Microphysiological systems: analysis of the current status, challenges and commercial future

The field of microphysiological systems (or organs-on-a-chip) experienced, in the past decade, a surge in publications and efforts towards commercialization. Such systems hold the promise to advance drug discovery, diagnostics, and many other areas. In this review we summarize and analyze the current status of the field, describe the commercial advances and discuss standing challenges and the commercial outlook of the field

Organs-on-Chips in Drug Development: The Importance of Involving Stakeholders in Early Health Technology Assessment

Organs-on-chips are three-dimensional, microfluidic cell culture systems that simulate the function of tissues and organ subunits. Organ-on-chip systems are expected to contribute to drug candidate screening and the reduction of animal tests in preclinical drug development and may increase efficiency of these processes. To maximize the future impact of the technology on drug development, it is important to make informed decisions regarding the attributes and features of organs-on-chips even though the technology is still in a stage of early development. It is likely that different stakeholders in organ-on-chip development, such as engineers, biologists, regulatory scientists, and pharmaceutical researchers, will have different perspectives on how to maximize the future impact of the technology. Various aspects of organ-on-chip development, such as cost, materials, features, cell source, read-out technology, types of data, and compatibility with existing technology, will likely be judged differently by different stakeholders. Early health technology assessment (HTA) is needed in order to facilitate the essential integration of such potentially conflicting views in the process of technology development. In this critical review we discuss the potential impact of organs-on-chips on the drug development process, and we use a pilot study to give examples of how different stakeholders have different perspectives on attributes of organ-on-chip technology. As a future tool in early HTA of organs-on-chips, we suggest the use of multicriteria decision analysis (MCDA), which is a formal method to deal with multiple and conflicting criteria in technology development. We argue that it is essential to design and perform a comprehensive MCDA for organ-on-chip development, and so the future impact of this technology in the pharmaceutical industry can be maximized

Allometric scaling in-vitro

About two decades ago, West and coworkers established a model which predicts that metabolic rate follows a three quarter power relationship with the mass of an organism, based on the premise that tissues are supplied nutrients through a fractal distribution network. Quarter power scaling is widely considered a universal law of biology and it is generally accepted that were in-vitro cultures to obey allometric metabolic scaling, they would have more predictive potential and could, for instance, provide a viable substitute for animals in research. This paper outlines a theoretical and computational framework for establishing quarter power scaling in three-dimensional spherical constructs in-vitro, starting where fractal distribution ends. Allometric scaling in non-vascular spherical tissue constructs was assessed using models of Michaelis Menten oxygen consumption and diffusion. The models demonstrate that physiological scaling is maintained when about 5 to 60% of the construct is exposed to oxygen concentrations less than the Michaelis Menten constant, with a significant concentration gradient in the sphere. The results have important implications for the design of downscaled in-vitro systems with physiological relevance

Characterization of Electromagnetic Valveless Micropump

This paper presents an electromagnetically-actuated micropump for microfluidic application. The system comprises two modules; an electromagnetic actuator module and a diffuser module. Fabrication of the diffuser module can be achieved using photolithography process with a master template and a PDMS prepolymer as the structural material. The actuator module consists of two power inductors and two NdFeB permanent magnets placed between the diffuser elements. The choice of this actuation principle merits from low operating voltage (1.5 Vdc) and the flow direction can be controlled by changing the orientation of the magnet vibration. Maximum volumetric flow rate of the fabricated device at zero backpressure is 0.9756 µLs-1 and 0.4659 µLs-1 at the hydrostatic backpressure of 10 mmH2O at 9 Hz of switching speed

Analysis of methods for physical and biological characterization and validation of microphysiological systems (MPSs)

