Bella Mirro, a fourth year student in Bioengineering who also minors in Chemistry, spoke with 34th Street Magazine about her many roles at Penn, including being Co–President of Shelter Health Outreach Program (SHOP), a Research Assistant in lab of Michal A. Elovitz, the Hilarie L. Morgan and Mitchell L. Morgan President’s Distinguished Professor in Women’s Health at Penn Medicine, and a Penn Engineering Council Marketing Team Member. In this Q&A, she discusses her research in women’s health and her passions for accessible healthcare, serving Philadelphia’s homeless community, and good food.
Each year, the the Department of Bioengineering seeks exceptional candidates to conduct summer research in bioengineering with the support of two scholarships: the Abraham Noordergraaf Student Summer Bioengineering Research Fund and the Blair Undergraduate Research Fund in the Department of Bioengineering. These scholarships provide a living stipend for students to conduct research on campus in a Penn research lab under the mentorship of a faculty member. The Abraham Noordergraaf Student Summer Bioengineering Research Fund provides financial support for undergraduate or graduate summer research opportunities in bioengineering with a preference for study in the area of cardiovascular systems. Dr. Noordergraaf, who died in 2014, was a founding member and first chair of Penn Bioengineering. The Blair Undergraduate Research Fund in the Department of Bioengineering supports three to five undergraduate research scholars each year with the support of Dr. James C. Blair II. After a competitive round of proposals, the following six scholars were chosen for the Summer 2022 semester. Keep reading below for the research abstracts and bios of the awardees.
The Blair Undergraduate Research Fund in the Department of Bioengineering (Blair Scholars)
Student: Ella Atsavapranee (BE Class of 2023)
PI: Michael J. Mitchell, J. Peter and Geri Skirkanich Assistant Professor of Innovation, Bioengineering
“Lipid nanoparticle-mediated delivery of RAS protease to inhibit cancer cell growth”
Mutations in RAS, a family of proteins found in all human cells, drive a third of cancers, including many pancreatic, colorectal, and lung cancers. However, there are still no therapies that can effectively prevent RAS from causing tumor growth. Recently, a protease was engineered to specifically degrade active RAS, offering a promising new tool for treating these cancers. However, many protein-based therapies still cannot be effectively delivered to patients. Lipid nanoparticles (LNPs), which were used in the Pfizer-BioNTech and Moderna COVID-19 vaccines, have emerged as a promising platform for safe and effective delivery of both nucleic acids and proteins. We formulated a library of LNPs using different cationic lipids. We characterized the LNPs by size, charge, and pKa, and tested their ability to deliver fluorescently labeled protease. The LNPs were able to encapsulate and deliver a RAS protease, successfully reducing proliferation of colon cancer cells.
Ella is a senior from Maryland studying bioengineering and chemistry. She works in Dr. Michael Mitchell’s lab, developing lipid nanoparticles to deliver proteins that reduce cancer cell proliferation. She has also conducted research on early-stage cancer detection and therapy monitoring (at Stanford University) and drug delivery across the blood-brain barrier for neurodegenerative diseases (at University of Maryland). She is passionate about translational research, science communication, and promoting diversity in STEM.
Student: Chiadika Eleh (BE and CIS Class of 2024)
PI: Eric J. Brown, Associate Professor of Cancer Biology, Perelman School of Medicine
“Investigating Viability in ATR and WEE1 Inhibitor Treated Ovarian Cancer Cells”
High-grade serous ovarian cancers (HGSOCs) are an aggressive subtype of ovarian cancer, accounting for up to 80% of all ovarian cancer-related deaths. More than half of HGSOCs are homologous recombination deficient; thus, they lack a favorable response when treated with common chemotherapeutic trials. Therefore, new treatment strategies must be developed to increase the life expectancy and quality of life of HGSOC patients. To address the lack of effective treatment options, the Brown Lab is interested in combining ATR and WEE1 inhibition (ATRi/WEE1i) to target HGSOC cells. It has previously been shown that low-dose ATRi/WEE1i is an effective treatment strategy for CCNE1-amplified ovarian cancer-derived PDX tumors (Xu et al., 2021, Cell Reports Medicine). Therefore, the next step is to characterize the HGSOC-specific response to ATRi/WEE1i treatment. This project aims to characterize the viability phenotype of ovarian cancer (OVCAR3) cells in the presence of ATRi/WEE1i in both single and combination treatments. With further research, Eleh hopes to prove the hypothesis low-dose combination ATRi/WEE1i treatment will result in the synergistic loss of viability in OVCAR3 cells. This goal will be achieved through the treatment of OVCAR3 cells with ranging doses of ATRi and Wee1i over 24 and 48 hour time intervals. We hope that this data will help set a treatment baseline that can be used for all OVCAR30-based viability experiments in the future.
