Monitoramento do horizonte tecnológico: MEDICAMENTOS para o tratamento de Doença Falciforme / Monitoring the technological horizon: MEDICINES for the treatment of sickle cell disease

CONDIÇÃO CLÍNICA: O termo doença falciforme representa um grupo de doenças hereditárias, com apresentação multissistêmica, caracterizada por hemácias de formato anormal (em formato de foice) resultante de uma alteração na estrutura da hemoglobina (Hb). As células afetadas apresentam uma maior fragilidade e são removidas da circulação e destruídas. Esse grupo de doenças se manifesta com episódios de agudização e traz repercussões que causam danos progressivos à maioria dos órgãos, incluindo cérebro, rins, pulmões, ossos e sistema cardiovascular. A anemia falciforme, um dos tipos de doença falciforme, é um dos distúrbios monogênicos graves mais comuns em todo o mundo. A anemia falciforme é causada pela presença de hemoglobina-S (HbS), ou hemoglobina falciforme, em homozigose (HbSS). A HbS ocorre quando há a substituição de uma única base nitrogenada no gene que codifica a subunidade beta da hemoglobina (ß-globina), levando a substituição de ácido glutâmico por valina na cadeia dessa proteína. Quando a HbS é desoxigenada, a troca dos aminoácidos gera interação hidrofóbica com outra molécula de hemoglobina, desencadeando uma agregação em grandes polímeros. A polimerização da HbS é o evento primário na patogênese molecular da doença falciforme, o que causa a distorção da forma da hemácia e diminuição acentuada de sua plasticidade com consequente rigidez celular. Essa rigidez prejudica a capacidade de as hemácias transitarem por pequenos vasos, o que causa vaso-oclusão, seguida de isquemia e infarto tecidual. O infarto pode ocorrer em qualquer parte do corpo e é responsável pela manifestação clínica mais precoce: a crise de dor aguda. TRATAMENTO: O gerenciamento da doença falciforme visa a melhora na perfusão tecidual, controle de dor e prevenção, e manejo de complicações associadas à anemia, crise vaso-oclusiva (CVO) e infecções. A terapia modificadora de doença com hidroxiureia é o tratamento mais eficaz para doença falciforme até o momento. O efeito primário da terapia com hidroxiureia está relacionado ao aumento da produção da hemoglobina-fetal (HbF), reduzindo os níveis de HbS e, consequentemente, a falcização de hemácias e vaso-oclusão. De acordo com uma revisão sistemática da Cochrane de estudos clínicos randomizados e quase-randomizados, os benefícios clínicos esperados do tratamento com hidroxiureia incluem a diminuição na frequência de episódios de dor, melhora nos valores de hemoglobina fetal e contagem de neutrófilos, redução de episódios de síndrome torácica aguda e da necessidade de transfusões sanguínea. ESTRATÉGIA DE BUSCA: Uma busca foi realizada no banco de dados eletrônico ClinicalTrials.gov, no dia 28 de março de 2022. O termo empregado foi "Sickle cell" visando a maior sensibilidade da busca. Foram considerados os ensaios clínicos de fase 2/3 e 3 de avaliação de medicamento para o tratamento da doença falciforme. Dos 893 cadastros de estudos, 92 atendiam ao critério de elegibilidade referente à fase da pesquisa. Destes, os estudos em andamento ou concluídos nos últimos cinco anos foram selecionados, restando 35 registros. Desses registros, foram excluídos aqueles que não envolviam medicamentos ou não estavam relacionados à causa-base da doença. Também foram excluídos registros envolvendo o medicamento hidroxiureia, uma vez que esse tratamento já está disponível no SUS. Deste modo, sete substâncias foram selecionadas. Uma oitava tecnologia, a L-glutamina, foi incluída por ter recebido autorização recente da FDA para a indicação abordada neste documento, embora seus registros de estudos não se enquadrem nos critérios de fase ou tempo de início e conclusão em cinco anos. MEDICAMENTOS: TECNOLOGIAS NOVAS: L-glutamina: A L-glutamina é um precursor para a síntese de glutationa, nicotinamida adenina dinucleotídeo (NAD) e arginina, substâncias que protegem os eritrócitos do dano oxidativo. No organismo, o NAD está presente em uma forma oxidada (NAD+) e reduzida (NADH). Ele é um cofator de oxidorredução (redox) onipresente nas hemácias e desempenha um papel central na manutenção do equilíbrio redox. É sabido que o estresse oxidativo contribui na fisiopatologia da doença falciforme. Crizanlizumabe: O crizanlizumabe é um anticorpo monoclonal humanizado seletivo de IgG2 kappa que promove a redução da frequência de CVOs e a inibição das interações multicelulares adesivas mediadas pela P-selectina. Esse medicamento está registrado nas agências pesquisadas, com a indicação para pacientes adultos e pediátricos a partir de 16 anos de idade