Mechanisms of Resistance to EGFR Inhibition in HNSCC

Head and neck squamous cell carcinoma (HNSCC) remains a deadly disease with poor prognosis. Developing novel, effective combination therapies have the potential to improve patient survival. However, advancing biomarkers in conjunction with combination therapy will also be essential for efficacy so as to match treatment to the patient. In my thesis, I investigated the hypothesis that co-targeting a specific mechanism of resistance with combination therapy would be more effective than either therapy alone. I focused on identifying resistance mechanisms to cisplatin and EGFR inhibition, which are common HNSCC treatments, in UM-SCC cell lines using CRISPR/Cas9 screening libraries. This approach identified genetic knockouts that sensitized cell lines to either cisplatin or EGFR inhibition. The results of a CRISPR/Cas9 screen nominated NOTCH pathway knockouts and specifically NOTCH1 knockouts as capable of sensitizing cisplatin-resistance cells. Further results suggest that the combination of Notch inhibitors and cisplatin therapy are capable of overcoming cisplatin resistance, and that inactivating mutations in NOTCH1 may be a biomarker of cisplatin sensitivity. I also used genome and kinome CRISPR libraries to identify genetic knockouts that sensitized resistant models to the EGFR inhibitors gefitinib and erlotinib. I observed that PIK3C2A may be an important linchpin in the PI3K pathway for mediating resistance, as well as identified an unexpected set of genes associated with KRAS signaling, nominating KRAS as a potential mediator of resistance to EGFR inhibition in HNSCC. Furthermore, my CRISPR/Cas9 screens also nominated FGF/FGFR knockouts as sensitizing cells to EGFR inhibition. Extending these discoveries, I investigated the potential of dual EGFR and FGFR inhibition by testing multiple UM-SCC cell lines. I observed that FGFR may be a more common compensatory mechanism that previously realized, with 14/22 (63%) of cell lines undergoing cell death when challenged with combination therapy. Surprisingly, neither copy number or expression of FGFRs predicted responsiveness to the combination of EGFR and FGFR inhibition. To explore the mechanism behind this response, I generated an EGFR K/O model and showed that FGFR signaling increases when EGFR protein is lost. Evaluation of the discovery in vivo demonstrated that dual inhibition of EGFR and FGFR was able to significantly decrease tumor volume in a xenograft mouse model, supporting the in vivo relevance of this combination for HNSCC. Consistent with the literature, dual EGFR and FGFR inhibition caused weight loss in animals suggesting a high level of toxicity; thus, this data suggests that new ways to target the pathway are critical to future clinical success. Finally, to evaluate potential clinical relevance, we analyzed transcriptome profiles of tumors from patients who received the EGFR inhibitor cetuximab. We observed changes in the FGFR receptors, KRAS signaling, and PI3K-mTOR signaling, consistent with our profiling data. Overall, my thesis work supports the hypothesis that there are specific and common compensatory pathways to EGFR inhibition and cisplatin, and that co-targeting EGFR or cisplatin with this compensatory pathway is more effective than monotherapy treatments. The CRISPR/Cas9 screens and transcriptome analysis have generated a wealth of data that can continue to be explored to develop novel, effective strategies for combination therapy. Collectively, this body of work represents a step forward in the understanding of how HNSCC tumors compensate in response to two prevalent therapies, and may provide the foundation for opportunities to advance combination therapies and improve survival of HNSCC patients.PhDCellular & Molecular BiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/151714/1/ludwigml_1.pdfDescription of ludwigml_1.pdf : Restricted to UM users only

Assessing Burnout and Well-Being in Higher Education Health Science Faculty

This study aimed to examine and compare the extent of burnout among health science faculty at a higher education institution and their self-reported perception of well-being during the COVID-19 pandemic. The design of the study was cross-sectional, descriptive survey research. An electronic questionnaire was developed to measure the constructs of burnout and well-being. Validated instruments used in the survey included the Oldenburg Burnout Inventory (OLBI) and the World Health Organization-5 Well-being Index. The Qualtrics ® platform was used to distribute the survey to all full-time faculty within the College of Health Sciences. 45 respondents from nursing, community and environmental health, kinesiology, social work, respiratory care, allied health sciences, and radiologic sciences completed the survey. Significant differences were observed in the extent of burnout and perception of well-being between faculty members who had clinical teaching responsibilities within their faculty role compared to those who did not, p = 0.005, Partial Eta Squared = 0.318. Faculty with a 9-month contract appointment had significantly lower OLBI-Disengagement scores (p = 0.024) and OLBI-Full Burnout scores (p = 0.047) compared with those with another contract length. There was a significantly negative relationship between the extent of burnout and perception of well-being. In this sample of health science faculty, burnout, as characterized by increased exhaustion and disengagement, was moderately prevalent and associated with poorer well-being

