Financial potential is being matched medical potential, as microenvironments play a role in new findings about stem cell treatment that may speed patients’ recovery from chemotherapy and radiation. And microfluidics advances have also led to a new tool for diabetes research.

Bone marrow stem cells produce the body’s blood and immune cells, but chemotherapy and radiation suppress that production, and it can take weeks or months for the blood and immune systems to recover. Results of a recent study by UCLA Broad Stem Cell Research Center scientists showed how a newly developed drug compound might hasten the blood system’s recovery after radiation and chemotherapy in mice. Some of the heretofore unknown capacity of microenvironments were critical to this type of investigation.

“The potency of this compound in animal models was very high,” said study co-author Dr. John Chute. “It accelerated the recovery of blood stem cells, white blood cells and other components of the blood system necessary for survival. If found to be safe in humans, it could lessen infections and allow people to be discharged from the hospital earlier.”

Looking at both mouse — and human stem cells (in dishes) — growth factors were observed to promote the recruitment of stem cells from degraded bone marrow, which fueled proliferation and reconstitution. “We’re very excited about the potential medical applications of these findings,” said Chute.

The new compound enabled the blood system to recover sooner from cancer treatments in mice because it was able to stop the molecular process that slows blood stem cell regeneration. “The new compound lifts molecular ‘brakes’ that normally slow the regeneration of blood stem cells,” according to the team.

The new compound speeded up the regeneration of both mouse and human blood stem cells after exposure to radiation. The bloodstream’s own regenerative capacity seemed to power up healing. Angiogenesis, the formation of new blood cells, is controlled by chemical signals, yet many of the mechanisms by which blood cell regeneration uses bone marrow have been unknown. The study highlights the process by which growth factors recruit stem cells from the bone marrow microenvironment. Advances detailing bone marrow’s microenvironments are a significant contribution.

Microfluidic devices can be monitored and imaged using fluorescent markers for tracking, and microenvironments needed to be precisely controlled for this investigation.

Among the mice that received high doses of radiation, almost all that were given the compound survived; more than half of those that did not receive the compound died. Mice that received chemotherapy but no compound had low levels of white blood cells and neutrophils — which fight bacteria — after two weeks; in mice that were treated, white blood cell counts recovered to normal levels within two weeks. The researchers are now refining the process, in preparation for human trials.

Microfluidics research also has another new arrow in its quiver, in the form of a new tool for diabetes research, developed by a team based at the Harvard Stem Cell Institute. The device improves aspects of studying diabetes, including upgrading the screening process prior to transplantation of insulin-producing cells into a patient.

The team noted that although microfluidic devices have been used to address research limitations in the past, previous adoption has been hampered by “incompatibility of most device materials with large-scale manufacturing. We designed and built a thermoplastic, microfluidic-based Islet-on-a-Chip compatible with commercial fabrication methods, that automates islet loading, stimulation, and insulin sensing.”

The design of their “Islet-on-a-Chip” is based on the human pancreas, where islets process non-stop, incoming information about glucose levels, and adjust insulin production accordingly. The new, automated, miniature device gives results in real-time, which can speed up clinical decision-making.

In addition to diabetes research, the device may prove useful in other areas. Its core technology can be modified to sense an array of microfluidic systems. Because it can detect cell secretions continuously, it can be employed to investigate how cells communicate using protein signals, throughout the body.

“It was exciting to see our lab’s method for measuring islet function taken forward from individual cells to much bigger groups of cells, and incorporated into a device that can be used widely in the community,” said co-author Dr. Michael Roper. “Now, we have a device that integrates glucose delivery, islet positioning and capture, reagent mixing, and insulin detection, and requires far fewer reagents. So labs can use it to do more experiments at the same cost, using a much shorter and easier process.”

Scalable, easier, shorter, and cost-contained. That says it all.

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Kathy Jean Schultz

Kathy Jean Schultz is a freelance medical science writer who focuses on medical innovations. She earned a Master’s Degree in Research Methodology from Hofstra University, and a Master’s Degree in Psychology from Long Island University. She is a member of the National Association of Science Writers, and the Association of Health Care Journalists. Her articles about organoids include "Would you trust a 3-D printed mini organ to test your drugs?" and "Stem cells not only slow disease, they come with their own safety test".