com doença falciforme. Sua eficácia e segurança foram avaliadas no estudo pivotal SUSTAIN (NCT01895361), um estudo clínico de fase 2, multicêntrico, duplo-cego, randomizado e controlado por placebo, que avaliou 198 pacientes com doença falciforme e com histórico de CVOs, por um período de 52 semanas. Voxelotor: O voxelotor é um inibidor da polimerização da HbS, desenvolvido para o tratamento da doença falciforme em adultos e crianças. No FDA está indicado para crianças de 4 a 11 anos. Na EMA está indicado como opção terapêutica em pacientes a partir de 12 anos de idade. O HOPE (NCT03036813), estudo que embasou os registros nas agências EMA e FDA é um estudo de fase 3, multicêntrico, duplo-cego, randomizado e controlado por placebo, projetado para avaliar a eficácia e segurança do voxelotor. TECNOLOGIAS EMERGENTES: LentiGlobin BB305 (bb1111): A terapia gênica lovotibeglogene autotemcel (LentiGlobin; bb1111), desenvolvida pela bluebird bio®, é um tratamento experimental para a doença falciforme. Até a última atualização deste informe, não havia registro deste medicamento nas agências regulatórias pesquisadas. O FDA, no entanto, concedeu designação de medicamento órfão, fast track, terapia avançada de medicina regenerativa e de doença pediátrica rara para o lovotibeglogene autotemcel para doença falciforme. CTX-001: A CTX-001 é uma terapia celular geneticamente modificada investigada como tratamento potencial voltado para doenças de causas genéticas. O medicamento recebeu designação de droga órfã pela EMA53 e FDA, que também concedeu designação de terapia avançada de medicina regenerativa e fast track54,55. Até a última atualização desse informe, o medicamento não havia recebido registro nas agências pesquisadas. Etavopivat: O etavopivat é um ativador da piruvato quinase eritrocitária (PKR), que está sendo investigado pela Forma Therapeutics® como terapia modificadora de doença para o tratamento da doença falciforme e de outras hemoglobinopatias. Que recebeu designação de droga órfã pelas agências EMA e FDA e fast track pelo FDA60,61. Até a última atualização deste informe, a terapia não possuía registro nas agências pesquisadas. Inclacumabe: O inclacumabe é um anticorpo monoclonal IgG4 totalmente humanizado investigado para a potencial redução da frequência de CVOs causadas pela doença falciforme e internações hospitalares associadas. O medicamento promove a diminuição da formação de agregados plaquetas-leucócitos mediada pela p-selectina, reduzindo a frequência e gravidade das CVOs63,64 . Até a última atualização deste informe, o inclacumabe ainda não possuía registro nas agências FDA e EMA. Mitapivat: O mitapivat é um ativador de piruvato quinase, que recebeu designação de droga órfã pelo FDA para o tratamento da doença falciforme. A ação do mitapivat para o tratamento da doença falciforme baseia-se na ativação alostérica da piruvato quinase de eritrócitos, promovendo a redução dos níveis de 2,3-DPG e da falcização das hemácias. Os resultados sobre o mitapivat publicados até o momento são referentes segurança e tolerabilidade em diferentes doses, obtidos através de um estudo de fase 1, intervencional e não randomizado (NCT04000165), realizado em 17 indivíduos entre 18 e 70 anos de idade, com diagnóstico confirmado de anemia falciforme. Estudos em andamento Os dados dos estudos em andamento e sem resultados publicados sobre o LentiGlobin BB305, crizanlizumabe, voxelotor, etavopivat, inclacumabe e mitapivat. CONSIDERAÇÕES FINAIS: Crizanlizumabe (SEG101), L-glutamina e voxelotor são as três tecnologias recentemente aprovadas pelas agências FDA e/ou EMA para o tratamento da doença falciforme. Esses medicamentos estão associados à boa tolerabilidade apesar da alta incidência de EAs. Seus benefícios clínicos relacionam-se à diminuição de crises de dor, CVOs e aumento dos níveis de hemoglobina. As evidências acerca das demais tecnologias discutidas neste informe devem ser analisadas com cautela uma vez que, mesmo aparentemente bem toleradas e com resultados clínicos que demonstram melhora no quadro da doença, precisam de resultados mais robustos para que seja possível estabelecer um perfil concreto a respeito da segurança e benefícios esperados. Para que ocorra a oferta desses medicamentos no SUS, é necessária a análise pela Conitec, conforme disposto na Lei nº 12.401/2011, que alterou a Lei nº 8.080/1990. Os relatórios de recomendação da Conitec levam em consideração as evidências científicas sobre eficácia, a acurácia, a efetividade e a segurança do medicamento, e, também, a avaliação econômica comparativa dos benefícios e dos custos em relação às tecnologias já incorporadas e o impacto da incorporação da tecnologia no SUS.