Identifying the Cytotoxic Effects of Mycobacteriophage Genes

A bacteriophage is a virus that infects and reproduces in bacteria. During productive infections—those that result in construction and release of infectious phage particles—key host cell metabolic processes are modified by the infecting phage and redirected toward making new phage particles. Protein-protein interactions are likely involved in this process. In this work, gene 80 of mycobacteriophage Vix, a gene cytotoxic to host strain Mycobacterium smegmatis, was studied. Our hypothesis was that an interaction between the Vix80 gene product and a host cell protein caused growth inhibition. Comparative analysis of the Vix80 protein sequence shows a conserved domain of unknown function 2786 (DUF2786) near the N-terminus. The Vix80 gene was dissected, and the N-terminal 66 residues, encompassing the entire DUF2786 domain, was found to be cytotoxic to M. smegmatis. DUF2786 was found to be homologous to a region of three M. smegmatis ORFs, two of which are related by alternate initiation points of the same sequence. Using in vitro protein pull-down and in vivo two-hybrid analyses, efforts are underway to look for possible interactions of Vix80 with itself and the three host proteins. As part of this process, different constructs of the Vix80 protein were expressed in Escherichia coli. We have determined that certain Vix80 constructs are also lethal to the E.coli host cell, while others permit growth, suggesting a conservation of cytotoxic function. Identifying the relevant phage and host gene products and understanding how phage exploit their host’s weaknesses could lead to new therapeutic options for many bacterial illnesses

The molecular landscape of the University of Michigan laryngeal squamous cell carcinoma cell line panel

BackgroundLaryngeal squamous cell carcinomas (LSCCs) have a high risk of recurrence and poor prognosis. Patient‐derived cancer cell lines remain important preclinical models for advancement of new therapeutic strategies, and comprehensive characterization of these models is vital in the precision medicine era.MethodsWe performed exome and transcriptome sequencing as well as copy number analysis of a panel of LSCC‐derived cell lines that were established at the University of Michigan and are used in laboratories worldwide.ResultsWe observed a complex array of alterations consistent with those reported in The Cancer Genome Atlas head and neck squamous cell carcinoma project, including aberrations in PIK3CA, EGFR, CDKN2A, TP53, and NOTCH family and FAT1 genes. A detailed analysis of FAT family genes and associated pathways showed disruptions to these genes in most cell lines.ConclusionsThe molecular profiles we have generated indicate that as a whole, this panel recapitulates the molecular diversity observed in patients and will serve as useful guides in selecting cell lines for preclinical modeling.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/151290/1/hed25803.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/151290/2/hed25803_am.pd

Comprehensive review of genetic factors contributing to head and neck squamous cell carcinoma development in low‐risk, nontraditional patients

BackgroundThe past 2 decades have seen an increased incidence of head and neck squamous cell carcinoma (HNSCC) in a nontraditional, low‐risk patient population (ie, ≤45 years of age, no substance use history), owing to a combination of human papillomavirus (HPV) infection and individual genetic variation.MethodsArticles positing genetic variants as contributing factors in HNSCC incidence in low‐risk, nontraditional patients were identified using a PubMed search, reviewed in detail, and concisely summarized herein.ResultsRecent data suggest that common polymorphisms in DNA repair enzymes, cell‐cycle control proteins, apoptotic pathway members, and Fanconi anemia‐associated genes likely modulate susceptibility to HNSCC development in low‐risk, nontraditional patients.ConclusionAt present, there is a lack of robust, comprehensive data on genetic drivers of oncogenesis in low‐risk patients and a clear need for further research on genetic alterations underlying the rising incidence of HNSCC in low‐risk, nontraditional patients.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/143606/1/hed25057_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/143606/2/hed25057.pd