• http://kathyjeanschultz.pressfolios.com/

The post Industry Predictions of Microfluidics Commercialization are Buoyed by Recent Advances in Diabetes and Cancer Research appeared first on The MicroFluidic Circle.

Figure 1. Personalized cancer therapy¹. Credit: The University of Texas MD Anderson Cancer Center

The field of microfluidics has come a long way after the introduction² in the early 1980s and found its application in a wide range of fields like molecular biology, cell biology, genomics, proteomics, astrobiology, optics, and fuel cells. Microfluidics also has shown great promise for cancer diagnosis and in understanding cancer biology with its inherent advantage of high sensitivity, high throughput, less sample consumption, low cost, and enhanced spatio-temporal control³. Microfluidic tools developed over the years for probing cancer have dealt with a small part of the big problem like isolating cancer cells from biopsied tissue sample or patient-derived bio-fluids (blood, pleural effusion), phenotype individual cells in terms of size, shape and deformability, extract the genetic materials like DNA, RNA and proteins from cells and sequencing them, culture cancer cells inside the microfluidic chip, analyze drug dose response etc.  Now is the time we should think of combining the pieces together and develop a complete, automated process for personalized cancer therapy.

An outright microfluidic tool for personalized cancer therapy should integrate all available individual tools and automate the process. This tool should have the capability to analyze single cells at a high throughput fashion to tackle the heterogeneity within a tumor. I believe, it will be surely very challenging but not impossible. Isolating cancer cells from patient samples is the first step of the process. The patient sample is either a piece of biopsied tissue or bio-fluids like blood or pleural effusion. The device should have the capability to isolate cancer cells from those samples. Sanjin et al⁴. has published a nice review article on sample preparation for single-cell analysis in microfluidics. Hattersley et al⁵. has shown the capability of microfluidics in maintaining a viable tissue sample for 72 hours in the device and then dissociate it to get 78±2.4% viable single cells.

Figure 2. Workflow for personalized cancer therapy microfluidic tool

Although the isolation of cancer cells (known as circulating tumor cells, CTCs) from blood is very difficult, mainly because of their insignificant numbers (1-10 CTCs per billions of blood cells in 1 mL of blood), noninvasive, label-free microfluidic tools like labyrinth⁶, vortex⁷ and spiral⁸ chips have shown great outcomes in isolating CTCs.  Since the number of cancer cells is very low in blood we need to choose a technique that can process blood at a high throughput manner with high yield and purity. Labyrinth device seems to be the right choice as it can process 2.5 mL blood per minutes, has a high yield (>90%) and purity (600 WBCs/mL). After separation of cancer cells from the blood next step would be identifying the cancer cells from the background white blood cells (WBCs). Current technique to identify cancer cell is immunostaining which is invasive and kills the cells barring further downstream analysis. To overcome this barrier researchers are trying to develop label-free techniques to identify cancer cells. For example, I am now working with Prof. Siva Vanapalli at Texas Tech University to develop deformability and optical property based techniques using digital holographic microscopy(DHM) to identify cancer cells. Immunostaining aliquot can be another way to handle the problem.

To understand the molecular basis of cancer and to select a specific therapy for a patient we need to combine the phenotypic data (whole cell) with genetic information. So, the next component of the workflow will include tools capable of phenotyping and genotyping at single cell resolution. Deformability cytometers (DCs)^9-11 developed in recent years have the capability to phenotype single cells based on deformability at very high throughput (100 cells/sec) and identify the subpopulations to address heterogeneity. Real-time deformability cytometer (RT-DC)⁹ can generate deformability data for single cells in real-time while multi-sample deformability cytometer (MS-DC)¹¹ can analyze 10 patient samples in a single experiment. The beauty of these deformability cytometers is that they are noninvasive and label-free. Cells collected from the outlet of DC devices can be used for genotyping. Microfluidic tools are also capable of genotyping cells as successfully demonstrated by Sohni et al.¹² and Chang et al.¹³

Another component of the microfluidic tool for personalized cancer therapy should have is drug response analysis to avoid unnecessary or ineffective treatment. A study shows that medical waste due to unnecessary or ineffective treatment in the US per year is 75 billion US dollars¹⁴. The microfluidic capability of doing drug response analysis is well established. Bithi et al.¹⁵ isolated single cancer cells into microfluidic chambers and tested the drug response and uptake kinetics. Park et al¹⁶. studied the cellular response to the small-molecule drug within a microfluidic dielectrophoresis device in real-time.