Strategies to Overcome the Barrier of Ischemic Microenvironment in Cell Therapy of Cardiovascular Disease.

The transplantation of various immune cell types are promising approaches for the treatment of ischemic cardiovascular disease including myocardial infarction (MI) and peripheral arterial disease (PAD). Major limitation of these so-called Advanced Therapy Medicinal Products (ATMPs) is the ischemic microenvironment affecting cell homeostasis and limiting the demanded effect of the transplanted cell products. Accordingly, different clinical and experimental strategies have been evolved to overcome these obstacles. Here, we give a short review of the different experimental and clinical strategies to solve these issues due to ischemic cardiovascular disease.

Human adipose-derived mesenchymal stem cell-based medical microrobot system for knee cartilage regeneration in vivo.

Targeted cell delivery by a magnetically actuated microrobot with a porous structure is a promising technique to enhance the low targeting efficiency of mesenchymal stem cell (MSC) in tissue regeneration. However, the relevant research performed to date is only in its proof-of-concept stage. To use the microrobot in a clinical stage, biocompatibility and biodegradation materials should be considered in the microrobot, and its efficacy needs to be verified using an in vivo model. In this study, we propose a human adipose-derived MSC-based medical microrobot system for knee cartilage regeneration and present an in vivo trial to verify the efficacy of the microrobot using the cartilage defect model. The microrobot system consists of a microrobot body capable of supporting MSCs, an electromagnetic actuation system for three-dimensional targeting of the microrobot, and a magnet for fixation of the microrobot to the damaged cartilage. Each component was designed and fabricated considering the accessibility of the patient and medical staff, as well as clinical safety. The efficacy of the microrobot system was then assessed in the cartilage defect model of rabbit knee with the aim to obtain clinical trial approval.

The use of micro-needle arrays to deliver cells for cellular therapies.