Chiadika Eleh is a Bioengineering and Computer Science junior and a member of Penn Engineering’s Rachleff Scholar program. As a Blair Scholar, she worked in Dr. Eric Brown’s cancer biology lab, where she studied cell cycle checkpoint inhibitors as a form of cancer treatment.
“Tbc1d2b regulates vascular formation during development and tissue repair after ischemia”
The mechanisms behind endothelial cells forming blood vessels remains unknown. We have identified Tbc1d2b as a protein that is integral to the regulation of vascular formation. In order to investigate the role of Tbc1d2b in tubule formation, fibrin gel bead assays will be conducted to evaluate how the presence of Tbc1d2b is required for angiogenesis. Fibrin gel bead assays simulate the extracellular matrix environment to support the in vitro development of vessels from human umbilical vein endothelial cells (HUVEC) coated on cytodex beads. In order to confirm the success of angiogenesis, immunostaining for Phalloidin and CD31 will be conducted. After confirmation that fibrin gel bead assays can produce in vitro tubules, sgRNA CRISPR knockout of Tbc1d2b will be performed on HUVEC cells which will then be used to conduct more fibrin gel bead assays. We hypothesize that HUVEC with the Tbc1d2b knockout phenotype will be unable to form tubules while wild type HUVEC will be able to.
Gloria Lee is a rising senior studying Bioengineering and Physics in the VIPER program from Denver, Colorado. Her research in Dr. Yi Fan’s lab focuses on the role that proteins play in cardiovascular tubule formation.
Abraham Noordergraaf Student Summer Bioengineering Research Fund (Noordergraaf Fellows)
Student: Gary Lin (Master’s in MEAM Class of 2023)
PI: Michelle J. Johnson, Associate Professor in Physical Medicine and Rehabilitation, Perelman School of Medicine, and in Bioengineering
“Development and Integration of Dynamically Modulating Control Systems in the Rehabilitation Using Community-Based Affordable Robotic Exercise System (Rehab CARES)”
As the number of stroke patients requiring rehabilitative care continues to increase, strain is being put onto the US health infrastructure which already has a shortage of rehabilitation practitioners. To help alleviate this pressure, a cost-effective robotic rehabilitative platform was developed to increase access to rehabilitative care. The haptic TheraDrive, a one-degree of freedom actuated hand crank that can apply assistive and resistive forces, was modified to train pronation and supination at the elbow and pinching of the fingers in addition to flexion and extension of the elbow and shoulder. Two controllers were created including an open-loop force controller and a closed-loop proportional-integral (PI) with adaptive control gains based on subject performance in therapy-game tasks as well as galvanic skin response. Stroke subjects (n=11) with a range of cognitive and motor impairment completed 4 therapy games in both adaptive and non-adaptive versions of the controllers (n=8) while measuring force applied on the TheraDrive handle. Resulting normalized average power versus Upper Extremity Fugl-Meyer (UE-FM) and Montreal Cognitive Assessment (MoCA) correlation analyses showed that power was strongly correlated with UE-FM in 2 of the conditions and moderately correlated with the other 6 while MoCA was moderate correlated to 2 of the conditions and weakly correlated to the rest. Mann-Whitney U-tests between adaptive and non-adaptive versions of each therapy game showed no significant differences with regards to power between controller types (p<0.05).
Gary is a master’s student in the School of Engineering studying Mechanical Engineering and Applied Mechanics with a concentration in Robotic and Mechatronic systems. His research primarily focuses on developing affordable rehabilitation robotics for use in assessment and game-based therapies post neural injury. Many of his interests revolve around the design of mechatronic systems and the algorithms used to control them for use in healthcare spaces.