Microphysiological systems (MPSs), also known as 'Organ-On-a-Chip' (OCC), have revolutionized the way of understanding biology. These systems, which include three-dimensional co-culture and microfluidic technology, aim to mimic human physiology with in-vitro culture systems. Their purpose is to increase the knowledge of biological processes, as well as to provide an effective diagnostic tool for the analysis of different drugs. Standardization of MPSs implies the robustness and reproducibility of these devices and is desirable for their industrialization, production and regularization. However, due to the early stage of academic and commercial development of this technology, no standardization procedure exists in the literature to date. Therefore, experts recommend focusing on the characterization or qualification of these devices. This characterization and qualification of microphysiological systems involves testing the different elements that make up the device to ensure that their configuration mimics the physiology of the human structures represented, behaving and providing values as similar as possible to those of the tissues in vivo. It is in this context that this thesis attempts to develop a characterization protocol applicable to any 'Organ-On-a-Chip' system based on the tests carried out and compiled in the literature of devices in the experimental or commercialization phase. Specifically, a characterization and qualification procedure is presented in which the membrane permeability is monitored in real time depending on device elements such as the presence or not of cell culture, the application or not of microfluids, among others. The choice of the assays to be performed, from among those described in the protocol, will depend on the elements of the OCC to be characterized.Els sistemes microfisiològics (MPSs), també coneguts com a "Organ-On-a-Chip" (OCC), han revolucionat la manera d'entendre la biologia. Aquests sistemes, que inclouen el co-cultiu tridimensional i la tecnologia microfluídica, pretenen imitar la fisiologia humana amb els sistemes de cultiu in-vitro. El seu objectiu és augmentar el coneixement dels processos biològics, així com proporcionar una eina de diagnòstic eficaç per a l'anàlisi de diferents fàrmacs. L'estandardització de MPSs implica la robustesa i reproductibilitat d'aquests dispositius i és desitjable per a la seva industrialització, producció i regularització. No obstant això, a causa de la primera etapa del desenvolupament acadèmic i comercial d'aquesta tecnologia, no existeix cap procediment d'estandardització en la literatura fins a data d’avui. Per tant, els experts recomanen centrar-se en la caracterització o qualificació d'aquests dispositius. Aquesta caracterització i qualificació de sistemes microfisiològics implica provar els diferents elements que componen el dispositiu per assegurar que la seva configuració imiti la fisiologia de les estructures humanes representades, comportant-se i proporcionant valors el més similars possible als dels teixits in-vivo. És en aquest context que aquesta tesi intenta desenvolupar un protocol de caracterització aplicable a qualsevol sistema "Organ-On-a-Chip" basat en les proves realitzades i compilades en la literatura de dispositius en la fase experimental o de comercialització. Concretament, es presenta un procediment de caracterització i qualificació en el qual la permeabilitat de la membrana es controla en temps real depenent dels elements del dispositiu com la presència o no del cultiu cel·lular, l'aplicació o no de microfluids, entre d'altres. L'elecció dels assaigs a realitzar, d'entre els descrits en el protocol, dependrà dels elements de l'OCC que el caracteritzin.Los sistemas microfisiológicos (MPSs), también conocidos como "Organ-On-a-Chip" (OCC), han revolucionado la forma de entender la biología. Estos sistemas, que incluyen el co-cultivo tridimensional y la tecnología microfluídica, pretenden imitar la fisiología humana con sistemas de cultivo in vitro. Su finalidad es aumentar el conocimiento de los procesos biológicos, así como proporcionar una herramienta de diagnóstico eficaz para el análisis de diferentes fármacos. La estandarización de los MPSs implica la robustez y reproducibilidad de estos dispositivos y es deseable para su industrialización, producción y regularización. Sin embargo, debido a la temprana etapa de desarrollo académico y comercial de esta tecnología, hasta la fecha no existe en la literatura ningún procedimiento de estandarización. Por ello, los expertos recomiendan centrarse en la caracterización o cualificación de estos dispositivos. Esta caracterización y cualificación de los sistemas microfisiológicos implica probar los diferentes elementos que componen el dispositivo para asegurar que su configuración imita la fisiología de las estructuras humanas representadas, comportándose y proporcionando valores lo más similares posibles a los de los tejidos in vivo. Es en este contexto en el que esta tesis trata de desarrollar un protocolo de caracterización aplicable a cualquier sistema 'Organ-On-a-Chip' basado en las pruebas realizadas y recopiladas en la literatura de dispositivos en fase experimental o de comercialización. En concreto, se presenta un procedimiento de caracterización y cualificación en el que se monitoriza en tiempo real la permeabilidad de la membrana en función de elementos del dispositivo como la presencia o no de cultivo celular, la aplicación o no de microfluidos, entre otros. La elección de los ensayos a realizar, de entre los descritos en el protocolo, dependerá de los elementos del OCC a caracterizar.Outgoin