Cell therapy is used to treat various diseases and to repair injuries. Cell delivery is a crucial process that delivers cells to target sites. Cells must be precisely delivered to a target site and the cells that are delivered must be localized to the target site to repair damaged tissue. For stem cell therapy, the most convenient method of cell delivery involves directly injecting cells into damaged tissue. Other strategies use carriers to transplant stem cells into damaged tissue. These are termed, stem cell delivery systems (SCDSs). Micro-needle arrays are minimally invasive transdermal delivery systems. The devices can pass through the stratum corneum barrier and deliver macromolecules into the skin. They can also access the microcirculation system in the skin. This study fabricates PMMA micro-needle using a two-stage micro-molding method. Cells are seeded on the micro-needle arrays and then transferred into the target tissue. Collagen hydrogel is used as a model biomimetic tissue. Cells are efficiently delivered to regions of interest, collagen hydrogel, by using this system. The delivery rate is about 83.2%. This demonstrates that micro-needle arrays allow very efficient delivery of cells.

Determination of antibiotic impurities in good manufacturing practices-grade cell therapy medicinal products.

Backrounds: According to the regulations of the health autorities, cell-based therapy products must be manufactured in good manufacturing production (GMP) facilities, fulfilling the required GMP standards. Products developed under the high quality control (QC) necessarity need to be approved for some QC tests. One of the main residual test is antibiotic test and this test should be validated. The aim of this study is to validate and determine the methods of detection of the antibiotic residue in the final product.Methods: Liquid Chromatography Tandem-Mass Spectrometry (LC-MS/MS) methods were used for the main steps of the production procedure, as well as the final products. Pharmaceutical Grade penicillin G and streptomycin sulfate were used as positive controls.Results: The results suggest that penicillin is broken down during cell culture and streptomycin is eliminated at the first washing step of the final product manufacture. It is shown in this study that LC-MS/MS method is one of the convenient method to test residual anibiotics and can be used to detect the antibiotic residues in cellular therapy products.Discussion: Since the antibiotic residues are eliminated in the final product and also it could be suggested that the methodology we followed is sufficiently safe and final product is pure.

A New Era for Cyborg Science Is Emerging: The Promise of Cyborganic Beings.

Living flesh, hacked beyond known biological borders, and sophisticated machineries, made by humans, are currently being united to address some of the impending challenges in medicine. Imagine biological systems made from smart biomaterials with the capacity to operate like smart machines to regulate insulin production in diabetic patients, or cardiac patches that can monitor and release important biological factors, on demand, to optimize the mending of broken hearts. It sounds like something from the realm of science fiction; however, this big gap between the real world and the world of fantasy and fiction is slowly being bridged. This piece sheds a much-needed light on this emerging area, as this futuristic concept is gaining momentum, at a speed, that soon will ignite a paradigm shift in disease management and the healthcare sector as an entirety.

Streamlined production of genetically modified T cells with activation, transduction and expansion in closed-system G-Rex bioreactors.

BACKGROUND: Gas Permeable Rapid Expansion (G-Rex) bioreactors have been shown to efficiently expand immune cells intended for therapeutic use, but do not address the complexity of the viral transduction step required for many engineered T-cell products. Here we demonstrate a novel method for transduction of activated T cells with Vectofusin-1 reagent. Transduction is accomplished in suspension, in G-Rex bioreactors. The simplified transduction step is integrated into a streamlined process that uses a single bioreactor with limited operator intervention. METHODS: Peripheral blood mononuclear cells (PBMCs) from healthy donors were thawed, washed and activated with soluble anti-CD3 and anti-CD28 antibodies either in cell culture bags or in G-Rex bioreactors. Cells were cultured in TexMACS GMP medium with interleukin (IL)-7 and IL-15 and transduced with RetroNectin in bags or Vectorfusin-1 in the G-Rex. Total viable cell number, fold expansion, viability, transduction efficiency, phenotype and function were compared between the two processes. RESULTS: The simplified process uses a single vessel from activation through harvest and achieves 56% transduction with 29-fold expansion in 11 days. The cells generated in the simplified process do not differ from cells produced in the conventional bag-based process functionally or phenotypically. DISCUSSION: This study demonstrates that T cells can be transduced in suspension. Further, the conventional method of generating engineered T cells in bags for clinical use can be streamlined to a much simpler, less-expensive process without compromising the quality or function of the cell product.