“Optogenetic Control of Developing Kidney Cells for Future Treatment of End-Stage Renal Disease”
This project sought to build from prior research in the Hughes Lab on the geometric and mechanical consequences of kidney form on cell and tissue-scale function. While the developmental trajectory of the kidney is well understood, little is currently known about many factors affecting nephron progenitor differentiation rate. Insufficient differentiation of nephron progenitor cells during kidney formation can result in lower nephron number and glomerular density, which is a risk factor for progression to end-stage renal disease later in life. Prior studies indicated that the amount of nephron differentiation – and thus function of the adult kidney – is correlated to the packing of ureteric tubule tips present at the surface of the kidney. Building off of research conducted in the Bugaj Lab, we found that inserting an optogenetic construct into the genome of human embryonic kidney (HEK) cells allowed us to manipulate the contraction of those cells through exposing them to blue light. Manipulating the contraction of the cells allows for the manipulation of the packing of ureteric tubule tips at the kidney surface. We used a lentiviral vector to transduce HEK293 cells with the optogenetic construct and witnessed visible contraction of the cells when they were exposed to blue light. Future work will include using CRISPR-Cas9 to introduce the optogenetic construct into IPS cells.
Priya is a junior studying bioengineering and had the opportunity to work on manipulating developing kidney cells using an optogenetic construct in the Hughes Lab this summer. She is thrilled to continue this research throughout the coming school year. Outside of the lab, Priya is involved with the PENNaach dance team and the Society of Women Engineers, as well as other mentorship roles.
Student: Cosette Tomita (Master’s in MEAM Class of 2023)
“Expression and Characterization of an Anti-Aβ42 scFv”
Background: Amyloid Beta (Aβ42) fibrils contribute to the pathology of Alzheimer’s Disease. Numerous monoclonal antibodies have been developed against Aβ42. In this study we have designed and expressed a short chain variable fragment specific to Aβ42 (Anti-Aβ42 scFv). To characterize our anti-Aβ42 scFv we have performed structural analysis using transmission electron microscopy (TEM) and binding kinetics using microscale thermophoresis (MST) compared to commercially available antibodies 6E10, Aducanumab, and an IgG isotype control. The goal of this study is to determine if labeling densities and binding constants for Aducanumab and anti-Aβ42 scFv are not significantly different.
Method: To characterize Aβ42 fibril associated antibodies we used negative stain TEM. Aβ42 fibrils were stained on a glow discharged copper grid, and incubated with gold conjugated anti-Aβ42 scFv, 6E10—which binds all Aβ species, aducanumab, or IgG isotype control. Labeling densities were calculated as the number of fibril-associated gold particles per 1 μm2 for each image. Next, we used microscale thermophoresis determine the binding kinetics. Antibodies or anti-Aβ42 scFv were labeled with Alexa Fluor-647 and unlabeled Aβ42 was titrated in a serial dilution over 16 capillaries. The average fluorescence intensity was plotted against the antibody or scFv concentration and the curves were analyzed using the GraphPad Prism software to calculate the dissociation constant (KD) values.
Results: We found a significant difference, tested with a one-way ANOVA (P <0.0001), in gold particle associated Aβ fibrils per 1 μm2 between anti-Aβ42 scFv, 6E10, aducanumab, and IgG isotype control. Further analysis of aducanumab and 6CO3 with unpaired student t-test indicates significant differences in fibril associated gold particles between aducanumab vs. 6E10 (P=0.0003), Aducanumab vs. Isotype control (P <0.0001), anti-Aβ42 scFv vs 6E10 (p=0.0072), and anti-Aβ42 scFv vs Isotype Control (P=0.0029) with no significant difference in labeling densities between Aducanumab and anti-Aβ42 scFv. The expected KD values from MST were 1.8μM for Aducanumab and anti-Aβ42 scFv, 10.3nM for 6E10 and no expected binding for the isotype control. The experimental KD values for anti-Aβ42 scFv and 6E10 are 0.1132μM and 1.467μM respectively. The KD value for Isotype control was undetermined, as expected, however, the KD for Aducanumab was undetermined due to suboptimal assay conditions. Due to confounding variables in the experimental set up such as the use of Aβ1-16 compared to Aβ42 and the use of different fluorophores—5-TAMRA, Alexa Fluor 647 or FITC— the experimental KD values were off by several orders of magnitude.