The Grizzly, November 29, 2001

UC Blueskies Calls for Help with Recycling • AIDS Legacy Continues in Poor Countries • Economics and Business Administration Panel Discussion Offers Students Advice on Finding Job Connections • Volunteers Wanted for NBA League Jam Session • UC Group Makes Plans for Alternative Spring Break • New California Law Enables Women to Obtain Emergency Contraceptive Pills without a Prescription • Opinions: So This is Christmas?; Family Bonding is Better than Trip to Spain; Letter of Apology • Messiah to Hit the Stage • Jazz Ensemble Keeps it Swinging • Comparative Pricing: Local Gasoline • Dougherty Finishes 123rd at Nationals • 8+6+9 = Success for UC Wrestling Team • Lady Bears Knocked off by Susquehanna • Duncan Selected as a Finalist for the Gagliardi Trophy • Captains Leading the Way to Victory for Bears Swimming • Bears Basketball Still Looking for First Winhttps://digitalcommons.ursinus.edu/grizzlynews/1502/thumbnail.jp

Project #91: Optimizing Vascular Access to Reduce CLABSI

Henry Ford Macomb Hospital experienced an increase in Central Line Associated Bloodstream Infections (CLABSI) in 2021. A significant portion were occurring in the MICU and were associated with Candida sp. Bloodstream infections negatively impact patient outcomes, provider workload, and are costly, with a median cost of $48,108 based on a meta-analysis conducted by AHRQ in 2017. By end of 2022, HFM aimed to reduce CLABSI incidence by 50%.https://scholarlycommons.henryford.com/qualityexpo2023/1004/thumbnail.jp

Report from IPITA-TTS opinion leaders meeting on the future of β-cell replacement

Peer reviewe

Improved nucleotide selectivity and termination of 3′-OH unblocked reversible terminators by molecular tuning of 2-nitrobenzyl alkylated HOMedU triphosphates

We describe a novel 3′-OH unblocked reversible terminator with the potential to improve accuracy and read-lengths in next-generation sequencing (NGS) technologies. This terminator is based on 5-hydroxymethyl-2′-deoxyuridine triphosphate (HOMedUTP), a hypermodified nucleotide found naturally in the genomes of numerous bacteriophages and lower eukaryotes. A series of 5-(2-nitrobenzyloxy)methyl-dUTP analogs (dU.I–dU.V) were synthesized based on our previous work with photochemically cleavable terminators. These 2-nitrobenzyl alkylated HOMedUTP analogs were characterized with respect to incorporation, single-base termination, nucleotide selectivity and photochemical cleavage properties. Substitution at the α-methylene carbon of 2-nitrobenzyl with alkyl groups of increasing size was discovered as a key structural feature that provided for the molecular tuning of enzymatic properties such as single-base termination and improved nucleotide selectivity over that of natural nucleotides. 5-[(S)-α-tert-Butyl-2-nitrobenzyloxy]methyl-dUTP (dU.V) was identified as an efficient reversible terminator, whereby, sequencing feasibility was demonstrated in a cyclic reversible termination (CRT) experiment using a homopolymer repeat of ten complementary template bases without detectable UV damage during photochemical cleavage steps. These results validate our overall strategy of creating 3′-OH unblocked reversible terminator reagents that, upon photochemical cleavage, transform back into a natural state. Modified nucleotides based on 5-hydroxymethyl-pyrimidines and 7-deaza-7-hydroxymethyl-purines lay the foundation for development of a complete set of four reversible terminators for application in NGS technologies

Pain Control in Breast Surgery: Survey of Current Practice and Recommendations for Optimizing Management—American Society of Breast Surgeons Opioid/Pain Control Workgroup

Introduction: The opioid epidemic in the United States is a public health crisis. Breast surgeons are obligated to provide good pain control for their patients after surgery but also must minimize administration of narcotics to prevent a surgical episode of care from becoming a patient's gateway into opioid dependence. Methods: A survey to ascertain pain management practice patterns after breast surgery was performed. A review of currently available literature that was specific to breast surgery was performed to create recommendations regarding pain management strategies. Results: A total of 609 surgeons completed the survey and demonstrated significant variations in pain management practices, specifically within regards to utilization of regional anesthesia (e.g., nerve blocks), and quantity of prescribed narcotics. There is excellent data to guide the use of local and regional anesthesia. There are, however, fewer studies to guide narcotic recommendations; thus, these recommendations were guided by prevailing practice patterns. Conclusions: Pain management practices after breast surgery have significant variation and represent an opportunity to improve patient safety and quality of care. Multimodality approaches in conjunction with standardized quantities of narcotics are recommended