Microfluidic Organ/Body-on-a-Chip Devices at the Convergence of Biology and Microengineering

Recent advances in biomedical technologies are mostly related to the convergence of biology with microengineering. For instance, microfluidic devices are now commonly found in most research centers, clinics and hospitals, contributing to more accurate studies and therapies as powerful tools for drug delivery, monitoring of specific analytes, and medical diagnostics. Most remarkably, integration of cellularized constructs within microengineered platforms has enabled the recapitulation of the physiological and pathological conditions of complex tissues and organs. The so-called organ-on-a-chip technology, which represents a new avenue in the field of advanced in vitro models, with the potential to revolutionize current approaches to drug screening and toxicology studies. This review aims to highlight recent advances of microfluidic-based devices towards a body-on-a-chip concept, exploring their technology and broad applications in the biomedical field.European Regional Development Fund-Project FNUSA-ICRC [CZ.1.05/1.1.00/02.0123]; Fundacao para a Ciencia e a Tecnologia (FCT), Portugal [UID/BIM/04773/2013]; Internal Research Grant Program, Universita Campus Bio-Medico di Romainfo:eu-repo/semantics/publishedVersio

A strategy for integrating essential three-dimensional microphysiological systems of human organs for realistic anticancer drug screening

Cancer is one of the leading causes of morbidity and mortality around the world. Despite some success, traditional anticancer drugs developed to reduce tumor growth face important limitations primarily due to undesirable bone marrow and cardiovascular toxicity. Many drugs fail in clinical development after showing promise in preclinical trials, suggesting that the available in vitro and animal models are poor predictors of drug efficacy and toxicity in humans. Thus, novel models that more accurately mimic the biology of human organs are necessary for high-throughput drug screening. Three-dimensional (3D) microphysiological systems can utilize induced pluripotent stem cell technology, tissue engineering, and microfabrication techniques to develop tissue models of human tumors, cardiac muscle, and bone marrow on the order of 1 mm3 in size. A functional network of human capillaries and microvessels to overcome diffusion limitations in nutrient delivery and waste removal can also nourish the 3D microphysiological tissues. Importantly, the 3D microphysiological tissues are grown on optically clear platforms that offer non-invasive and non-destructive image acquisition with subcellular resolution in real time. Such systems offer a new paradigm for high-throughput drug screening and will significantly improve the efficiency of identifying new drugs for cancer treatment that minimize cardiac and bone marrow toxicity

