Tag Archives: Michael L. Shuler

Human-on-a-chip predicts in vivo results based on in vitro model … for the first time

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If successful the hope is that ‘human-on-a-chip’ will replace most, if not all, animal testing. This July 3, 2019 Hesperos news release (also on EurekAlert) suggests scientists are making serious gains in the drive to replace animal testing (Note: For anyone having difficulty with the terms, pharmacokinetics and pharmacodynamics, there are definitions towards the end of this posting, which may prove helpful),

Hesperos Inc., pioneers* of the “human-on-a-chip” in vitro system has announced the use of its innovative multi-organ model to successfully measure the concentration and metabolism of two known cardiotoxic small molecules over time, to accurately describe the drug behavior and toxic effects in vivo. The findings further support the potential of body-on-a-chip systems to transform the drug discovery process.

In a study published in Nature Scientific Reports, in collaboration with AstraZeneca, Hesperos described how they used a pumpless heart model and a heart:liver system to evaluate the temporal pharmacokinetic/pharmacodynamic (PKPD) relationship for terfenadine, an antihistamine that was banned due to toxic cardiac effects, as well as determine its mechanism of toxicity.

The study found there was a time-dependent, drug-induced response in the heart model. Further experiments were conducted, adding a metabolically competent liver module to the Hesperos Human-on-a-Chip® system to observe what happened when terfenadine was converted to fexofenadine. By doing so, the researchers were able to determine the driver of the pharmacodynamic (PD) effect and develop a mathematical model to predict the effect of terfenadine in preclinical species. This is the first time an in vitro human-on-a-chip system has been shown to predict in vivo outcomes, which could be used to predict clinical trial outcomes in the future.

“The ability to examine PKPD relationships in vitro would enable us to understand compound behavior prior to in vivo testing, offering significant cost and time savings,” said Dr. Shuler, President and CEO, Hesperos, Inc and Professor Emeritus, Cornell University. “We are excited about the potential of this technology to help us ensure that potential new drug candidates have a higher probability of success during the clinical trial process.”

Understanding the inter-relationship between pharmacokinetics (PK), the drug’s time course for absorption, distribution, metabolism and excretion, and PD, the biological effect of a drug, is crucial in drug discovery and development. Scientists have learned that the maximum drug effect is not always driven by the peak drug concentration. In some cases, time is a critical factor influencing drug effect, but often this concentration-effect-time relationship only comes to light during the advanced stages of the preclinical program. In addition, often the data cannot be reliably extrapolated to humans.

“It is costly and time consuming to discover that potential drug candidates may have poor therapeutic qualities preventing their onward progression,” said James Hickman, Chief Scientist at Hesperos and Professor at the University of Central Florida. “Being able to define this during early drug discovery will be a valuable contribution to the optimization of potential new drug candidates.”

As demonstrated with the terfenadine experiment, the PKPD modelling approach was critical for understanding both the flux of compound between compartments as well as the resulting PD response in the context of dynamic exposure profiles of both parent and metabolite, as indicated by Dr. Shuler.

In order to test the viability of their system in a real-world drug discovery setting, the Hesperos team collaborated with scientists at AstraZeneca, to test one of their failed small molecules, known to have a CV [cardiovscular?] risk.

One of the main measurements used to assess the electrical properties of the heart is the QT interval, which approximates the time taken from when the cardiac ventricles start to contract to when they finish relaxing. Prolongation of the QT interval on the electrocardiogram can lead to a fatal arrhythmia known as Torsade de Pointes. Consequently, it is a mandatory requirement prior to first-in-human administration of potential new drug candidates that their ability to inhibit the hERG channel (a biomarker for QT prolongation) is investigated.

In the case of the AstraZeneca molecule, the molecule was assessed for hERG inhibition early on, and it was concluded to have a low potential to cause in vivo QT prolongation up to 100 μM. In later pre-clinical testing, the QT interval increased by 22% at a concentration of just 3 μM. Subsequent investigations found that a major metabolite was responsible. Hesperos was able to detect a clear PD effect at concentrations above 3 μM and worked to determine the mechanism of toxicity of the molecule.

