Lab-on-a-Chip and Microfluidics 2020 Agenda

Welcome, Introduction, Topics Addressed and Conference Opening by Conference Chairperson
Leanna Levine, Founder & CEO, ALine, Inc., United States of America

Microfluidics for Interrogating Intact-Tumor Biopsies
Albert Folch, Professor of Bioengineering, University of Washington, United States of America

Cancer remains a major healthcare challenge worldwide. It is now well established that cancer cells constantly interact with fibroblast cells, endothelial cells, immune cells, signaling molecules, and the extracellular matrix in the tumor microenvironment (TME). Present tools to study drug responses and the TME have not kept up with drug testing needs. The number of clinical trials of combination therapies has been climbing at an unsustainable rate, with 3,362 trials launched since 2006 to test PD-1/PD-L1-targeted monoclonal antibodies alone or in combination with other agents. We have developed a microfluidic platform (called Oncoslice) for the delivery of multiple drugs with spatiotemporal control to live tumor biopsies, which retain the TME. We have developed the use of Oncoslice for the delivery of small-molecule cancer drug panels to glioblastoma (GBM) xenograft slices as well as to slices from patient tumors (GBM and colorectal liver metastasis). In addition, we have developed a precision slicing methodology that allows for producing large numbers of cuboidal micro-tissues (“cuboids”) from a single tumor biopsy. We have been able to trap cuboids in arrays of microfluidic traps in a multi-well platform. This work will potentially allow for the high-throughput application of drugs to intact human tumor tissues.

Microfluidic Technologies For Liquid Biopsy
Daniel Chiu, A. Bruce Montgomery Professor of Chemistry, University of Washington, United States of America

This presentation will describe microfluidic technologies we developed for liquid biopsy and precision medicine. Specifically, a rare-cell isolation instrument we call eDAR (ensemble decision aliquot ranking), a nanofluidic technology for the high-sensitivity sorting of exosomes, and a digital-nucleic-acid detection and analysis platform based on our SD (self-digitization) chip. I will outline the workings of these microfluidic platforms, describe their performance, and discuss the clinical questions we are addressing with these technologies.

Precision Biology: Deep Profiling of Single Cells Using Electrophoretic Cytometry
Amy Herr, Professor, University Of California Berkeley, United States of America

Underpinning single-cell measurement tools, microfluidic design offers the throughput, multiplexing, and quantitation needed for rich, multi-dimensional data. Genomics and transcriptomics are leading examples. Yet, while proteins are the dynamic, downstream effectors of function, the immunoassay remains the de facto standard (flow cytometry, mass cytometry, immunofluorescence). We posit that to realize the full potential of high-dimensionality cytometry, new approaches to protein measurement are needed. I will describe our ‘electrophoretic cytometry’ tools that increase target selectivity beyond simple immunoassays. Enhanced selectivity is essential for targets that lack high quality immunoreagents – as is the case for the vast majority of protein forms (proteoforms).  I will share our results on highly multiplexed single-cell western blotting and single-cell isoelectric focusing that resolves single charge-unit proteoform differences. In fundamental engineering and design, I will discuss how the physics and chemistry accessible in microsystems allows both the “scale-down” of electrophoresis to single cells and the “scale-up” to concurrent analyses of large numbers of cells. Precise reagent control allows for integration of cytometry with sophisticated sample preparation – the unsung hero of measurement science. Lastly, I will link our bioengineering research to understanding the role of protein signaling and truncated isoforms in development of breast cancer drug resistance and understanding protein signaling in individual circulating tumor cells. Taken together, we view microfluidic design strategies as key to advancing protein measurement performance needed to address unmet gaps in quantitative biology and precision medicine.

Microengineering a Physiologic Colon Replica
Nancy Allbritton, Frank and Julie Jungers Dean of the College of Engineering and Professor of Bioengineering, University of Washington in Seattle, United States of America