An Atmosphere-Breathing Refillable Biphasic Device for Cell Replacement Therapy.

Cell replacement therapy is emerging as a promising treatment platform for many endocrine disorders and hormone deficiency diseases. The survival of cells within delivery devices is, however, often limited due to low oxygen levels in common transplantation sites. Additionally, replacing implanted devices at the end of the graft lifetime is often unfeasible and, where possible, generally requires invasive surgical procedures. Here, the design and testing of a modular transcutaneous biphasic (BP) cell delivery device that provides enhanced and unlimited oxygen supply by direct contact with the atmosphere is presented. Critically, the cell delivery unit is demountable from the fixed components of the device, allowing for surgery-free refilling of the therapeutic cells. Mass transfer studies show significantly improved performance of the BP device in comparison to subcutaneous controls. The device is also tested for islet encapsulation in an immunocompetent diabetes rodent model. Robust cell survival and diabetes correction is observed following a rat-to-mouse xenograft. Lastly, nonsurgical cell refilling is demonstrated in dogs. These studies show the feasibility of this novel device for cell replacement therapies.

Smart polymers for cell therapy and precision medicine.

Soft materials have been developed very rapidly in the biomedical field over the past 10 years because of advances in medical devices, cell therapy, and 3D printing for precision medicine. Smart polymers are one category of soft materials that respond to environmental changes. One typical example is the thermally-responsive polymers, which are widely used as cell carriers and in 3D printing. Self-healing polymers are one type of smart polymers that have the capacity to recover the structure after repeated damages and are often injectable through needles. Shape memory polymers are another type with the ability to memorize their original shape. These smart polymers can be used as cell/drug/protein carriers. Their injectability and shape memory performance allow them to be applied in bioprinting, minimally invasive surgery, and precision medicine. This review will describe the general materials design, characterization, as well as the current progresses and challenges of these smart polymers.

A bioresorbable biomaterial carrier and passive stabilization device to improve heart function post-myocardial infarction.

The limited regenerative capacity of the heart after a myocardial infarct results in remodeling processes that can progress to congestive heart failure (CHF). Several strategies including mechanical stabilization of the weakened myocardium and regenerative approaches (specifically stem cell technologies) have evolved which aim to prevent CHF. However, their final performance remains limited motivating the need for an advanced strategy with enhanced efficacy and reduced deleterious effects. An epicardial carrier device enabling a targeted application of a biomaterial-based therapy to the infarcted ventricle wall could potentially overcome the therapy and application related issues. Such a device could play a synergistic role in heart regeneration, including the provision of mechanical support to the remodeling heart wall, as well as providing a suitable environment for in situ stem cell delivery potentially promoting heart regeneration. In this study, we have developed a novel, single-stage concept to support the weakened myocardial region post-MI by applying an elastic, biodegradable patch (SPREADS) via a minimal-invasive, closed chest intervention to the epicardial heart surface. We show a significant increase in %LVEF 14 days post-treatment when GS (clinical gold standard treatment) was compared to GS + SPREADS + Gel with and without cells (p ≤ 0.001). Furthermore, we did not find a significant difference in infarct quality or blood vessel density between any of the groups which suggests that neither infarct quality nor vascularization is the mechanism of action of SPREADS. The SPREADS device could potentially be used to deliver a range of new or previously developed biomaterial hydrogels, a remarkable potential to overcome the translational hurdles associated with hydrogel delivery to the heart.

Bringing Stem Cell-Based Therapies for Type 1 Diabetes to the Clinic: Early Insights from Bioprocess Economics and Cost-Effectiveness Analysis.