Conclusion: We have illustrated similar labeling densities between Aducanumab and our anti-Aβ42 scFv. In the future, we will further optimize the MST assay conditions and compare the KD values obtained by MST with other techniques such as surface plasma resonance.
Cosette was born and raised in Chicago land area. Go Sox! She attended University of Missouri where she majored in Chemistry and Biology. She synthesized sigma-2 radiotracers and developed advanced skills in biochemical techniques in Dr. Susan Lever’s lab. After graduation, she moved to NJ to work at Lantheus, a radiopharmaceutical company. She missed academia and the independence of program and project development, so she came to work at the Penn Cyclotron facility before entering the Bioengineering master’s program.
Cells in complex organisms undergo frequent changes, and researchers have struggled to monitor these changes and create a comprehensive profile for living cells and tissues. Historically researchers have been limited to only 3-5 markers due to spectral overlaps in fluorescence microscopy, an essential tool required for imaging cells. With only this small handful of markers, it is difficult to monitor protein expressions of live cells and a comprehensive profile of cellular dynamics cannot be created. However, a new study in Nature Biotechnology addresses these limitations by demonstrating a new method for comprehensive profiling of living cells.
Jina Ko, Assistant Professor in Bioengineering in the School of Engineering and Applied Science and in Pathology and Laboratory Medicine in the Perelman School of Medicine, conducted postdoctoral research at Massachusetts General Hospital (MGH) and the Wyss Institute at Harvard University, and the work for this study was done under the supervision of Jonathan Carlson M.D., Ph.D. and Ralph Weissleder M.D., Ph.D. of MGH. Ko’s lab at Penn develops novel technologies using bioengineering, molecular biology, and chemistry to address diagnostic challenges for precision medicine.
To address these limitations in microscopy, the team developed a new chemistry tool which was highly gentle to cells. This “scission-accelerated fluorophore exchange (or SAFE)” method utilizes “click” chemistry, a type of chemistry that follows examples found in nature to create fast and simple reactions. This new SAFE method functions with non-toxic conditions to living cells and tissues, whereas previous methods have used harsh chemicals that would strip off fluorophores and consequently would not work with living cells and tissues.
With the development of SAFE, the authors demonstrated that researchers can now effectively perform multiple cycles of cell profiling and can monitor cellular changes over the course of their observations. Instead of the previous limitation of 3-5 markers total, SAFE allows for many more cycles and can keep track of almost as many markers as the researcher wants. One can now stain cells and quench/release fluorophores and repeat the cycle multiple times for multiplexing on living cells. Each cycle can profile 3 markers, and so someone interested in profiling 15 markers could easily perform 5 cycles to achieve this much more comprehensive cell profile. With this breakthrough in more detailed imaging of cells, SAFE demonstrates broad applicability for allowing researchers to better investigate the physiologic dynamics in living systems.
César de la Fuente, Presidential Assistant Professor in Psychiatry, Bioengineering, Microbiology, and in Chemical and Biomolecular Engineering has been honored with a 2022 Young Investigator Award by the Royal Spanish Society of Chemistry (RSEQ) for his pioneering research efforts to combine the power of machines and biology to help prevent, detect, and treat infectious diseases.
With the goal of supporting partnerships between minority-serving educational institutions and leading materials science research centers, NSF’s Partnership for Research & Education in Materials (PREM) program funds innovative research programs and provides institutional support to increase recruitment, retention, and graduation by underrepresented groups as well as providing underserved communities access to materials research and education.
‘Research at the frontier’
With this PREM award, known as the Advancing Device Innovation through Inclusive Research and Education (ADIIR) program, researchers from Penn and UPR’s Humacao and Cayey campuses will conduct research on the properties of novel carbon-based materials with unique properties, and will study the effects of surface modification in new classes of sensors, detectors, and purification devices.