The ability of these systems to assess cardiac function non-invasively in the presence of both parent molecule and metabolite over time, using multiplexed and repeat drug dosing regimes, provides an opportunity to run long-term studies for chronic administration of drugs to study their potential toxic effects.

Hesperos, Inc. is the first company spun out from the Tissue Chip Program at NCATS (National Center for Advancing Translational Sciences), which was established in 2011 to address the long timelines, steep costs and high failure rates associated with the drug development process. Hesperos currently is funded through NCATS’ Small Business Innovation Research program to undertake these studies and make tissue chips technology available as a service based company.

“The application of tissue chip technology in drug testing can lead to advances in predicting the potential effects of candidate medicines in people,” said Danilo Tagle, Ph.D., associate director for special initiatives at NCATS.

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About Hesperos
Hesperos, Inc. is a leader in efforts to characterize an individual’s biology with human-on-a-chip microfluidic systems. Founders Michael L. Shuler and James J. Hickman have been at the forefront of every major scientific discovery in this realm, from individual organ-on-a-chip constructs to fully functional, interconnected multi-organ systems. With a mission to revolutionize toxicology testing as well as efficacy evaluation for drug discovery, the company has created pumpless platforms with serum-free cellular mediums that allow multi-organ system communication and integrated computational PKPD modeling of live physiological responses utilizing functional readouts from neurons, cardiac, muscle, barrier tissues and neuromuscular junctions as well as responses from liver, pancreas and barrier tissues. Created from human stem cells, the fully human systems are the first in vitro solutions that accurately utilize in vitro systems to predict in vivo functions without the use of animal models, as featured in Science. More information is available at http://www.
hesperosinc.com

Years ago I went to a congress focused on alternatives to animal testing (August 22, 2014 posting) and saw a video of heart cells in a petri dish (in vitro) beating in a heartlike rhythm. It was something like this,

ipscira
Published on Oct 17, 2010 https://www.youtube.com/watch?v=BqzW9Jq-OVA

I found it amazing as did the scientist who drew my attention to it. After, it’s just a collection of heart cells. How do they start beating and keep time with each other?

Getting back to the latest research, here’s a link and a citation for the paper,

On the potential of in vitro organ-chip models to define temporal pharmacokinetic-pharmacodynamic relationships by Christopher W. McAleer, Amy Pointon, Christopher J. Long, Rocky L. Brighton, Benjamin D. Wilkin, L. Richard Bridges, Narasimham Narasimhan Sriram, Kristin Fabre, Robin McDougall, Victorine P. Muse, Jerome T. Mettetal, Abhishek Srivastava, Dominic Williams, Mark T. Schnepper, Jeff L. Roles, Michael L. Shuler, James J. Hickman & Lorna Ewart. Scientific Reports volume 9, Article number: 9619 (2019) DOI: https://doi.org/10.1038/s41598-019-45656-4 Published: 03 July 2019

This paper is open access.

I happened to look at the paper and found good definitions of pharmacokinetics and pharmacodynamics. I know it’s not for everyone but if you’ve ever been curious about the difference (from the Introduction of On the potential of in vitro organ-chip models to define temporal pharmacokinetic-pharmacodynamic relationships),

Integrative pharmacology is a discipline that builds an understanding of the inter-relationship between pharmacokinetics (PK), the drug’s time course for absorption, distribution, metabolism and excretion and pharmacodynamics (PD), the biological effect of a drug. In drug discovery, this multi-variate approach guides medicinal chemists to modify structural properties of a drug molecule to improve its chance of becoming a medicine in a process known as “lead optimization”.

…

*More than one person and more than one company and more than one country claims pioneer status where ‘human-on-a-chip’ is concerned.

Two-organ tests (body-on-a-chip) show liver damage possible from nanoparticles

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This is the first time I’ve seen testing of two organs for possible adverse effects from nanoparticles. In this case, the researchers were especially interested in the liver. From an Aug. 12, 2014 news item on Azonano,

Nanoparticles in food, sunscreen and other everyday products have many benefits. But Cornell [University] biomedical scientists are finding that at certain doses, the particles might cause human organ damage.

