Penn Integrates Knowledge Professor Kevin Johnson takes the stage at 24th Engaging Minds. (Image: Ben Asen)
This past weekend in New York City, the University of Pennsylvania showcased its 24th Engaging Minds event, the first in person since 2019. It was hosted by Penn Alumni.
Three Penn Integrates Knowledge University Professors — Kevin Johnson, Lance Freeman and Dolores Albarracín, — each discussed their research. The audience, at least 600 in person and remote, heard about using city planning to promote racial equity, about how conspiracy theories come to life and propagate, and about the need for physicians to communicate effectively with patients and families.
Following brief remarks from Penn Alumni President Ann Reese, University President Liz Magill introduced the event. “As many of you know, I’ve been thinking a lot and speaking often about what makes Penn Penn,” she said. “What are our distinctive strengths? What are the unique contributions to society that we have made in the past and can make in the future? And where do we go from the extraordinary position we are in now?”
Magill went on to express gratitude for the speakers and invited the audience to think about how the researchers’ work and expertise furthered what she described as the “twin principles of truth and opportunity.”
He took the audience through his family history, education and training, pausing at a point on the timeline when he was a young physician-scientist who had just explained a new medical topic to a journalist. “I felt really good about the conversation — and then the article came out,” Johnson said.
In the piece, he had been cast as saying that the medical community was over-treating this condition, “which is not what I said.” He realized in that moment that as a physician, he had been taught to communicate what a study finds, not how to act based on those findings. That experience shifted his thinking on how to communicate scientific topics, and he has spent decades trying to move the needle on how others in his field perceive this.
“As scientists we face obstacles. We face the obstacle of scale, so, small projects that we’re asked to generalize. We face the issue of trust. And then we face the issue of values,” Johnson said. “I’ll add a fourth, which is format; the way we choose to reach specific audiences will be different.”
Read more about the 24th Engaging Minds at Penn Today.
Kevin Johnson is the David L. Cohen University of Pennsylvania Professor in the Departments of Biostatistics, Epidemiology and Informatics and Computer and Information Science. As a Penn Integrates Knowlegde (PIK) University Professor, Johnson also holds appointments in the Departments of Bioengineering and Pediatrics, as well as in the Annenberg School of Communication.
The 2022 iGEM team from left to right: June Ahn, Shreya Villimanalan, Adiva Daniar, Wangari Mbuthia, Cristina Perez and Moses Zeidan.
Congratulations to the 2022 University of Pennsylvania iGEM Team who took home a gold medal in the iGEM Grand Jamboree. This international competition of multidisciplinary teams of graduate and undergraduate students presenting original projects in synthetic biology culminated in the in-person Jamboree event held in Paris, France in October 2022. Over 370 judges awarded prizes and medals to the 350+ teams representing over 40 countries.
The 2022 Penn team was awarded a Gold Medal for their project “Photocreate,” a toolbox to control intercellular communication using optogenetics. Their plasmid constructs are designed to control protein secretion, display and shedding using a photocleavable protein, Phocl. The full abstract reads:
Intercellular communication is primarily studied using synthetic protein-level circuits. These circuits currently lack the spatial and temporal control necessary for targeted and time-sensitive applications. To address this gap, we developed Photocrete, a toolbox of protein constructs for light-inducible control of protein display, secretion, and shedding. We expanded upon RELEASE (Vlahos et al.), a modular and generalizable protein circuit which utilizes an ER retention motif and an exogenous protease to control protein secretion. We optogenetically modified RELEASE by replacing different components with the photocleavable protein PhoCl, allowing us to control the mammalian secretion pathway at distinct nodes with finely-tuned light administration regimens. Preliminary results indicate integration of Photocrete into the secretion pathway, but more research is necessary to determine optimal light administration settings. The potential for high spatial and temporal control of Photocrete could allow researchers to perform various signaling studies and develop therapeutics at a new level of precision.
