Research

Lili has been performing research on cancer biology and neurodegenerative diseases. Below are some of the main projects that she has worked on.

 

Protein Quality Control System that Protects Against Neurodegenerative Diseases

During her Ph.D. research, Lili explored the connection between cancer signaling pathway and protein quality control system for neurodegenerative diseases. Protein is essential for all forms of lives. All proteins need to be folded into a correct structure to become functional. Proteins that fail to form correct structure, namely misfolded proteins, are not only dysfunctional, but also toxic to cells. Misfolded protein are the underlying cause to many neurodegenerative diseases including Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis (ALS), Huntington’s disease (HD), and spinocerebellar ataxias (SCA). In the case of polyglutamine diseases such as Huntington’s disease (HD) and spinocerebellar ataxias (SCA), the mutated gene contains a repeat of the CAG nucleotide triplet. C, A and G represent three out of four different nucleotides in DNA strands, which encode amino acid glutamine. A repeat of CAG results in a polyglutamine tract. The polyglutamine-tract proteins are deleterious and may cause the death of neurons. Lili used polyglutamine proteins as major misfolded protein models in her research.

 

Other_CAG the death chain_web

CAG: The death chain
This work was created in 2010 and exhibited at the Philadelphia Science Festival at 2011. A neuron cell is being choked by an iron chain. The chain is made of the letters “CAG”, which represents metaphorically the cause of neuron cell damage in Huntington’s disease the CAG repeat in the mutated gene. The green blob inside the neuron represents the protein aggregate formed by disease-associated proteins, which are commonly seen in neurodegenerative diseases.

Healthy cells have defensive mechanisms to remove aberrant proteins. However, these mechanisms are poorly understood. It is also unclear why some misfolded proteins can escape the system and cause diseases. For this project, Lili identified a novel cellular protein quality control mechanism that recognizes and removes many types of toxic misfolded proteins. The key players in this system are promyelocytic protein (PML), a protein identified initially in promyelocytic leukemia, and another protein, RNF4. Lili showed that PML protein has a special region that can recognize and bind to misfolded proteins. Lili demonstrated the activity of PML to label misfolded proteins with a small protein tag called SUMO (small ubiquitin-like modifier). SUMO modified misfolded proteins are handed off to RNF4, which places a second tag called ubiquitin to the misfolded proteins. Ubiquitin is a signal recognized by a protein degradation machinery, proteasome. As a result of the tagging relay, the misfolded proteins are degraded by proteasome. Using cell culture models, Lili have showed that this system can effectively remove pathogenic proteins that cause amyotrophic lateral sclerosis (ALS), Huntington’s disease (HD), and spinocerebellar ataxias (SCA). This work was published on Molecular Cell, 2014.

 

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Protein Quality Control Relay
Lili’s research on protein quality control system was highlighted as the cover of Molecular Cell, July, 2014. Cellular protein quality control system comprised of PML (pink) and RNF4 (purple) proteins. This system selects misfolded proteins (deformed boxes), tag them with degradation signals (red and white labels), and send them for destruction by proteasome (green). The red tag represents SUMO and the white tag represents ubiquitin.

 

Mechanism of the Anti-Cancer Drug Lonidamine

One of the projects for Lili’s postdoc research is the mechanisms of anti-cancer drugs, especially those targeting cancer cell metabolism. Cancer cell metabolism is very different from normal cell metabolism. These special metabolisms are normally featured with high demand of nutrients in order to provide energy to support the rapid growth and proliferation of cancer cells. Therefore, cancer cell metabolic pathways can be potentially targeted for anti-cancer treatment. The recent research on preclinical and clinical research has further shed light on its future clinical application. Lonidamine (1-(2,4-dichlorobenzyl)-1H-indazole-3-carboxylic acid) is an anti-cancer drug known for its ability to inhibit cancer cell metabolism. It has been used in many clinical trials and can effectively improve the efficacy of other anti-cancer agents. However, the mechanism by which that Lonidamine induces cancer cell death was unclear. For this project, Lili discovered that Lonidamine targets respiratory complex II, an essential enzyme complex in mitochondria where the biochemical process of respiration and energy production occur. Lili further defined the exact targeting site of Lonidamine on complex II. The inhibition of complex II by Lonidamine markedly altered several essential metabolic pathways in cancer cells. It also led to production of large amount of reactive oxygen species (ROS) through complex II. The high levels of ROS induced by Lonidamine are toxic to cancer cells, which significantly contributes to its cancer-cell killing activity. This work was published on Journal of Biological Chemistry, 2016.