Differentiation of pluripotent stem cells (PSCs) into ß cells could provide insulin independence for type 1 diabetes (T1D) patients. This approach would reduce the clinical complications that most patients managed on intensive insulin therapy (IIT) face. However, bottlenecks of PSC manufacturing and limited engraftment of encapsulated cells hinder the long-term effectiveness of these therapies. A bioprocess decision-support tool is combined with a disease state-transition model to evaluate the cost-effectiveness of the stem cell-based therapy against IIT. Clinical effectiveness is assessed in quality-adjusted life years (QALYs). Manufacturing costs per patient reduce from $430 000 to $160 000 with optimization of batch size and annual demand. For 96% of the patients, cell therapy improves the quality of life compared to IIT. Cost savings are achieved for 2% of the population through prevention of renal disease. The therapy is cost-effective for 3.4% of patients when a willingness to pay (WTP) of up to $150 000 per QALY is considered. A 75% cost reduction in the cell therapy price increases cost-effectiveness likelihood to 51% at $100 000 per QALY. This study highlights the need for scalable manufacturing platforms for stem cell therapies, as well as to prioritizing access to the therapy to patients with an increased likelihood of costly complications.

Modeling biological and economic uncertainty on cell therapy manufacturing: the choice of culture media supplementation.

AIM: To evaluate the cost-effectiveness of autologous cell therapy manufacturing in xeno-free conditions. MATERIALS & METHODS: Published data on the isolation and expansion of mesenchymal stem/stromal cells introduced donor, multipassage and culture media variability on cell yields and process times on adherent culture flasks to drive cost simulation of a scale-out campaign of 1000 doses of 75 million cells each in a 400 square meter Good Manufacturing Practices facility. RESULTS & CONCLUSION: Passage numbers in the expansion step are strongly associated with isolation cell yield and drive cost increases per donor of $1970 and 2802 for fetal bovine serum and human platelet lysate. Human platelet lysate decreases passage numbers and process costs in 94.5 and 97% of donors through lower facility and labor costs. Cost savings are maintained with full equipment depreciation and higher numbers of cells per dose, highlighting the number of cells per passage step as the key cost driver.

An immune cell spray (ICS) formulation allows for the delivery of functional monocyte/macrophages.

Macrophages are key cells of the innate immune system and act as tissue resident macrophages (TRMs) in the homeostasis of various tissues. Given their unique functions and therapeutic use as well as the feasibility to derive macrophages in vitro from hematopoietic stem cell (HSC) sources, we propose an "easy-to-use" immune cell spray (ICS) formulation to effectively deliver HSC-derived macrophages. To achieve this aim, we used classical pump spray devices to spray either the human myeloid cell line U937 or primary murine HSC-derived macrophages. For both cell types used, one puff could deliver cells with maintained morphology and functionality. Of note, cells tolerated the spraying process very well with a recovery of more than 90%. In addition, we used osmotic preconditioning to reduce the overall cell size of macrophages. While a 800 mosm hyperosmolar sucrose solution was able to reduce the cell size by 27%, we identified 600 mosm to be effective to reduce the cell size by 15% while maintaining macrophage morphology and functionality. Using an isolated perfused rat lung preparation, the combinatorial use of the ICS with preconditioned and genetically labeled U937 cells allowed the intra-pulmonary delivery of cells, thus paving the way for a new cell delivery platform.

Versatile graphene biosensors for enhancing human cell therapy.

Technological advances in engineering and cell biology stimulate novel approaches for medical treatment, in particular cell-based therapy. The first cell-based gene therapy against cancer was recently approved by the US Food and Drug Administration. Progress in cancer diagnosis includes a blood test detecting five cancer types. Numerous stem cell phase I/II clinical trials showing safety and efficacy will soon pursue qualifying criteria for advanced therapy medicinal products (ATMP), aspiring to join the first stem-cell therapy approved by the European Medicines Agency. Cell based therapy requires extensive preclinical characterisation of biomarkers indicating mechanisms of action crucial to the desired therapeutic effect. Quantitative analyses monitoring critical functions for the manufacture of optimal cell and tissue-based clinical products include successful potency assays for implementation. The challenge to achieve high quality measurement is increasingly met by progress in biosensor design. We adopt a cell therapy perspective to highlight recent examples of graphene-enhanced biointerfaces for measurement of biomarkers relevant to cancer treatment, diagnosis and tissue regeneration. Graphene based biosensor design problems can thwart their use for health care transformative point of care testing and real-time applications. We discuss concerns to be addressed and emerging solutions for establishing clinical grade biosensors to accelerate human cell therapy.