The challenges for combining the individual tools together will be multifold as the operating conditions, sample preparation, addition of reagents and data analysis for each of them are different. Individual devices designed for a specific application may require certain modifications to be compatible with the workflow of personalized cancer therapy tool and real-world applications. Full process will require automation through artificial intelligence (AI), machine learning (ML) and image processing. I believe the day is not too far when we will see microfluidics is delivering as promised.

<sup>1. MD Anderson Cancer Center. Personalized cancer therapy (2018).

2. Terry, S. C., Jerman, J. H. & Angell, J. B. A gas chromatographic air analyzer fabricated on a silicon wafer. IEEE Transactions on Electron Devices 26, 1880-1886, doi:10.1109/T-ED.1979.19791 (1979).

3. Zhang, Z. & Nagrath, S. Microfluidics and cancer: are we there yet? Biomedical microdevices 15, 595-609, doi:10.1007/s10544-012-9734-8 (2013).

4. Hosic, S., Murthy, S. K. & Koppes, A. N. Microfluidic Sample Preparation for Single Cell Analysis. Analytical chemistry 88, 354-380, doi:10.1021/acs.analchem.5b04077 (2016).

5. Hattersley, S. M., Dyer, C. E., Greenman, J. & Haswell, S. J. Development of a microfluidic device for the maintenance and interrogation of viable tissue biopsies. Lab Chip 8, 1842-1846, doi:10.1039/b809345h (2008).

6. Lin, E. et al. High-Throughput Microfluidic Labyrinth for the Label-free Isolation of Circulating Tumor Cells. Cell Systems 5, 295-304.e294, doi:10.1016/j.cels.2017.08.012 (2017).

7. Che, J., Yu, V., Garon, E. B., Goldman, J. W. & Di Carlo, D. Biophysical isolation and identification of circulating tumor cells. Lab on a Chip 17, 1452-1461, doi:10.1039/C7LC00038C (2017).

8. Di Carlo, D. Inertial microfluidics. Lab on a Chip 9, 3038-3046, doi:10.1039/B912547G (2009).

9. Otto, O. et al. Real-time deformability cytometry: on-the-fly cell mechanical phenotyping. Nat Meth 12, 199-202, doi:10.1038/nmeth.3281  (2015).

10. Gossett, D. R. et al. Hydrodynamic stretching of single cells for large population mechanical phenotyping. Proceedings of the National Academy of Sciences 109, 7630-7635, doi:10.1073/pnas.1200107109 (2012).

11. Ahmmed, S. M. et al. Multi-sample deformability cytometry of cancer cells. APL Bioengineering 2, 032002, doi:10.1063/1.5020992 (2018).

12. Sohni, Y. R., Burke, J. P., Dyck, P. J. & O’Kane, D. J. Microfluidic chip-based method for genotyping microsatellites, VNTRs and insertion/deletion polymorphisms. Clinical Biochemistry 36, 35-40, doi:https://doi.org/10.1016/S0009-9120(02)00420-4 (2003).

13. Chang, Y.-M., Ding, S.-T., Lin, E.-C., Wang, L. & Lu, Y.-W. A microfluidic chip for rapid single nucleotide polymorphism (SNP) genotyping using primer extension on microbeads. Sensors and Actuators B: Chemical 246, 215-224, doi:https://doi.org/10.1016/j.snb.2017.01.160 (2017).

14. Lowrey, A. Study of US Health Care System Finds Both Waste and Opportunity to Improve (2012, Sep 11).

15. Bithi, S. S. & Vanapalli, S. A. Microfluidic cell isolation technology for drug testing of single tumor cells and their clusters. Scientific Reports 7, 41707, doi:10.1038/srep41707 (2017).