Vascularization in organ- and body-on-a-chip platforms

In tissue culture, soluble molecules such as gases, nutrients and waste are transported by diffusion. This limits the size of the tissue: if the dimensions of the tissue construct exceed the diffusion range, the inner parts of the tissue are deprived of nutrients. In the human body, long distance transport is covered by blood circulation. Vascularization of cultured tissue can help achieve adequate mass transport throughout tissue engineered constructs. Organ- and body-on-a-chip platforms are cell culture devices based on microfluidics, i.e., manipulation of small volumes of fluids. In comparison to traditional cell and tissue culture, the on-chip devices allow precise control of environmental parameters, such as perfusion of medium, and require only small amounts of reagents. The development of vascular networks is largely dependent on blood flow. Therefore, organ-on-a-chip platforms with controlled flow conditions are especially suitable in studies involving vascularization. This bachelor’s thesis discusses methods to build vasculature in organ-on-a-chip platforms. One of the two main approaches is guided self-organization of endothelial cells into vascular networks, mimicking the angiogenic processes in vivo. In the other approach, a pre-patterned blood vessel scaffold is fabricated first and then seeded with cells. In addition to research on blood vessel development and function, vascularized organ-on-a-chip devices can be applied in improving in vitro organ models and connecting multiple models together as body-on-a-chip platforms. Because nearly every human tissue type includes vasculature, it can be considered an essential component in all future in vitro models of human biology.Kudosviljelyssä liukoiset molekyylit, kuten ravinteet, kaasut ja metaboliajäte, kulkeutuvat diffuusion avulla. Tämä rajoittaa kudosviljelmän kokoa: diffuusio ei ulotu liian suuren kudoskappaleen sisäosiin, jolloin ne eivät saa tarvitsemiaan ravinteita. Elimistössä molekyylit kulkeutuvat pitkiä matkoja verenkierron välityksellä. Vaskularisaatio eli verisuonituksen lisääminen kudosviljelmään edesauttaa tehokasta ravinteiden ja kaasujen kuljetusta. Kudos- ja monikudosmallinnuksella (engl. organ- ja body-on-a-chip) tarkoitetaan mikrofluidistiikkaan perustuvia solukasvatuslaitteistoja. Verrattuna perinteiseen soluviljelyyn niiden etuja ovat kasvatusolosuhteiden, kuten nestevirtauksen, tarkka säätely sekä pieni reagenssien kulutus. Veren virtaus ja sen aiheuttamat voimat säätelevät verisuoniston kehitystä, minkä vuoksi organ-on-a-chip-laitteistot soveltuvat hyvin verisuonitettujen kudosmallien kasvattamiseen. Kandidaatintyö selvittää ja kuvaa keinoja, joilla verisuonitusta voidaan rakentaa organ-on-a-chip-malleihin. Pääasiallisia lähestymistapoja on kaksi: endoteelisoluja voidaan ohjata järjestäytymään itse verkostoksi mukaillen luonnollista angiogeneesiä, tai solut voidaan istuttaa valmiiksi muotoiltuun verisuonistoa muistuttavaan tukirakenteeseen. Verisuoniston kehityksen ja toiminnan tutkimisen lisäksi verisuonitettuja organ-on-a-chip-laitteistoja voidaan hyödyntää kudosmallien elinkelpoisuuden ja toiminnallisuuden parantamiseen sekä useiden elinmallien yhdistämiseen. Koska lähes jokaisessa ihmisen kudostyypissä on verisuonitusta, vaskularisaatiota olisi hyvä hyödyntää yhä useammissa in vitro -kudosmalleissa

Technological advances for analyzing the content of organ-on-a-chip by mass spectrometry

Three-dimensional (3D) cell cultures, including organ-on-a-chip (OOC) devices, offer the possibility to mimic human physiology conditions better than 2D models. The organ-on-a-chip devices have a wide range of applications, including mechanical studies, functional validation, and toxicology investigations. Despite many advances in this field, the major challenge with the use of organ-on-a-chips relies on the lack of online analysis methods preventing the real-time observation of cultured cells. Mass spectrometry is a promising analytical technique for real-time analysis of cell excretes from organ-on-a-chip models. This is due to its high sensitivity, selectivity, and ability to tentatively identify a large variety of unknown compounds, ranging from metabolites, lipids, and peptides to proteins. However, the hyphenation of organ-on-a-chip with MS is largely hampered by the nature of the media used, and the presence of nonvolatile buffers. This in turn stalls the straightforward and online connection of organ-on-a-chip outlet to MS. To overcome this challenge, multiple advances have been made to pre-treat samples right after organ-on-a-chip and just before MS. In this review, we summarised these technological advances and exhaustively evaluated their benefits and shortcomings for successful hyphenation of organ-on-a-chip with MS

Valoaktivoituvien ICG-Doksorubisiini-liposomien valmistaminen ja Quasi-Vivo® -pohjaisen kaksisolumallin kehittäminen lääkkeiden teho- ja toksisuuskokeisiin