 

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Anti-Cancer Drug and Mitochondrial Respiration
Lili’s research on the mechanism of lonidamine was highlighted as the cover of Journal of Chemical Biology, January, 2016. The image shows the antitumor agent lonidamine (yellow) inhibits the succinate-ubiquinone reductase activity of complex II (orange) on the mitochondria membrane. . Lonidamine induces ROS generation through complex II and alters glutamine metabolism. The blue and green molecules are mitochondrial complex I and III.

Development of LC-MS methods

Dr. Blair’s lab is specialized in Liquid chromatography–mass spectrometry (LC-MS) and one of Lili’s project is to develop LC-MS methods for metabolism and biomarker analysis. LC-MS is an analytical chemistry technique which combines the separating power of liquid chromatography (LC) and the mass detection power of mass spectrometry (MS). LC-MS has been widely used for detection and identification of chemicals in many fields including pharmaceuticals, environment, foods, and industrial materials. One of an important application of LC-MS in the preclinical and clinical research is metabolic analysis. Recently, the successes of metabolic biomarkers and anti-cancer drugs that target certain metabolic pathways have drawn great attention from scientists and general public. To further promote the research in metabolic field, it is urgent to develop even more sensitive and accurate LC-MS methodology. Many metabolites of low molecular weight are especially challenging for LC-MS analysis. In order to develop a LC-MS method to overcome this challenge, we creatively added a highly volatile reagent, hexafluoroisopropanol (HFIP), in the LC solvent. Upon the separation of LC, one key factor for effective MS analysis is to convert the chemicals for analysis to charged molecules. When HFIP which is positively charged in the solvent rapidly evaporates at the interface between LC and MS, many metabolites left in the solvent droplet are able to gain negative charge effectively. As a result, the sensitivity of MS was boosted up to 10-fold for many metabolites for analysis. In addition, HFIP also enhances the LC performance. We have shown that our improved method is highly sensitive and can be applied to analysis to a wide range of metabolic analysis for cells, tissues or patient samples. In addition to develop LC-MS method for metabolic analysis, Lili is now working on develop LC-MS method for analyzing protein biomarkers from biological samples.

 

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HFIP enhances ionization at the MS source
Lili’s work on LC-MS method development was highlighted as the cover of Rapid Communication of Mass Spectrometry, 2016. The image shows hexafluoroisopropanol (HFIP) enhances negative ionization of metabolite molecules at the MS ESI source.

 

Representative Collaborative Projects:

tRNAs in cell death

Ribonucleic acid (RNA) is one of the three major biological molecules (along with DNA and proteins) that are essential for the function of cells. tRNAs (transfer RNA) play a very important role in protein synthesis. In 2010, we discovered an alternative function for tRNAs in apoptosis (the process of programmed cell death). We demonstrated that tRNAs directly bind to the protein cytochrome c and inhibit cytochrome c-initiated apoptosome formation, which initiates apoptosis. This was a collaborative project between Dr. Xiaolu Yang and Dr. Gideon Dreyfuss.

 

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tRNAs in cell death
Lili’s research on protein quality control system was highlighted as the cover of Molecular Cell, July, 2014. Cellular protein quality control system comprised of PML (pink) and RNF4 (purple) proteins. This system selects misfolded proteins (deformed boxes), tag them with degradation signals (red and white labels), and send them for destruction by proteasome (green). The red tag represents SUMO and the white tag represents ubiquitin.

 

Regulation of T cell metabolism in cancer immunotherapy

Recently cancer immunotherapy has become a potential game-changer for treating many types of cancer. Among different immunotherapy strategies, one of the promising one is the adoptive T cell therapy using infusion of genetically redirected autologous T cells – the chimeric antigen receptor (CAR) T cells. We explored the effect of signaling domains of coreceptors CD28 and 4-1BB on the metabolic characteristics of human therapeutic CAR T cells. The regulation of metabolism in T cells is critical for the control of their expansion, exhaustion and memory formation. We discovered that 4-1BB in the CAR architecture promoted the formation of CD8+ central memory T cells. Biochemical assays and mass spectrometry (MS) analysis revealed that cells with 4-1BB domain developed significantly elevated respiratory capacity, increased fatty acid oxidation and enhanced mitochondrial biogenesis. In contrast, CD28 domains in the CAR architecture led to outgrowth of effector memory cells. These cells showed enhanced glycolytic metabolism. Our study has provided mechanistic insights into the regulation of CAR-T cell fates by different signaling domains through metabolism. Importantly, it also provided potential means for designing engineered T cells towards different fates for therapeutic purposes. This was a collaborative project between Dr. Blair, Dr. Milone and Dr. June. The paper was published on Immunity, 2016.

 

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Regulation of CAR T cell metabolism
This research was highlighted as the cover of Immunity, February, 2016. T cells (yellow) expressing CARs (brown wrench) engaging antigen-bearing tumor cells (bottom left). The image depicts the activation of CAR, which regulates T cell metabolism through mitochondrial adaptations.