Organ-on-chips are miniaturized devices that arrange living cells to simulate functional subunits of tissues and organs. These microdevices provide exquisite control of tissue microenvironment for the investigation of organ-level physiology and disease. A 3D polarized epithelium using primary human gastrointestinal stem cells was developed to fully recapitulate gastrointestinal epithelial architecture and physiology. A planar monolayer comprised of stem/proliferative and differentiated primary cells is cultured on a shaped hydrogel scaffold with an array of crypt-like structures replicating the intestinal architecture. These planar layers display physiologic drug transport and metabolism and immunologically appropriate responses. Facile co-culture with other cell types such as immune cells or myofibrolasts is readily achieved. A dense mucus layer is formed on the luminal epithelial surface that is impermeable to bacteria and acts a barrier to toxins. The in vitro mucus has remarkably similar in its biophysical properties to that produced in vivo. Imposition of chemical gradients across the crypt long axis yields a polarized epithelium with a stem-cell niche and differentiated cell zone. The stem cells proliferate, migrate and differentiate along the crypt axis as they do in vivo. An oxygen gradient across the tissue mimic permits luminal culture of anaerobic bacteria while maintaining an oxygenated stem cell niche. This in vitro human colon crypt array replicates the architecture, luminal accessibility, tissue polarity, cell migration, and cellular responses of in vivo intestinal crypts. This bioanalytical platform is envisioned as a next-generation system for assay of microbiome-behavior, drug-delivery and toxin-interactions with the intestinal epithelia.

Workflow Automation with Digital and Customizable Microfluidics: Challenges and Opportunities
Anita Rogacs, Head of Life Sciences Strategy and R&D, HP Labs, United States of America

Life Sciences analytical workflows benefit greatly from microfluidic automation, leading to improvement in accuracy, multiplexing, efficiency, and cost. Yet microfluidics has faced significant barriers to adoption in both diagnostics and drug discovery, due to constrained designs and long and costly development pipelines. HP is addressing this challenge by leveraging our digital and customizable mode of microfluidics, eliminating the need to design a “new chip” for every new workflow, target and reagent.

Exploiting Cardiac Microphysiological Systems For COVID-19 Drug Screening
Kevin Healy, Jan Fandrianto and Selfia Halim Distinguished Professorship in Engineering, University of California, Berkeley, United States of America

Our work has emphasized creating both healthy and diseased cardiac and liver microphysiological systems (MPS) or ‘organ chips’, to address the costly and inefficient drug discovery process. While MPS are poised to disrupt the drug development process and significantly reduce the cost of bringing a new drug candidate to market, the technology is more robust and creates a whole new paradigm in how to conduct safety pharmacology science, and advances medicine in revolutionary ways. An emerging use of MPS is in the evaluation of repurposed drugs to treat COVID-19. While repurposed drugs are typically FDA-approved for monotherapy, most have not been tested in polytherapy with  anti-inflammatory or antibiotic medications typically employed as part of intensive care protocols. Since COVID-19 patient morbidity is highly correlated with myocardial injury, independent of pre-existing cardiovascular disease, this presentation will address examples of exploiting our human cardiac MPS as unique testbeds for rapidly assessing the cardiac liability of polytherapy of repurposed COVID-19 drugs. Preclinical data generated will inform clinical trial design for polytherapies for COVID-19 patients, particularly regarding risks of potential drug-drug interactions or identifying appropriate exclusion criteria, monitoring strategies, and dose adjustments to minimize cardiac liabilities.

Optofluidic Systems For On-Chip Molecular Diagnostics
Holger Schmidt, Narinder Kapany Professor of Electrical Engineering, University of California-Santa Cruz, United States of America

Optofluidic devices have emerged as the basis for ultra-sensitive, amplification-free analysis of single molecules. We will describe the core features and capabilities of this approach and discuss recent advances in the areas of integration of multiple functions on a single chip, optical detection and analysis with maximized efficiency, and representative examples such as multiplex detection of single antibiotic resistant bacteria and SARS-CoV-2 detection.

Microfluidic-based Diagnostic Approaches for COVID-19
Dino Di Carlo, Armond and Elena Hairapetian Chair in Engineering and Medicine, Professor and Vice Chair of Bioengineering, University of California-Los Angeles, United States of America

Microfluidic technologies have laid a foundation for a number of diagnostic solutions that are especially relevant during the COVID-19 pandemic. I will discuss technological platforms being applied to rapidly detect the SARS-CoV-2 virus, characterize antibodies produced by individuals in response to infection, and triage patients based on detection of abnormal host immune activation. Integration of microfluidic and machine learning approaches enable these tests to have rapid turnaround times which are critical for  clinical scenarios involving diagnosing infected individuals at the point-of-care, broadly screening for immunity / vaccination effectiveness, and triaging patients entering the emergency room based on the amount of care needed.