16. Park, I. S. et al. Real-Time Analysis of Cellular Response to Small-Molecule Drugs within a Microfluidic Dielectrophoresis Device. Anal Chem 87, 5914-5920, doi:10.1021/ac5041549 (2015).</sup>

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Shamim Ahmmed

Shamim Ahmmed is a final year Ph.D. student at microfluidics lab lead by Prof. Siva Vanapalli at Texas Tech University. His training is in Chemical Engineering, microfluidics and image processing. Shamim has unique skills for developing microfluidic tools for phenotyping cancer cells. He led three microfluidic projects from inception to publishing results by collaborating with three cross-functional groups which resulted in multiple publications and presentations. Shamim’s current project involves developing a microfluidic tool that can enumerate circulating tumor cells based on deformability and confirm that through on-chip immunostaining.

• mdsa253@gmail.com
• http://sivaav.wixsite.com/microfluidics

The post Combining the Pieces in Microfluidics for Personalized Cancer Therapy appeared first on The MicroFluidic Circle.

New research suggests microfluidic factors might also determine whether the stem cell is stressed or not stressed. Stem cells that are growing and developing under stress from radiation treatment were observed to differ from stem cells developing in less-stressed environments— that is, with no radiation. A recent animal study suggests stem cells might be able to switch between a “normal-growth” version of themselves, to a “growth-under-stress” version, if that is what the body needs.

Dr. John Chute, of the UCLA Broad Stem Cell Research Center and a professor of hematology/oncology, investigates differences between the microfluidic environments of normal-growth stem cells, compared to those of growth-under-stress stem cells. The Chute team’s goal is to illuminate why normal-growth cells can switch to become growth-under-stress cells, as reported in a recent Cell Stem Cell article. In a press release, Chute observed that “Although the switch occurs, the reason for the change is a mystery.”

Treatments such as radiation and chemotherapy leave human blood-forming cells dangerously stressed and depleted.  A growth-under-stress version of stem cell treatment might be able to heal that depletion much better than a normal-growth version.

The new findings spur questions about whether it might be possible to predict the stress level of stem cells, and to administer the most effective stem cell treatments to radiation patients, in order to speed recovery.

Just as soil nutrients sustain a plant stem, the microfluidic environment surrounding stem cells nurtures them. According to Chute, “In stem cell research, two important questions are, ‘What are the micro-environment cells that regulate stem cells?’ and ‘How do they do it?’” U.S. National Institutes of Health scientists agree: “Microfluidics offers a systematic way to study the decision-making process of stem cells.” In addition, analyses of stem cells based on the microfluidics that nurture them “can be done in a much deeper and wider way” than without them.

NIH scientists have also observed that it is ultimately their microfluidic complexities that predict how stem cells will become one particular body part or another. To gain a precise understanding of how body-part differentiation happens, microfluidic analyses are a necessity.

That necessity is nowhere more evident than in efforts to find out how switching between normal-growth and growth-under-stress stem cells happens. The possible impact on recovery from cancer treatment could be immense. That impact supports industry analyses that the microfluidics market, for which the 2016 global evaluation was $4.76 billion U.S. dollars, will grow to $12.45 billion by 2025.

Numbers tell the story. A surging biotechnology sector paired with the simultaneously increasing global burdens of disease are estimated to drive up market growth. For example, according to the 2017 World Health Organization data, the number of patients suffering from diabetes worldwide was estimated at 422 million in 2014 — and microfluidic advances contribute to innovative diabetes treatments.

The endless frontier of research targets also points to growth on many fronts, from stem-cell stress levels to brain cells, fibrosis and bone joints.

Currently, some new companies that develop stem cells for brain research — deliverable to scientists in both industry and academia — are gaining clients because many facilities do not have the resources to generate neural stem cells themselves. The new companies assemble stem cell types onto microfluidic chips that duplicate human tissue, as well as predict physiological processes. As their novel production challenges are ironed out, these companies will develop and deliver in short time frames.

As reported in a recent Financial Post article, Canada’s Genomics Application Partnership Program (GAPP) supports collaborations specifically to bridge the gap between research and commercialization, and is now funding a $6.5 million microfluidics project to develop fibrosis treatments.