A recently published study in Lab on a Chip by the Royal Society of Chemistry and led by senior research associate Mandy Esch shows that nanoparticles injure liver cells when they are in microfluidic devices designed to mimic organs of the human body. The injury was worse when tested in two-organ systems, as opposed to single organs – potentially raising concerns for humans and animals.

Anne Ju’s Aug. 11, 2014 article for Cornell University’s Chronicle describes the motivation for this work and the research itself in more detail,

“We are looking at the effects of what are considered to be harmless nanoparticles in humans,” Esch said. “These particles are not necessarily lethal, but … are there other consequences? We’re looking at the non-lethal consequences.”

She used 50-nanometer carboxylated polystyrene nanoparticles, found in some animal food sources and considered model inert particles. Shuler’s lab specializes in “body-on-a-chip” microfluidics, which are engineered chips with carved compartments that contain cell cultures to represent the chemistry of individual organs.

In Esch’s experiment, she made a human intestinal compartment, a liver compartment and a compartment to represent surrounding tissues in the body. She then observed the effects of fluorescently labeled nanoparticles as they traveled through the system.

Esch found that both single nanoparticles as well as small clusters crossed the gastrointestinal barrier and reached liver cells, and the liver cells released an enzyme called aspartate transaminase, known to be released during cell death or damage.

It’s unclear exactly what damage is occurring or why, but the results indicate that the nanoparticles must be undergoing changes as they cross the gastrointestinal barrier, and that these alterations may change their toxic potential, Esch said. Long-term consequences for organs in proximity could be a concern, she said.

“The motivation behind this study was twofold: one, to show that multi-organ, in vitro systems give us more information when testing for the interaction of a substance with the human body, and two … to look at nanoparticles because they have a huge potential for medicine, yet adverse effects have not been studied in detail yet,” Esch said.

Mary Macleod’s July 3, 2014 article for Chemistry World features a diagram of the two-organ system and more technical details about the research,

Schematic of the two-organ system [downloaded from http://www.rsc.org/chemistryworld/2014/07/nanoparticle-liver-gastrointestinal-tract-microfluidic-chip]

Schematic of the two-organ system [downloaded from http://www.rsc.org/chemistryworld/2014/07/nanoparticle-liver-gastrointestinal-tract-microfluidic-chip]

HepG2/C3A cells were used to represent the liver, with the intestinal cell co-culture consisting of enterocytes (Caco-2) and mucin-producing (HT29-MTX) cells. Carboxylated polystyrene nanoparticles were fluorescently labelled so their movement between the chambers could be tracked. Levels of aspartate transaminase, a cytosolic enzyme released into the culture medium upon cell death, were measured to give an indication of liver damage.

The study saw that single nanoparticles and smaller nanoparticle aggregates were able to cross the GI barrier and reach the liver cells. The increased zeta potentials of these nanoparticles suggest that crossing the barrier may raise their toxic potential. However, larger nanoparticles, which interact with cell membranes and aggregate into clusters, were stopped much more effectively by the GI tract barrier.

The gastrointestinal tract is an important barrier preventing ingested substances crossing into systemic circulation. Initial results indicate that soluble mediators released upon low-level injury to liver cells may enhance the initial injury by damaging the cells which form the GI tract. These adverse effects were not seen in conventional single-organ tests.

Here’s a link to and a citation for the paper,

Body-on-a-chip simulation with gastrointestinal tract and liver tissues suggests that ingested nanoparticles have the potential to cause liver injury by Mandy B. Esch, Gretchen J. Mahler, Tracy Stokol, and Michael L. Shuler. Lab Chip, 2014,14, 3081-3092 DOI: 10.1039/C4LC00371C First published online 27 Jun 2014

This paper is open access until Aug. 12, 2014.

While this research is deeply concerning, it should be noted the researchers are being very careful in their conclusions as per Ju’s article, “It’s unclear exactly what damage is occurring or why, but the results indicate that the nanoparticles must be undergoing changes as they cross the gastrointestinal barrier, and that these alterations may change their toxic potential … Long-term consequences for organs in proximity could be a concern … .”