Thanks to this collaboration of more than 20 years, both institutions have made significant scientific and educational progress aided by biannual symposia and regular pre-pandemic travel between both institutions before the pandemic, resulting in a rich portfolio of publications, conference presentations, patents, students trained, and outreach programs.
“Together we have been publishing good papers that have impact, and we’ve really cultivated a culture of collaboration and friendship between our institutions,” says Penn’s Arjun Yodh, former director of the LRSM. “Our goal is to carry out research at the frontier and, in the process, nurture promising students from Puerto Rico and Penn.”
Ivan Dmochowski, a chemistry professor at Penn who has been involved with PREM for several years, says that this program has helped his group connect with experts in Puerto Rico whose skills complement his group’s interests in protein engineering. Dmochowski has also hosted UPR faculty members and students in his lab and also travelled to Puerto Rico before the pandemic to participate in research symposia, seminars, and outreach events.
“I’ve had students who have benefitted from being a co-author on a paper or having a chance to mentor students, and the faculty we’ve interacted with are exceptional,” Dmochowski says. “There’s a lot of benefit for both me and my students, and I’ve enjoyed our interactions both personally and scientifically.”
Penn’s Daeyeon Lee, a chemical and biomolecular engineering professor who has been involved with PREM for several years, regularly hosts students and faculty from UPR while working on nanocarbon-based composite films for sensor applications. The success of this collaboration relies on unique materials made by researchers at UPR combined with a method for processing them into composite structures developed in Lee’s lab.
“What I really admire about people at PREM, both faculty and students, is their passion,” says Lee. “I think that’s had a really positive impact on my students and postdocs who got to interact with them because they got to see the passion that the students brought.”
Manuela Teresa Raimondi was appointed Visiting Professor in Bioengineering in the Associated Faculty of the School of Engineering and Applied Science for the 2020-2021 academic year. Raimondi received her Ph.D. in Bioengineering in 2000 from Politecnico di Milano, Italy. She is currently a Full Professor of Bioengineering at Politecnico di Milano in the Department of Chemistry, Materials and Chemical Engineering “G. Natta”, where she teaches the course “Technologies for Regenerative Medicine” in the Biomedical Engineering graduate program.
Raimondi is the founder and Director of the Mechanobiology Lab and of the Interdepartmental Live Cell Imaging lab. She has pioneered the development of cutting edge tools for cell modelling, ranging from micro-engineered stem cell niches, to miniaturized windows for in vivo intravital imaging, to microfluidic culture systems to engineer tissue-equivalents and organoids for cell modelling and drug discovery. Her platforms are currently commercialized by her start-up, MOAB srl. Her research is funded by the European Research Council (ERC), by The National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs), by the European Commission, and by the European Space Agency.
“Getting to Penn was quite the challenge with the various travel restrictions and the pandemic, but I am used to overcoming adverse odds and I am really excited to be here now,” says Dr. Raimondi. “In this challenging time, when many new barriers are coming up, I think building bridges and new scientific collaborations is even more important. I very much look forward to being part of the Penn research community.”
During her sabbatical at Penn, Raimondi is investigating her hypothesis that stem cells pluripotency reprogramming can be guided by mechanical cues. Over the past five years, she has cultured many different stem cell types in the “Nichoids,” the synthetic stem cell niche she developed, and gathered robust evidence on how physical constraints at the microscale level upregulate pluripotency. Raimondi is hosted in the Bioengineering and Biomaterials Lab of Riccardo Gottardi, Assistant Professor in Bioengineering and in Pediatrics at the Perelman School of Medicine, where she is helping to refine human stem cell sources that could be minimally manipulated for translational tissue engineering for a safe and effective use in regenerative therapies, as a key issue for clinical translation is the maintenance or enhancement of multipotency during cell expansion without exogenous agents or genetic modification.
“Dr. Raimondi is a trailblazer in Italy in regenerative medicine who has introduced many new concepts in a sometimes musty academic environment and has shattered a number of glass ceilings,” says Dr. Gottardi. “I think her sabbatical at Penn is a great opportunity for her and for the Penn community to build new and exciting trans-Atlantic collaborations.”