The PLRC Grant Review committee has selected the awardees for the 2021-2022 Pilot and Feasibility Program grants. Our thanks to all who submitted proposals.
Congratulations to the following awardees of the 2021 PLRC Pilot and Feasibility Program grants:
Amanda Maree Clark, PhD – Research Assistant Professor, Department of Pathology, Division of Medicine
“Exosomes from hepatic non-parenchymal cells promote dormancy and therapeutic resistance of liver metastases”
Distant metastasis is the major cause of cancer-related mortality. Dauntingly, it can emerge years to decades after a seeming ‘cure’ of the primary tumor. The most common site of metastasis for solid tumors (excluding lymph nodes) is the liver and its involvement correlates poorly with patient survival. Metastasis begins with cells within the primary tumor undergoing a cancer-associated epithelial-tomesenchymal
transition (cEMT), which enables motility to intravasate into the circulation followed by extravasation into distant organs. At the metastatic organ (i.e. the liver), the tumor cells again change behavior to survive, in part by undergoing a partial cancer-associated mesenchymal-to-epithelial reverting transition (cMErT), and a proportion may enter a dormant state. Such tumor cells may remain in this state for years before forming clinically detectable metastases. The mechanisms as to why some tumor cells initially enter a dormant state at metastatic sites instead of outgrowing immediately remains unclear.
Evidence is emerging that cells and cues within the metastatic microenvironment are critical regulators of dormancy, with the immune and stromal systems being the predominant players in these interactions. Within the liver, the relevant candidate cells are collectively termed non-parenchymal cells (NPCs); principal subtypes include liver sinusoidal endothelial cells (LSEC), Kupffer cells (KC), and hepatic stellate cells (HStC). The role activated NPCs play in promoting the outgrowth of metastatic tumor cells in the liver is well characterized. However, more recent preliminary data from our lab found that inclusion of quiescent NPCs in the hepatic
niche was associated with reduced outgrowth of metastatic tumor cells and that their exosomes induced Ecadherin expression, a prototypical marker of cMErT and promoter of therapeutic resistance. Consequently, this leads us to hypothesize that exosomes from quiescent NPCs (NPCexo) regulate dormancy and impart therapeutic resistance in metastatic tumor cells upon colonizing the liver. This hypothesis will be examined by the following Aims which seek to build upon the preliminary findings and dissect the individual contributions of NPC subtypes. Discerning the key liver NPC subtype(s) involved will help improve our understanding of the mechanisms promoting dormancy and also help devise strategies to maintain them in a quiescent, suppressed state.
Hamza Obaid Yazdani, MD – Research Associate Professor, Department of Surgery, Division of Medicine
“Role of Exercise Training in Preconditioning the Liver against Ischemia/ Reperfusion Injury in an Orthotopic Murine Liver Transplant Model”
Orthotopic liver transplantation (OLT) is the gold standard of care in patients with end-stage liver disease and many patients with liver tumors. Hepatic ischemia/reperfusion injury (IRI), an innate immune-driven inflammatory response, is a major obstacle to the success of OLT. During liver transplantation, the liver undergoes at least two serious insults- a period of cold ischemic storage—followed by reperfusion and a warm ischemia injury1,2. During both warm I/R and cold storage, stressed and dying hepatocytes in donor grafts undergo metabolic reprogramming that skew the cells towards aerobic glycolysis and release of cytokines and danger-associated molecular patterns (DAMPs). DAMPs, such as HMGB1 and Histone H3, can, upon organ transplantation, activate inflammatory networks in the recipient which may contribute to poor early graft function, and may predispose the graft to acute and/or chronic rejection3,4. Although significant progress has been made in understanding the hepatic response to cold and warm ischemia and reperfusion, translational applications have been slow to emerge. The few agents found to be protective against liver IRI in animal models, have shown limited success when tested in randomized trials in human transplants. Thus, new therapeutic strategies to minimize IRI are warranted to improve outcomes. Exercise training (ExT) confers a sustainable protection against IRI in several animal models of ischemia, such as cardiac or cerebral ischemia, and has been associated with improved cardiovascular outcomes in patients with solid organ transplants5,6. We have recently shown that chronic ExT, in a murine model of warm liver IRI, significantly decreased liver injury, a protection associated by suppression of activated liver sinusoidal endothelial cells (LSECs), marked reduction of hepatic cell death and the appearance of a distinct population of infiltrating macrophages expressing an M2 phenotype, thereby altering the complex inflammatory chain initiated by the invasion of the IRI liver by neutrophils and neutrophil extracellular traps (NETs). Indeed, ExT rendered mouse neutrophils less capable of Netosis7. Thus, the chain of liver damaging inflammatory events accompanying liver I/R can be modified by a prior course of ExT. The fact that one can reduce the inflammatory changes of liver I/R by ExT, suggests that livers from ExT mice may be resistant to damage during cold and/or warm ischemia during transplantation. Detailed mechanistic study of damage and its alleviation by ExT may lead to better hepatic preservation.