An oxygen plasma treated poly(dimethylsiloxane) bioscaffold coated with polydopamine for stem cell therapy.

In this study, 3D macroporous bioscaffolds were developed from poly(dimethylsiloxane) (PDMS) which is inert, biocompatible, non-biodegradable, retrievable and easily manufactured at low cost. PDMS bioscaffolds were synthesized using a solvent casting and particulate leaching (SCPL) technique and exhibited a macroporous interconnected architecture with 86 ± 3% porosity and 300 ± 100 µm pore size. As PDMS intrinsically has a hydrophobic surface, mainly due to the existence of methyl groups, its surface was modified by oxygen plasma treatment which, in turn, enabled us to apply a novel polydopamine coating onto the surface of the bioscaffold. The addition of a polydopamine coating to bioscaffolds was confirmed using composition analysis. Characterization of oxygen plasma treated-PDMS bioscaffolds coated with polydopamine (polydopamine coated-PDMS bioscaffolds) showed the presence of hydroxyl and secondary amines on their surface which resulted in a significant decrease in water contact angle when compared to uncoated-PDMS bioscaffolds (35 ± 3%, P < 0.05). Seeding adipose tissue-derived mesenchymal stem cells (AD-MSCs) into polydopamine coated-PDMS bioscaffolds resulted in cells demonstrating a 70 ± 6% increase in viability and 40 ± 5% increase in proliferation when compared to AD-MSCs seeded into uncoated-PDMS bioscaffolds (P < 0.05). In summary, this two-step method of oxygen plasma treatment followed by polydopamine coating improves the biocompatibility of PDMS bioscaffolds and only requires the use of simple reagents and mild reaction conditions. Hence, our novel polydopamine coated-PDMS bioscaffolds can represent an efficient and low-cost bioscaffold platform to support MSC therapies.

Manufacturing human mesenchymal stem cells at clinical scale: process and regulatory challenges.

Human mesenchymal stem cell (hMSC)-based therapies are of increasing interest in the field of regenerative medicine. As economic considerations have shown, allogeneic therapy seems to be the most cost-effective method. Standardized procedures based on instrumented single-use bioreactors have been shown to provide billion of cells with consistent product quality and to be superior to traditional expansions in planar cultivation systems. Furthermore, under consideration of the complex nature and requirements of allogeneic hMSC-therapeutics, a new equipment for downstream processing (DSP) was successfully evaluated. This mini-review summarizes both the current state of the hMSC production process and the challenges which have to be taken into account when efficiently producing hMSCs for the clinical scale. Special emphasis is placed on the upstream processing (USP) and DSP operations which cover expansion, harvesting, detachment, separation, washing and concentration steps, and the regulatory demands.

TGF-ß1/CD105 signaling controls vascular network formation within growth factor sequestering hyaluronic acid hydrogels.