Plastic-based Nanofluidic Sensor for the Detection of Rare Nucleic Acids and Determining Their Sequence Variations from Liquid Biopsy Markers
Steve Soper, Foundation Distinguished Professor, Director, Center of BioModular Multi-scale System for Precision Medicine, The University of Kansas, United States of America

Liquid biopsy markers (circulating tumor cells, CTCs; extracellular vesicles, EVs; and cell free DNA, cfDNA) are becoming extremely popular to manage a variety of cancer-related diseases due to the minimally invasive nature of their acquisition. However, the challenge with liquid biopsy markers is their rarity; for example, it is not uncommon to secure 1-100 CTCs per mL of whole blood supplying about 6-600 pg of genomic DNA. Because platforms like next generation sequencing require >30 ng of input DNA, significant amounts of amplification of the input are required that can generate a biased representation of the genome. To mitigate this issue, we have produced a mixed-scale nanofluidic sensor featuring a baffle area, high surface area pillar arrays, and nanometer flight tubes.  The pixel arrays can perform solid-phase ligase detection reactions (spLDRs) to score the presence of DNA mutations found in a diseased patient even when the mass of the marker is low (<1 ng), but does not require PCR amplification for the analysis. The spLDR can also expression profile mRNAs following reverse transcription. Successfully formed spLDR products are identified using a molecular-dependent time-of-flight (TOF) through a polymer nanofluidic channel flanked by two in-plane nanopores. Simulations (COMSOL) were used to guide the design and fabrication of the nanofluidic sensor to determine the loading efficiency and transport patterns of spLDR products from the pillar array into the flight tubes by evaluating operational parameters when using either hydrodynamic or electrokinetic flow. The nanofluidic sensor was fabricated from a Si master patterned using a combination of focused ion beam (FIB) milling and photolithography with inductively coupled plasma reactive ion etching. The Si master was used to produce resin stamps that were then used to transfer the relevant structures to a plastic via thermal nanoimprint lithography (NIL). The operational features of the device will be presented as well as detecting point mutations in KRAS genes from CTCs’ genomic DNA as well as mRNA expression profiling.

Microfluidic Cell Engineering for Precision Therapy
Abraham Lee, Professor, University of California Irvine, United States of America

Precision medicine is the paradigm to develop treatments for patients based on molecular-targets that are effective in vivo when administered.  That is, one must not only be able to identify molecular and cellular targets that are the source of disease but also understand how these targets behave in the body based on physiological principles.  Recent developments in microfluidics have contributed to burgeoning precision medicine fields such as liquid biopsy, immunotherapy, single cell analysis, genotyping and gene sequencing, and microphysiological systems.  A specific focus of my talk would be on microfluidic devices for engineering cells for developing targeted cell therapies and immunotherapies. These microfluidic technologies have the potential to close the loop from detection to diagnosis, to therapy.

Free-Surface Microfluidics and SERS for High Performance Sample Capture and Analysis
Carl Meinhart, Professor, University of California-Santa Barbara, United States of America

Nearly all microfluidic devices to date consist of some type of fully-enclosed microfluidic channel. The concept of ‘free-surface’ microfluidics has been pioneered at UCSB during the past several years, where at least one surface of the microchannel is exposed to the surrounding air.  Surface tension is a dominating force at the micron scale, which can be used to control effectively fluid motion. There are a number of distinct advantages to the free surface microfluidic architecture.  For example, the free surface provides a highly effective mechanism for capturing certain low-density vapor molecules.  This mechanism is a key component (in combination with surface-enhanced Raman spectroscopy, i.e. SERS) of a novel explosives vapor detection platform, which is capable of sub part-per-billion sensitivity with high specificity.

Tracking Tumor Extracellular Vesicles Using Microfluidics
Shannon Stott, Assistant Professor, Massachusetts General Hospital & Harvard Medical School, United States of America

Advances in microfluidic technologies and molecular profiling have propelled the rapid growth and interest in achieving a ‘liquid biopsy’ in cancer. The ability to manipulate fluidics flows towards the goal of interrogating millions of particles per second provides a significant advantage over other technological approaches. Through a collaborative effort between bioengineers, biologists, and clinicians, my laboratory at Massachusetts General Hospital has developed microfluidic devices to isolate and characterize these EVs from whole blood. For this presentation, data will be presented on our effort to use our devices to serially track EVs in glioblastoma patients over time. Further, we will share details on some of our latest technologies in development. Through the microfluidic isolation of blood based biomarkers from patients, our goal is to obtain complementary data to the current standard of care to help better guide treatment and identify new biomarkers and putative therapeutic targets.