The 2022 iGEM team includes undergraduates June Ahn (B.S. in Biochemistry, Physics and Nutrition), Adiva Daniar (B.S.E. in Bioengineering, minor in Engineering Entrepreneurship), Wangari Mbuthia (B.S.E. in Bioengineering), Cristina Perez (B.S.E. in Bioengineering, minor in Physics), Shreya Vallimanalan (B.S.E. in Bioengineering, minor in Computational Neuroscience), an d Moses Zeidan (B.S.E. in Bioengineering, minor in Chemistry and Spanish). They were mentored by graduate students David Gonzalez-Martinez, Gabrielle Ho, Zikang Huang, and Will Benman. Their faculty advisor is Lukasz Bugaj, Assistant Professor in Bioengineering.
Read the full results of the 2022 iGEM Competition here.
The Heilmeier Award honors a Penn Engineering faculty member whose work is scientifically meritorious and has high technological impact and visibility. It is named for the late George H. Heilmeier, a Penn Engineering alumnus and member of the School’s Board of Advisors, whose technological contributions include the development of liquid crystal displays and whose honors include the National Medal of Science and Kyoto Prize.
Bassett, who also holds appointments in Physics & Astronomy in Penn Arts & Sciences and in Neurology and Psychiatry in the Perelman School of Medicine, is a pioneer in the field of network neuroscience, an emerging subfield which incorporates elements of mathematics, physics, biology and systems engineering to better understand how the overall shape of connections between individual neurons influences cognitive traits. They lead the Complex Systems lab, which tackles problems at the intersection of science, engineering and medicine using systems-level approaches, exploring fields such as curiosity, dynamic networks in neuroscience, and psychiatric disease.
Bassett will deliver the 2022-23 Heilmeier Award Lecture in Spring 2023.
Dan Huh’s BIOLines Lab develops several different kinds of organ-on-a-chip systems, such as this blinking-eye-on-a-chip.
If you’d read about it in a science fiction novel, you might not have believed it. Human organs and organ systems — from lungs to blood vessels to blinking eyes — bio-miniaturized and stored on a plastic chip no larger than a matchbook.
But that’s the breathing, blinking reality at the Biologically Inspired Engineering Systems (BIOLines) Laboratory in the Department of Bioengineering in the School of Engineering and Applied Sciences at the University of Pennsylvania, a bona fide pioneer of what is now widely known as “organ-on-a-chip” technology. Proponents hope these devices can one day help scientists around the world learn more about the body’s inner workings and ultimately improve disease prevention and treatment.
“The century-old practice of cell culture is to grow living cells isolated from the human body in hard plastic dishes and keep them bathed in copious amounts of culture media under static conditions, and that is drastically different than the complex, dynamic environment of native tissues in which these cell reside,” said Dan Dongeun Huh, Ph.D., BIOLines’ principal investigator and an associate professor of Bioengineering in Penn’s School of Engineering and Applied Science. “What makes this organ-on-a-chip technology so unique and powerful is that it enables us to reverse-engineer living human tissues using microengineered devices and mimic their intricate biological interactions and physiological functions in ways that have not been possible using traditional cell culture techniques. This represents a major advance in our ability to model and understand the inner workings of complex physiological systems in the human body.”
Generally speaking, organ-on-a-chip devices are made of clear silicone rubber — the same material used to make contact lenses — and can vary in size and design. Embedded within are microfabricated three-dimensional chambers lined with different human cell types, arranged and propagated to ultimately form a structure complex enough to actually mimic the essential elements of a functioning organ.
With partners at the Perelman School of Medicine, BIOLines recently developed a newer variation of the organ-on-a-chip: one that replicates the interface between maternal tissue and the cells of the placenta at the critical moments in early pregnancy when the embryo is implanting in the uterus. Huh and Penn Medicine physicians led a study using the “implantation-on-a-chip” to observe things that would otherwise have been virtually unobservable.