Cell-based strategies for the treatment of ischemic diseases are at the forefront of tissue engineering and regenerative medicine. Cell therapies purportedly can play a key role in the neovascularization of ischemic tissue; however, low survival and poor cell engraftment with the host vasculature following implantation limits their potential to treat ischemic diseases. To overcome these limitations, we previously developed a growth factor sequestering hyaluronic acid (HyA)-based hydrogel that enhanced transplanted mouse cardiosphere-derived cell survival and formation of vasculature that anastomosed with host vessels. In this work, we examined the mechanism by which HyA hydrogels presenting transforming growth factor beta-1 (TGF-ß1) promoted proliferation of more clinically relevant human cardiosphere-derived cells (hCDC), and their formation of vascular-like networks in vitro. We observed hCDC proliferation and enhanced formation of vascular-like networks occurred in the presence of TGF-ß1. Furthermore, production of nitric oxide (NO), VEGF, and a host of angiogenic factors were increased in the presence of TGF-ß1. This response was dependent on the co-activity of CD105 (Endoglin) with the TGF-ßR2 receptor, demonstrating its role in the process of angiogenic differentiation and vascular organization of hCDC. These results demonstrated that hCDC form vascular-like networks in vitro, and that the induction of vascular networks by hCDC within growth factor sequestering HyA hydrogels was mediated by TGF-ß1/CD105 signaling.

Survival of Mesenchymal Stem Cells in Different Methods of Nebulization.

We compared survival of bone marrow mesenchymal stem cells after compressor, ultrasound, and mesh nebulization of the cell suspension over 10 min. Viability of stromal cells was best preserved after compressor nebulization (72%). Cell survival after ultrasonic nebulization was significantly lower (20%). After mesh nebulization, no live cells were found. Thus, compressor nebulization is the most preferable method of the production of cell aerosol for their delivery to the lower respiratory tract.

Bioprocessing automation in cell therapy manufacturing: Outcomes of special interest group automation workshop.

Phacilitate held a Special Interest Group workshop event in Edinburgh, UK, in May 2017. The event brought together leading stakeholders in the cell therapy bioprocessing field to identify present and future challenges and propose potential solutions to automation in cell therapy bioprocessing. Here, we review and summarize discussions from the event. Deep biological understanding of a product, its mechanism of action and indication pathogenesis underpin many factors relating to bioprocessing and automation. To fully exploit the opportunities of bioprocess automation, therapeutics developers must closely consider whether an automation strategy is applicable, how to design an 'automatable' bioprocess and how to implement process modifications with minimal disruption. Major decisions around bioprocess automation strategy should involve all relevant stakeholders; communication between technical and business strategy decision-makers is of particular importance. Developers should leverage automation to implement in-process testing, in turn applicable to process optimization, quality assurance (QA)/ quality control (QC), batch failure control, adaptive manufacturing and regulatory demands, but a lack of precedent and technical opportunities can complicate such efforts. Sparse standardization across product characterization, hardware components and software platforms is perceived to complicate efforts to implement automation. The use of advanced algorithmic approaches such as machine learning may have application to bioprocess and supply chain optimization. Automation can substantially de-risk the wider supply chain, including tracking and traceability, cryopreservation and thawing and logistics. The regulatory implications of automation are currently unclear because few hardware options exist and novel solutions require case-by-case validation, but automation can present attractive regulatory incentives.

Minimally invasive and targeted therapeutic cell delivery to the skin using microneedle devices.

BACKGROUND: Translation of cell therapies to the clinic is accompanied by numerous challenges, including controlled and targeted delivery of the cells to their site of action, without compromising cell viability and functionality. OBJECTIVES: To explore the use of hollow microneedle devices (to date only used for the delivery of drugs and vaccines into the skin and for the extraction of biological fluids) to deliver cells into skin in a minimally invasive, user-friendly and targeted fashion. METHODS: Melanocyte, keratinocyte and mixed epidermal cell suspensions were passed through various types of microneedles and subsequently delivered into the skin. RESULTS: Cell viability and functionality are maintained after injection through hollow microneedles with a bore size ≥ 75 µm. Healthy cells are delivered into the skin at clinically relevant depths. CONCLUSIONS: Hollow microneedles provide an innovative and minimally invasive method for delivering functional cells into the skin. Microneedle cell delivery represents a potential new treatment option for cell therapy approaches including skin repigmentation, wound repair, scar and burn remodelling, immune therapies and cancer vaccines.