGill University (2012)
Undergraduate Student Research Award, National Sciences and Engineering Research Council of Canada (2011)
Bachelor of Science, McGill University (2012)
Doctor of Philosophy, University of Toronto (2017)
DNA Polymerase θ Increases Mutational Rates in Mitochondrial DNA.
ACS chemical biology
2018; 13 (4): 900–908
Replication and maintenance of mitochondrial DNA (mtDNA) is essential for cellular function, yet few DNA polymerases are known to function in mitochondria. Here, we conclusively demonstrate that DNA polymerase θ (Polθ) localizes to mitochondria and explore whether this protein is overexpressed in patient-derived cells and tumors. Polθ appears to play an important role in facilitating mtDNA replication under conditions of oxidative stress, and this error-prone polymerase was found to introduce mutations into mtDNA. In patient-derived cells bearing a pathogenic mtDNA mutation, Polθ expression levels were increased, indicating that the oxidative conditions in these cells promote higher expression levels for Polθ. Heightened Polθ expression levels were also associated with elevated mtDNA mutation rates in a selected panel of human tumor tissues, suggesting that this protein can influence mutational frequencies in tumors. The results reported indicate that the mitochondrial function of Polθ may have relevance to human disease.
View details for DOI 10.1021/acschembio.8b00072
View details for PubMedID 29509408
View details for PubMedCentralID PMC5914477
Characterization of Trypanosoma cruzi MutY DNA glycosylase ortholog and its role in oxidative stress response.
Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases
2017; 55: 332–42
Trypanosoma cruzi is a protozoan parasite and the causative agent of Chagas disease. Like most living organisms, it is susceptible to oxidative stress, and must adapt to distinct environments. Hence, DNA repair is essential for its survival and the persistence of infection. Therefore, we studied whether T. cruzi has a homolog counterpart of the MutY enzyme (TcMYH), important in the DNA Base Excision Repair (BER) mechanism. Analysis of T. cruzi genome database showed that this parasite has a putative MutY DNA glycosylase sequence. We performed heterologous complementation assays using this genomic sequence. TcMYH complemented the Escherichia coli MutY- strain, reducing the mutation rate to a level similar to wild type. In in vitro assays, TcMYH was able to remove an adenine that was opposite to 8-oxoguanine. We have also constructed a T. cruzi lineage that overexpresses MYH. Although in standard conditions this lineage has similar growth to control cells, the overexpressor is more sensitive to hydrogen peroxide and glucose oxidase than the control, probably due to accumulation of AP sites in its DNA. Localization experiments with GFP-fused TcMYH showed this enzyme is present in both nucleus and mitochondrion. QPCR and MtOX results reinforce the presence and function of TcMYH in these two organelles. Our data suggest T. cruzi has a functional MYH DNA glycosylase, which participates in nuclear and mitochondrial DNA Base Excision Repair.
View details for DOI 10.1016/j.meegid.2017.09.030
View details for PubMedID 28970112
Mitochondria-penetrating peptides conjugated to desferrioxamine as chelators for mitochondrial labile iron.
2017; 12 (2): e0171729
Desferrioxamine (DFO) is a bacterial siderophore with a high affinity for iron, but low cell penetration. As part of our ongoing project focused on DFO-conjugates, we synthesized, purified, characterized and studied new mtDFOs (DFO conjugated to the Mitochondria Penetrating Peptides TAT49-57, 1A, SS02 and SS20) using a succinic linker. These new conjugates retained their strong iron binding ability and antioxidant capacity. They were relatively non toxic to A2780 cells (IC50 40-100 μM) and had good mitochondrial localization (Rr +0.45 -+0.68) as observed when labeled with carboxy-tetramethylrhodamine (TAMRA) In general, mtDFO caused only modest levels of mitochondrial DNA (mtDNA) damage. DFO-SS02 retained the antioxidant ability of the parent peptide, shown by the inhibition of mitochondrial superoxide formation. None of the compounds displayed cell cycle arrest or enhanced apoptosis. Taken together, these results indicate that mtDFO could be promising compounds for amelioration of the disease symptoms of iron overload in mitochondria.
View details for DOI 10.1371/journal.pone.0171729
View details for PubMedID 28178347
View details for PubMedCentralID PMC5298241
Mitochondrial DNA repair and replication proteins revealed by targeted chemical probes.