We hope you will join us for the 2022 Grace Hopper Distinguished Lecture by Dr. Jennifer Lewis, presented by the Department of Bioengineering and hosted by Dani S. Bassett, J. Peter Skirkanich Professor in Bioengineering, Electrical and Systems Engineering, Physics & Astronomy, Neurology and Psychiatry.
Date: Thursday, December 8, 2022
Start Time: 3:30 PM EST
Location: Glandt Forum, Singh Center for Nanotechnology, 3205 Walnut Street, Philadelphia, PA 19104
Join us after the live lecture for a light reception!
Daphna Shohamy, Ph.D.
Speaker: Daphna Shohamy, Ph.D.
Kavli Professor of Brain Science, Co-Director of the Kavli Institute for Brain Science, Professor in the Department of Psychology & Zuckerman Mind Brain Behavior Institute Columbia University
From robots to humans, the ability to learn from experience turns a rigid response system into a flexible, adaptive one. In the past several decades, major advances have been made in understanding how humans and other animals learn from experience to make decisions. However, most of this progress has focused on rather simple forms of stimulus-response learning, such as automatic responses or habits. In this talk, I will turn to consider how past experience guides more complex decisions, such as those requiring flexible reasoning, inference, and deliberation. Across a range of behavioral contexts, I will demonstrate a critical role for memory in such decisions and will discuss how multiple brain regions interact to support learning, what this means for how memories are used, and the consequences for how decisions are made. Uncovering the pervasive role of memory in decision-making challenges the way we think about what memory is for, suggesting that memory’s primary purpose may be to guide future behavior and that storing a record of the past is just one way to do so.
Dr. Shohamy Bio:
Daphna Shohamy, PhDis a professor at Columbia University where she co-directs the Kavli Center for Neural Sciences and is Associate Director of the Zuckerman Mind, Brain Behavior Institute. Dr. Shohamy’s work focuses on the link between memory, and decision-making. Combining brain imaging in healthy humans with studies of patients with neurological and psychiatric disorders, Dr. Shohamy seeks to understand how the brain transforms experiences into memories; how memories shape decisions and actions; and how motivation and exploration affect human behavior.
Information on the GraceHopper Lecture: In support of its educational mission of promoting the role of all engineers in society, the School of Engineering and Applied Science presents the GraceHopper Lecture Series. This series is intended to serve the dual purpose of recognizing successful women in engineering and of inspiring students to achieve at the highest level.
Rear Admiral GraceHopper was a mathematician, computer scientist, systems designer and the inventor of the compiler. Her outstanding contributions to computer science benefited academia, industry and the military. In 1928 she graduated from Vassar College with a B.A. in mathematics and physics and joined the Vassar faculty. While an instructor, she continued her studies in mathematics at Yale University where she earned an M.A. in 1930 and a Ph.D. in 1934. GraceHopper is known worldwide for her work with the first large-scale digital computer, the Navy’s Mark I. In 1949 she joined Philadelphia’s Eckert-Mauchly, founded by the builders of ENIAC, which was building UNIVAC I. Her work on compilers and on making machines understand ordinary language instructions lead ultimately to the development of the business language, COBOL. GraceHopper served on the faculty of the Moore School for 15 years, and in 1974 received an honorary degree from the University. In support of the accomplishments of women in engineering, each department within the School invites a prominent speaker for a one or two-day visit that incorporates a public lecture, various mini-talks and opportunities to interact with undergraduate and graduate students and faculty.
Twin siblings and scholars Dani S. Bassett of Penn and Perry Zurn of American University collaborated over half a dozen years to write “Curious Minds: The Power of Connection.” (Image: Tony and Tracy Wood Photography)
Twin academics Dani S. Basset, J. Peter Skirkanich Professor and director of the Complex Systems Lab, and Perry Zurn, a professor of philosophy at American University, were recently featured as guests on NPR radio show “Detroit Today” to discuss their new book, “Curious Mind: The Power of Connection.”
In their book, Basset and Zurn draw on their previous research, as well as an expansive network of ideas from philosophy, history, education and art to explore how and why people experience curiosity, as well as the different types it can take.