Shuchang (Silvia) Liu, PhD – Assistant Professor, Department of Pathology, Division of Medicine
“Development of bioinformatics pipeline to detect fusion transcripts in liver cancer samples via long-read sequencing technology”
Hepatocellular carcinoma (HCC) is among the top six major malignancies worldwide. It accounts for approximately 80% of all liver cancer cases and approximately 500,000 new cases are diagnosed annually. However, the underlying mechanism of HCC pathogenesis is complex and still not fully understood.
Fusion transcripts are fused RNAs that can either translate into chimeric proteins or alter the gene expression. Studies show that fusion transcripts have high concurrence rate in HCC samples and are closely correlated with cancer recurrence. Genes composing fusion transcripts are mostly oncogenes that can act as biomarkers for cancer prediction and therapeutic targets. To identify fusion transcripts, traditional short-read (ranging 50-200bp) sequencing technology requires extremely high coverage to detect fusion junction points and to predict transcript isoforms. To conquer the obstacle, long-read sequencing technologies are developed to generate reads with length of several-thousand base pairs. However, it brings challenges for the bioinformatic analysis to detect fusion transcripts on long-read data.
Our proposal aims to develop a bioinformatics pipeline specific for long-read sequencing data to detect fusion transcripts/ isoforms and predict the corresponding open reading frames. We will also evaluate the pipeline performance by both public cancer data and in-silico simulation data (Aim 1). In the next step, we will apply the developed software into HCC patient tumor samples to detect fusion transcripts. The identified fusions will be validated by experimental methods and annotated by liver tissue-specific database for biological function interpretation (Aim 2). The pilot data generated by this proposal will aim for peer-reviewed journal publication and future R01 grant application.
Bharat Bhushan, PhD – Research Assistant Professor, Department of Pathology
“Epidermal growth factor receptor in cell death signaling during acute liver injury and its therapeutic implications”
Acetaminophen (APAP) is one of the most commonly used drug across the world and APAP overdose is the topmost cause of acute liver failure (ALF) in the United States. N-acetylcysteine (NAC) is the only pharmacological treatment for APAP overdose and is effective only in early presenting patients. There is an urgent need to explore alternative pharmacological approaches which are effective even at a later stage as majority of the patients seek medical attention late. Epidermal growth factor receptor (EGFR) is long known to be crucial for hepatocyte proliferation and its proliferative roles have been widely studied utilizing partial hepatectomy model (in healthy liver). However, ours and other previous studies have shown that the microenvironment of the injured liver differ remarkably from normal liver and the role of EGFR specifically in liver injury models have not been extensively explored. Apart from the conventional role of EGFR in regulating hepatocyte proliferation, studies indicate its potential role in modulating mechanisms of liver injury as well. In the presence of toxic insults to hepatocytes in culture, sustained activation of EGFR can also promote hepatocyte death. In alignment with these reports, our previously published study and strong preliminary studies indicate a novel role of EGFR in promoting liver injury in the clinical relevant murine model of ALF (i.e. APAP overdose). Our studies show that APAP overdose in mice causes sustained and dose-dependent activation of EGFR. Surprisingly, strong EGFR activation preceded necrosis in APAP-treated mice and primary human hepatocytes, and the extent of EGFR activation correlated with the severity of liver injury. Further, treatment with EGFR inhibitor canertinib after non-lethal APAP overdose, almost completely abolished APAP-induced liver injury (AILI). Most importantly, canertinib treatment in combination with NAC significantly decreased AILI even after late intervention, when NAC alone was ineffective. Lastly, in our preliminary study, a single dose of afatinib, a clinically-approved EGFR inhibitor with less toxicity, also significantly attenuated AILI and even improved liver regeneration secondary to remarkable decrease in liver injury. Based on these strong preliminary findings, the proposed specific aim will comprehensively investigate the explicit role of EGFR in AILI, utilizing hepatocyte-specific EGFR deletion strategy in adult mice. Moreover, our studies will explore viability of EGFR inhibition as a therapeutic strategy for AILI utilizing a clinically-approved inhibitor, afatinib. Lastly, our studies will investigate how EGFR is rapidly activated (even preceding hepatocyte death) during AILI and further explore the signaling mechanisms how EGFR regulates acute liver injury. Overall, these studies will provide insights on novel aspect of EGFR biology in regulating acute liver injury, utilizing a clinically significant ALF model. Moreover, successful completion of these studies will also be useful to determine potential of repurposing a clinically available EGFR inhibitor (afatinib) to treat APAP-induced ALF.