Nature chemical biology
2016; 12 (7): 567–73
Efficient and accurate replication and repair of mitochondrial DNA is essential for cellular viability, yet only a minimal complement of mitochondrial proteins with relevant activities have been identified. Here, we describe an approach to screen for new pathways involved in the maintenance of mitochondrial DNA (mtDNA) that leverages the activities of DNA-damaging probes exhibiting specific subcellular localization. By conducting a siRNA screen of known nuclear DNA maintenance factors, and monitoring synergistic effects of gene depletion on the activity of mitochondria-specific DNA-damaging agents, we identify a series of proteins not previously recognized to act within mitochondria. These include proteins that function in pathways of oxidative DNA damage repair and dsDNA break repair, along with a novel mitochondrial DNA polymerase, POLθ, that facilitates efficient DNA replication in an environment prone to oxidative stress. POLθ expression levels affect the mutational rate of mitochondrial DNA, but this protein also appears critical for efficient mtDNA replication.
View details for DOI 10.1038/nchembio.2102
View details for PubMedID 27239789
Mitochondrial Chemical Biology: New Probes Elucidate the Secrets of the Powerhouse of the Cell.
Cell chemical biology
2016; 23 (8): 917–27
Mitochondria are energy-producing organelles with essential functions in cell biology, and mitochondrial dysfunction is linked to a wide range of human diseases. Efforts to better understand mitochondrial biology have been limited by the lack of tools for manipulating and detecting processes occurring within the organelle. Here, we highlight recent significant advances in mitochondrial chemical biology that have produced new tools and techniques for studying mitochondria. Specifically, we focus on the development of chemical tools to perturb mitochondrial biochemistry, probes allowing precise measurement of mitochondrial function, and new techniques for high-throughput characterization of the mitochondrial proteome. Taken together, these advances in chemical biology will enable exciting new directions in mitochondrial research.
View details for DOI 10.1016/j.chembiol.2016.06.012
View details for PubMedID 27478157
Peptide-Mediated Delivery of Chemical Probes and Therapeutics to Mitochondria.
Accounts of chemical research
2016; 49 (9): 1893–1902
Mitochondria are organelles with critical roles in key processes within eukaryotic cells, and their dysfunction is linked with numerous diseases including neurodegenerative disorders and cancer. Pharmacological manipulation of mitochondrial function is therefore important both for basic science research and eventually, clinical medicine. However, in comparison to other organelles, mitochondria are difficult to access due to their hydrophobic and dense double membrane system as well as their negative membrane potential. To tackle the challenge of targeting these important subcellular compartments, significant effort has been put forward to develop mitochondria-targeted systems capable of transporting bioactive cargo into the mitochondrial interior. Systems now exist that utilize small molecule, peptide, liposome, and nanoparticle-based transport. The vectors available vary in size and structure and can facilitate transport of a variety of compounds for mitochondrial delivery. Notably, peptide-based delivery scaffolds offer attractive features such as ease of synthesis, tunability, biocompatibility, and high uptake both in cellulo and in vivo. Owing to their simple and modular synthesis, these peptides are highly adaptable for delivering chemically diverse cargo. Key design features of mitochondria-targeted peptides include cationic charge, which allows them to harness the negative membrane potential of mitochondria, and lipophilicity, which permits favorable interaction with hydrophobic membranes of mitochondria. These peptides have been covalently tethered to target therapeutic agents, including anticancer drugs, to enhance their drug properties, and to provide probes for mitochondrial biology. Interestingly, mitochondria-targeted DNA damaging agents demonstrate high potency and the ability to evade resistance mechanisms and off-target effects. Moreover, a combination of mitochondria-targeted DNA damaging agents was applied to an siRNA screen for the elucidation of poorly understood mitochondrial DNA repair and replication pathways. In this work, a variety of novel proteins were identified that are essential for the maintenance of mitochondrial nucleic acids. Mitochondria-targeted peptides have also been used to increase the therapeutic window of antibacterial drugs with significant mammalian toxicity. Given the evolutionary similarity of mitochondria and bacteria, peptides are effective transporters that can target both of these entities. These antimicrobial peptides are highly effective even in difficult to target intracellular bacteria which reside within host cells. This peptide-based approach to targeting mitochondria has provided a variety of insights into the "druggability" of mitochondria and new biological processes that could be future drug targets. Nevertheless, the mitochondrial-targeting field is quite nascent and many exciting applications of organelle-specific conjugates remain to be explored. In this Account, we highlight the development and optimization of the mitochondria-penetrating peptides that our laboratory has developed, the unique applications of mitochondria-targeted bioactive cargo, and offer a perspective on important directions for the field.
View details for DOI 10.1021/acs.accounts.6b00277
View details for PubMedID 27529125
Molecular vehicles for mitochondrial chemical biology and drug delivery.