Basset, who holds appointments in the Departments of Bioengineering and Electrical and Systems Engineering, as well as the Department of Physics and Astronomy in Penn Arts & Science, and the Departments of Neuroscience and Psychiatry in Penn Perelman’s School of Medicine, and Zurn spoke with “Detroit Today” producer Sam Corey about what types of things make people curious, and how to stimulate more curiosity in our everyday lives.
According to the twin experts, curiosity is not a standalone facet of one’s personality. Basset and Zurn’s work has shown that a person’s capacity for inquiry is very much tied to the overall state of their health.
“There’s a lot of scientific research focusing on intellectual humility and also openness to ideas,” says Bassett. “And there are really interesting relationships between someone’s openness to ideas, someone’s intellectual humility and their curiosity and also their wellbeing or flourishing,”
An infographic explains the magnetic and catalytic properties of the iron oxide nanoparticles and their assembly into bristle and floss-like forms. (Image: Melissa Pappas/Penn Engineering)
Koo shared findings from one of his recent studies conducted in collaboration with Penn Engineering, which showed that a shapeshifting robotic microswarm can brush and floss teeth.
“Routine oral care is cumbersome and can pose challenges for many people, especially those who have a hard time cleaning their teeth” says Koo. “You have to brush your teeth, then floss your teeth, then rinse your mouth; it’s a manual, multistep process. The big innovation here is that the robotics system can do all three in a single, hands-free, automated way.”
The building blocks of these microrobots are iron oxide nanoparticles that have both catalytic and magnetic activity. Using a magnetic field, researchers could direct their motion and configuration to form either bristlelike structures that sweep away dental plaque from the broad surfaces of teeth, or elongated strings that can slip between teeth like a length of floss.
“Nanoparticles can be shaped and controlled with magnetic fields in surprising ways,” says Edward Steager, a senior research investigator at Penn Engineering and co-corresponding author. “We form bristles that can extend, sweep, and even transfer back and forth across a space, much like flossing. The way it works is similar to how a robotic arm might reach out and clean a surface. The system can be programmed to do the nanoparticle assembly and motion control automatically.”
Eight researchers from the Perelman School of Medicine have received research grants designed to invest in high-risk, high-reward projects.
Bushra Raj, Assistant Professor of Cell and Developmental Biology in the Perelman School of Medicine and member of the Penn Bioengineering Graduate Group, was one of three Penn winners of the NIH Director’s New Innovator Award for independent projects developed by early-career investigators. More additional Penn scientists who received NIH Director’s Transformative Research Award for a project focusing on cancer research.
Raj’s project focuses on “testing a novel technology that uses CRISPR/Cas gene-editing tools to genomically record inputs from two signaling pathways in the developing zebrafish brain.”
Established in 2009, the Transformative Research Award promotes cross-cutting, interdisciplinary science and is open to individuals and teams of investigators who propose research that could potentially create or challenge existing paradigms.
“Padilla came to the CiPD training program earlier this year with a Ph.D. in Chemistry from the University of Wisconsin-Madison. He is currently a postdoctoral fellow in the lab of Dr. Michael J. Mitchell of Penn’s Department of Bioengineering, where his research focuses on developing new materials to enhance the efficacy and safety of biological therapeutics. While passionate about research, he also has a strong interest in developing mentoring relationships and in teaching. At Wisconsin, Marshall earned a certificate in research, teaching, and learning, in which he conducted a research project on developing positive metacognitive practices in introductory organic chemistry. Additionally, he taught a course on mentoring in a research setting, and is passionate about promoting diversity and inclusiveness in biomedical sciences.”
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)
Ella Atsavapranee
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.
Chiadika Eleh
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)
Gary Lin
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.
Priya Shah
Student: Priya Shah (BE Class of 2024)
PI: Alex J. Hughes, Assistant Professor in Bioengineering
“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.
Cosette Tomita
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.