Perinteiset 2D-solunkasvatusmenetelmät ja kokeelliset alustat eivät usein pysty simuloimaan eri solutyyppien luonnollista kemiallista ja fysiologista ympäristöä. Tekijöitä, jotka voivat vaikuttaa merkittävästi solujen erilaistumiseen, kasvuun, solunsisäisiin rakenteisiin tai metaboliseen aktiivisuuteen ovat esimerkiksi hapen saatavuus, viestiaineet, kemialliset gradientit, paine, nesteen virtaus ja alustojen topografia. Modulaarisia bioreaktoreita, kuten Quasi-Vivo®-järjestelmää, voidaan käyttää simuloimaan näitä tekijöitä. Liposomit ovat fosfolipidikaksoiskerroksesta muodostuvia partikkeleja, joiden sisällä on vesitilavuus. Niitä voidaan muokata monin eri tavoin, lataamalla niitä kuljettamaan vesi- tai rasvaliukoisia molekyylejä, muokkaamalla niiden transitiolämpötilaa, tai päällystämällä niitä eri tarpeiden mukaan. Doksorubisiini on tehokas ja yhä laajassa käytössä oleva sytostaatti, jolla kuitenkin vapaana lääkeaineena annosteltuna on vakavia haittoja, erityisesti sydäntoksisuus. Tässä työssä tavoitteena on selvittää sopivat valmistusparametrit ja todeta riittävä säilyvyys valoaktivoituville ICG-Doksorubisiini-liposomeille, jotta niitä voidaan käyttää tulevissa in vitro kokeissa. Tämän lisäksi selvitetään HepG2 solulinjan selviäminen virtauksen alla Quasi-Vivo® -laitteistossa ja yhdistetään HepG2 ja A549 solulinjat yhdeksi kaksisolumalliksi. Lopuksi suoritetaan yksinkertainen valotuskoe aiemmin valmistetuilla liposomeilla tässä solumallissa, ja tarkastellaan, miten vaikutus näkyy koko systeemissä. Liposomien, joiden ICG- ja doksorubisiini-enkapsulaatio on yli 70%, valmistaminen onnistuu esitetyllä protokollalla luotettavasti ja toistettavasti, ja nämä liposomit säilyvät käyttökelpoisina ainakin 14 vuorokautta säilytettynä pimeässä, 4°C lämpotilassa. A549 ja HepG2 solulinjojen kasvattaminen ja yhdistäminen samaan laitteistoon yhteiseen kasvatusliuokseen virtauksen alle onnistuu, eikä kummankaan solulinjan kasvussa huomata eroa viljelyyn staattisissa olosuhteissa. Kun valotetaan laitteistoon annosteltuja liposomeja, huomataan alustavien tulosten perusteella merkittävää tehon lisäystä valotetussa järjestelmässä pimeään verrattuna, sekä valotetuissa kammiossa että niissä, jotka siihen on Quasi-Vivo® -putkiston kautta yhdistetty.Traditional 2D cell cultivating vessels and experimental models cannot often simulate natural chemical and physical environment of different cell types. For example, availability of oxygen, chemical gradients, messaging molecules, fluid pressure, flow and surface topography are factors that may affect significantly in cell differentiation, growth, cellular structure, and metabolism. Modular bioreactors like Quasi-Vivo® -system can be used to simulate these factors. Liposomes are particles of phospholipid bilayer with aqueous space enclosed within. They can be modified in numerous ways, like loading them with hydrophobic and hydrophilic molecules, changing their transition temperature or coating them according to different needs. Doxorubicin is effective and widely used cytostatic agent, but when administered as a free drug it has often severe side-effects, like cardiotoxicity. Goal of this thesis is to determine appropriate manufacturing parameters and verify adequate shelf-life of ICG-Doxorubicin liposomes, that they are applicable for future in vitro experiments. Then survival of HepG2 cell line under flow in Quasi-Vivo®-equipment is determined, after which A549 and HepG2 will be then combined into one two-cell model. Finally, a simple illumination experiment in this cell model with previously made liposomes is conducted, and the effect in whole system is examined. Using protocol presented in this thesis it is possible to produce successfully and repeatedly liposomes with both ICG and doxorubicin encapsulation over 70%. Their shelf-life was at least 14 days when stored in 4°C protected from light. This was determined to be sufficient for in vitro testing. Cultivating A549 and HepG2 cell lines combined in the same system with shared media and fluid flow conditions was successful. Neither of the cell lines show significant difference in viability when compared to static control. When light-activating liposomes are administered to the system and then illuminated, from preliminary results we can see significant difference in drug effect. Both illuminated chambers and off-target chambers connected via Quasi-Vivo® show increased suppression, which shows promise that this in vitro model would be useful for future experiments