ACS chemical biology
2014; 9 (2): 323–33
The mitochondria within human cells play a major role in a variety of critical processes involved in cell survival and death. An understanding of mitochondrial involvement in various human diseases has generated an appreciable amount of interest in exploring this organelle as a potential drug target. As a result, a number of strategies to probe and combat mitochondria-associated diseases have emerged. Access to mitochondria-specific delivery vectors has allowed the study of biological processes within this intracellular compartment with a heightened level of specificity. In this review, we summarize the features of existing delivery vectors developed for targeting probes and therapeutics to this highly impermeable organelle. We also discuss the major applications of mitochondrial targeting of bioactive molecules, which include the detection and treatment of oxidative damage, combating bacterial infections, and the development of new therapeutic approaches for cancer. Future directions include the assessment of the therapeutic benefit achieved by mitochondrial targeting for treatment of disease in vivo. In addition, the availability of mitochondria-specific chemical probes will allow the elucidation of the details of biological processes that occur within this cellular compartment.
View details for DOI 10.1021/cb400821p
View details for PubMedID 24410267
Re-directing an alkylating agent to mitochondria alters drug target and cell death mechanism.
2013; 8 (4): e60253
We have successfully delivered a reactive alkylating agent, chlorambucil (Cbl), to the mitochondria of mammalian cells. Here, we characterize the mechanism of cell death for mitochondria-targeted chlorambucil (mt-Cbl) in vitro and assess its efficacy in a xenograft mouse model of leukemia. Using a ρ° cell model, we show that mt-Cbl toxicity is not dependent on mitochondrial DNA damage. We also illustrate that re-targeting Cbl to mitochondria results in a shift in the cell death mechanism from apoptosis to necrosis, and that this behavior is a general feature of mitochondria-targeted Cbl. Despite the change in cell death mechanisms, we show that mt-Cbl is still effective in vivo and has an improved pharmacokinetic profile compared to the parent drug. These findings illustrate that mitochondrial rerouting changes the site of action of Cbl and also alters the cell death mechanism drastically without compromising in vivo efficacy. Thus, mitochondrial delivery allows the exploitation of Cbl as a promiscuous mitochondrial protein inhibitor with promising therapeutic potential.
View details for DOI 10.1371/journal.pone.0060253
View details for PubMedID 23585833
View details for PubMedCentralID PMC3621862
Targeting mitochondrial DNA with a platinum-based anticancer agent.
Chemistry & biology
2013; 20 (11): 1323–28
An analog of the anticancer drug cisplatin (mtPt) was delivered to mitochondria of human cells using a peptide specifically targeting this organelle. mtPt induces apoptosis without damaging nuclear DNA, indicating that mtDNA damage is sufficient to mediate the activity of a platinum-based chemotherapeutic. This study demonstrates the specific delivery of a platinum drug to mitochondria and investigates the effects of directing this agent outside the nucleus.
View details for DOI 10.1016/j.chembiol.2013.08.010
View details for PubMedID 24183971
View details for PubMedCentralID PMC4082333
The 5' binding MID domain of human Argonaute2 tolerates chemically modified nucleotide analogues.
Nucleic acid therapeutics
2013; 23 (1): 81–87
Small interfering RNAs (siRNAs) can trigger potent gene silencing through the RNA interference (RNAi) pathway. The RNA-induced silencing complex (RISC) is key to this targeted mRNA degradation, and the human Argonaute2 (hAGO2) endonuclease component of RISC is responsible for the actual mRNA cleavage event. During RNAi, hAGO2 becomes loaded with the siRNA guide strand, making several key nucleic acid-enzyme interactions. Chemically modified siRNAs are now widely used in place of natural double-stranded RNAs, and understanding the effects chemical modifications have on guide strand-hAGO2 interactions has become particularly important. Here, interactions between the 5' nucleotide binding domain of hAGO2, MID, and chemically modified nucleotide analogues are investigated. Measured dissociation constants reveal that hAGO2 does not discriminate between nucleotide analogues during binding, regardless of the preferred sugar conformation of the nucleotide analogues. These results correlate well with cell-based gene silencing results employing siRNAs with 5'-modified guide strands. Additionally, chemical modification with 2'-deoxy-2'-fluoroarabino nucleic acid (2'F-ANA) and 2'-deoxy-2'-fluororibonucleic acid (2'F-RNA) at the passenger strand cleavage site of siRNAs has been shown to prevent hAGO2-mediated strand cleavage, an observation that appears to have little impact on overall gene silencing potency.
View details for DOI 10.1089/nat.2012.0393
View details for PubMedID 23289589