According to U.S. National Institutes of Health scientists, the limitations of a bone joint and cartilage repair are fueling the development of stem cell therapies for weakened cartilage, and this work relies “upon microfluidic technology.”

Predictions about microfluidic commercialization encompass the fact that there were more than 15.5 million cancer survivors in the U.S. in 2016, and this number might be more than 20 million by 2026. About 7 million U.S. patients have had bone treatments such as hip or knee replacements. By 2030, U.S. cartilage-related knee replacement surgeries are projected to total 3.5 million per year.  More than 70,000 people worldwide live with cystic fibrosis.

Analysts need only to do the math.

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Kathy Jean Schultz

Kathy Jean Schultz is a freelance medical science writer who focuses on medical innovations. She earned a Master’s Degree in Research Methodology from Hofstra University, and a Master’s Degree in Psychology from Long Island University. She is a member of the National Association of Science Writers, and the Association of Health Care Journalists. Her articles about organoids include "Would you trust a 3-D printed mini organ to test your drugs?" and "Stem cells not only slow disease, they come with their own safety test".

• http://kathyjeanschultz.pressfolios.com/

The post Microfluidic Environments Nurture Stem Cells on Their Journey Toward Commercialization appeared first on The MicroFluidic Circle.

CTCs are challenging to capture, isolate and characterize in nature. First, CTCs are extremely rare in patients’ blood samples. One CTC usually exists among a background of millions of blood cells. Furthermore, CTCs are highly heterogeneous in physical characteristics and biological properties. No separation technology which is based on a single capture mechanism can produce pure and representative CTC subpopulations. In the traditional liquid biopsy, CTCs are isolated either by immunoaffinity strategies or by biophysical features differentiation. However, existing macro-scale isolation systems suffer important drawbacks, such as low capture efficiency, incomplete automation and low viability of captured CTCs². As a promising alternative, microfluidic technologies have gained tremendous interest in the field. Microfluidic technologies create devices that are at or smaller than the cellular length scale and enable accurate capturing and manipulation at single cell level. These technologies also offer precise control of fluid flow, which can greatly facilitate affinity reactions and physical separation. Moreover, on a microfluidic chip, CTC capturing and next-step analysis can be integrated to minimize intermediate sample handling and shorten the processing time. Above all, microfluidic approaches allow gentle isolation of live cells and thus enable many downstream analyses that rely on captured live CTCs³.

Microfluidic devices apply a wide range of principles for CTC isolations, including those that have been widely used in macro-scale systems such as magnetic-based separation, affinity chromatography and density gradient centrifugation, and novel mechanisms such as microfiltration in 2D and 3D by size and deformability⁴, inertial focusing, electrophoresis, and acoustophoresis. The performance of each isolation strategy is usually assessed by three figures of merits: capture efficiency(how many CTCs captured out of the total number of CTCs), capture purity (the ratio of captured CTCs to background contaminating cells) and throughput (how quickly a device can process a sample). Microfluidic devices have reported greater than 90% efficiency and purity, and throughput higher than 7.5 mL/hr¹. However, there are always trade-offs among these three metrics. Due to the substantial heterogeneity in CTCs, no single isolation principle fits all applications. Combining two or more strategies are required to achieve high efficiency, purity, and throughput.

At the University of California, Los Angeles, Dino Di Carlo, Ph.D., and colleagues have developed High-Throughput Vortex Chip (Vortex-HT) technology, which uses parallel microfluidic vortex chambers to accumulate the larger circulating tumor cells from flowing blood. Vortex-HT reportedly generates less contamination with white blood cells than other technologies and isolates cells in a smaller output volume. Source: GEN