Could nanoparticles in your mouthwash affect for your cells?

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The first news item I’m going to highlight was posted on Nanowerk, March 8, 2012 and is focused on the use of silver nanoparticles in mouthwashes and dentures to prevent yeast infections,

Yeasts which cause hard-to-treat mouth infections are killed using silver nanoparticles in the laboratory, scientists have found. These yeast infections, caused by Candida albicans and Candida glabrata target the young, old and immuno-compromised. Professor Mariana Henriques, University of Minho [Portugal], and her colleagues hope to test silver nanoparticles in mouthwash and dentures as a potential preventative measure against these infections.

Professor Henriques and her team, who published their research in the Society for Applied Microbiology’s journal Letters in Applied Microbiology(“Silver nanoparticles: influence of stabilizing agent and diameter on antifungal activity against Candida albicans and Candida glabrata biofilms”), looked at the use of different sizes of silver nanoparticles to determine their anti-fungal properties …

The scientists used artificial biofilms in conditions which mimic those of saliva as closely as possible. They then added different sizes and concentrations of silver nanoparticles and found that different sizes of nanoparticles were equally effective at killing the yeasts. Due to the diversity of the sizes of nanoparticles demonstrating anti-fungal properties the researchers hope this will enable the nanoparticles to be used in many different applications.

Some researchers have expressed concerns around the safety of nanoparticle use but the authors stress this research is at an early stage and extensive safety trials will be carried out before any product reaches the market. [emphasis mine]

Following on the notion of safety and gargling silver nanoparticles, coincidentally, there was another news item also dated March 8, 2012 on Nanowerk, this one about the impact that nanoparticles may have on nutrient uptake,

Nanoparticles are everywhere. From cosmetics and clothes, to soda and snacks. But as versatile as they are, nanoparticles also have a downside, say researchers at Binghamton University and Cornell University in a recent paper published in the journal Nature Nanotechnology (“Oral exposure to polystyrene nanoparticles affects iron absorption”). These tiny particles, even in low doses, could have a big impact on our long-term health.

According to lead author of the article, Gretchen Mahler, assistant professor of bioengineering at Binghamton University, much of the existing research on the safety of nanoparticles has been on the direct health effects. But what Mahler, Michael L. Shuler of Cornell University and a team of researchers really wanted to know was what happens when someone gets constant exposure in small doses – the kind you’d get if you were taken a drug or supplement that included nanoparticles in some form. [e.g. silver nanoparticles in your mouthwash or on your dentures]

“We thought that the best way to measure the more subtle effects of this kind of intake was to monitor the reaction of intestinal cells,” said Mahler. “And we did this in two ways: in vitro, through human intestinal-lining cells that we had cultured in the lab; and in vivo, through the intestinal linings of live chickens. Both sets of results pointed to the same thing – that exposure to nanoparticles influences the absorption of nutrients into the bloodstream.”

As for why the researchers focused on iron and tested polystyrene nanoparticles (from the news item),

The uptake of iron, an essential nutrient, was of particular interest due to the way it is absorbed and processed through the intestines. The way Mahler and the team tested this was to use polystyrene nanoparticles because of its easily traceable fluorescent properties.

“What we found was that for brief exposures, iron absorption dropped by about 50 percent,” said Mahler. “But when we extended that period of time, absorption actually increased by about 200 percent. It was very clear – nanoparticles definitely affects iron uptake and transport.”

While acute oral exposure caused disruptions to intestinal iron transport, chronic exposure caused a remodeling of the intestinal villi – the tiny, finger-like projections that are vital to the intestine’s ability to absorb nutrients – making them larger and broader, thus allowing iron to enter the bloodstream much faster.

As to whether these changes are good or bad the researchers don’t speculate. They do have plans for more testing,

calcium,
copper,
zinc, and
fat-soluble vitamins A, D, E and K

They don’t mention any changes in the types of nanoparticles they might be testing in future.

In any event, our bodies have changed a lot over the centuries, you just have to visit a pyramid in Egypt or a museum that holds medieval armour to observe that humans were once much shorter than we are today.