Despite the massive surge of innovation and research development in microfluidic CTC isolation devices in recent years, only a few of them have successfully undergone commercialization and been pushing towards clinical uses. Microfluidics-based CTC isolation systems face several major barriers to commercialization. To begin with, many microfluidic devices operate with complex peripheral fluidic components, such as pumps, pneumatic systems, connectors, and tubing. Their interfaces are usually not user-friendly. How to integrate the fluidic systems and simplify the user interface is the first step to an easy-to-use product⁵. Next, similar to other types of microfluidic systems, many published microfluidic CTC devices suffer from short lifespans due to material degradation, clotting, and fouling problems⁶. The associated device reliability and robustness issues also pose major challenges for commercialization. Last but not least, many microfluidic devices cannot produce high throughput, due to the miniaturized length scales. The ability of processing 7.5 mL of blood samples within a short time is required for CTC clinical applications. Yet few microfluidics-based systems are able to meet this requirement. For these reasons, the advancement of microfluidics-based CTC technologies should focus on developing efficient integration and user-friendly interface, enhancing reliability and robustness, and improving sample throughput as wells as capture efficiency and purity.

The author here shares multiple perspectives on opportunities and challenges of microfluidic technologies for circulating tumor cell analysis. Microfluidics-based CTC technologies have arisen rapidly as promising alternatives to traditional macro-scale CTC isolation and analysis systems. We have discussed the advantages of microfluidics-enabled platforms, current separation principles, and the barriers to successful commercialization. For future development, it is essential to recognize the substantial heterogeneity in CTCs’ characteristics. Isolation strategies should be designed with regard to different cancer types and even sub-types. It is also important to develop performance profiles (efficiency, purity, throughput, viability and etc.) based on cancer types that can allow researchers and clinic users to evaluate and compare different technologies¹. Furthermore, research development and commercialization should continue exploiting the integration advantages of microfluidics systems. As current CTC research focus shifts from enumeration to characterization, integrated microfluidics systems that not only isolate cells but also perform downstream analysis can provide valuable information about cancer progress and metastasis. In addition, these integrated systems can potentially reduce assay cost significantly. It is worth noting that microfluidics systems need to overcome the aforementioned short life spans, reliability and robustness issues. For integrated systems, these issues will become even larger challenges to commercialization. Finally, it is surprising that a macro-scale immunoaffinity system developed by CellResearch remains the only FDA-approved CTC assessment platform since 2004.No other systems or microfluidic devices have been approved by FDA since then. Considering regulatory approvals, the field of microfluidic CTC technologies should drive more clinical relevant tests to increase the number of applications in clinical use and the impact in the medical industry². We look forward to seeing the first and more microfluidics-enabled CTC platforms to obtain FDA approval in the next few years.

<sup>1. Ferreira, M. M., Ramani, V. C. & Jeffrey, S. S. Circulating tumor cell technologies. Mol Oncol 10, 374–394 (2016)

2. Li, P., Stratton, Z. S., Dao, M., Ritz, J. & Huang, T. J. Probing circulating tumor cells in microfluidics. Lab on a Chip 13, 602 (2013)

3. Dong, Y. et al. Microfluidics and Circulating Tumor Cells. J Mol Diagn 15, 149–157 (2012)

4. Au, S. H. et al. Microfluidic isolation of circulating tumor cell clusters by size and asymmetry. Scientific Reports 7, (2017)

5. Shields, C. W., Ohiri, K. A., Szott, L. M. & López, G. P. Translating microfluidics: Cell separation technologies and their barriers to commercialization. Cytometry Part B – Clinical Cytometry 92, 115–125 (2017)

6. Wyatt Shields IV, C., Reyes, C. D. & López, G. P. Microfluidic cell sorting: a review of the advances in the separation of cells from debulking to rare cell isolation. Lab Chip 15, 1230–1249 (2015)</sup>

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Yatian Qu

Dr. Yatian Qu is a microfluidics research scientist at Purigen Biosystems. She is working on developing hands-free solutions for extracting and quantifying nucleic acids from biological samples. She received her M.S. (2012) and Ph.D. (2016) in Mechanical Engineering from Stanford University. She has diverse research experiences in microfluidics, including developing novel electrophoresis techniques to extract nucleic acids and proteins from biological samples and developing high-performance water desalination systems using porous carbon materials. She is also the co-founder of Bay Area Microfluidics Network, an organization that brings together leaders in microfluidics to foster innovation and collaboration in Silicon Valley.

• yatianqu@gmail.com
• http://yatianqu.com/

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