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Cellular therapies, including hematopoietic stem cells (HSCs), mesenchymal stem cells (MSCs), and non-stem cell-based therapies like CAR-T cells, have gained prominence in therapeutic applications due to their regenerative and immunomodulatory properties. Despite the benefits observed in patients, these therapies are often accompanied by certain disadvantages that limit their clinical use. In contrast, cell-free therapies, such as acellular scaffolds, small molecules, RNA molecules, and MSC-derived factors such as extracellular vesicles (EVs), present a promising alternative for treating a wide range of diseases. These therapies offer several advantages, including minimal immunogenicity, defined composition, cost-effectiveness, and ease of storage for extended periods. This chapter will provide a comprehensive overview of the various cell-free therapeutic approaches in the context of different diseases and explore how these methods can revolutionize treatment, offering a significant advancement over traditional cellular therapies.
*Address all correspondence to: deepakjha736@gmail.com
1. Introduction to cellular therapies
Cell therapy, or cellular therapy, refers to the introduction of autologous or allogenic viable cells into a patient in order to treat a medical condition [1].
There are two major types of cell-based therapies, namely: (A) stem cell and (B) non-stem cell-based therapies. In the former case, adult stem cells such as hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs) are the most widely used in clinical settings to treat a variety of conditions such as disorders of the blood and immune system, cancer, and degenerative diseases, to name a few [2]. Currently, the only stem cell-based therapy approved by the “U.S.” Food and Drug Administration (FDA) is hematopoietic (or blood) stem cell and progenitor cell (derived from cord blood) transplantation for treating certain cancers and blood disorders [3, 4]. Clinical trials for the treatment of other conditions employing MSCs are underway [5, 6]. Despite being a promising candidate to treat a variety of medical conditions, stem cell-based therapies are also associated with certain limitations, which are classified into four categories (Figure 1).
Intrinsic factor—Rejection of the transplanted cells based on the origin of cells (autologous or allogenic).
Cell characteristics—(a) Tumorigenic potential, due to the ability of the stem cells to self-renew, and (b) ability of the stem cells to change into inappropriate cell types.
Extrinsic factors—(a) Infections, arising due to lack of donor history, (b) potential for the contamination of the product arising due to the cell handling procedures, raw materials (growth media components, chemicals, etc.), and (c) low cell viability during administration.
Clinical characteristics—(a) Undesired immune response (for example, Graft versus Host Disease (GVHD), arising due to allogenic stem cell transplantation), (b) injection site reaction, (c) administration route, if inappropriate, can result in engraftment at an unwanted location, (d) lack of therapeutic efficacy arising due to several factors (for example, use of immunosuppressives), (e) Lack of knowledge regarding optimal dosage and timing of injections, (f) Less observable clinical benefits due to low engraftment of cells [7, 8, 9, 10].
Figure 1.
Challenges associated with stem cell-based therapies. (Source: BioRender).
Non-stem cell-based therapies are generally somatic cells (T cells or genetically modified T cells, fibroblasts, and keratinocytes, to name a few) that are isolated from humans, expanded in vitro and administered back to the patients in order to effectuate the medical condition [11, 12, 13, 14]. Currently, there are only 13 non-stem cell-based products available that are approved by the FDA, of which 60% of the products employ Chimeric Antigen Receptor T cells (CAR-T cells) to treat B cell lymphoma [15, 16], and the remaining 40% are scaffold-based cellular products (Table 1) [13].
CD19-directed genetically modified autologous T cells
ABECMA®
B cell maturation antigen (BCMA)-directed genetically modified autologous T cells
PROVENGE®
Autologous CD54+ cells activated with PAP-GM-CSF
GINTUIT®
Allogenic cultured keratinocytes and fibroblasts in bovine collagen
LAVIV®
Autologous cultured fibroblast cells
MACI®
Autologous cultured chondrocytes on porcine collagen membrane
Table 1.
FDA approved non-stem cell-based cellular products [11].
Scaffold-based cellular products are by far considered to be safe and an effective alternative to current cell-based treatments (which mainly focus on delivering the cells either systemically or intradermally) in favoring enhanced cell viability, retention, and functionality at the wound site (Table 1) [17].
In the past decade there has been increasing evidence that it is the bioactive factors secreted from the transplanted cells that matter more during the treatment than the cell transplantation alone [18, 19]. Cell-free therapies have been quite extensively studied for their potential application in cancer, regenerative medicine, and inflammatory diseases, to name a few [18, 20]. This kind of therapy is now considered the next-generation of therapeutic technology. Cell-free therapies overcome the limitations and risks posed by the cell-based therapies in that: [21, 22, 23, 24]
It lowers the risk associated with pulmonary embolism after intravenous (IV) administration.
It avoids the risk of tumorigenic potential, as cell-free products cannot self-replicate.
It is non-immunogenic due to the limited number of antigenic components.
It avoids exogenous infections, which are more prominent in cell-based therapies.
It can be stored for relatively long periods of time without any toxic cryopreservant such as DMSO.
It is able to infiltrate the target organ more effectively (targeted drug delivery).
Therapeutic efficacy can be equivalent to or more effective than cell-based therapy.
Broadly, there are two major types of cell-free strategies: (a) acellular material-based scaffolds [25] and (b) endogenous cell targeting through delivery of bioactive factors [26]. These two fields are the subjects of extensive research nowadays (Figure 2).
Figure 2.
Cell-free products and their advantages (Source: BioRender).
3.1 Acellular material-based scaffold
The field of using acellular material-based scaffolds has significantly expanded over the past few decades owing to its potential application ranging from promoting tissue formation through mechanical support (traditional approach) to organizing and vascularizing tissues (scaffold containing bioactive molecules) (current approach) and to modulating endogenous cellular processes such as inflammation, direct differentiation, and recruiting cells for repair (emerging approach). Additionally, they also provide structural support to the exogenously applied cells to attach, grow, and differentiate in vivo [25].
Acellular biomaterials are aimed at enhancing the repair process of native cell populations after injury or with disease, for example, repairing cartilage tissue, treating patients after myocardial infarction, and bone tissue engineering [27, 28, 29]. Although cell-seeded scaffolds have given promising results in Tissue Engineering and Regenerative Medicine (TERM), there are quite a few challenges in order to maximize the clinical benefits [30]. Some of the major challenges include:
Cell type, seeding, and distribution on a scaffold
Design of the scaffold (composition, i.e., natural or synthetic polymers, topography, and architecture), which influences cell attachment and its behavior.
In contrast, acellular-based scaffolds have extremely low immunogenicity due to the absence of the employment of any allogenic cells on a scaffold, i.e., these types of acellular scaffolds are loaded with all the necessary cues and signals in order to drive the repair process efficiently [31]. The current limitations of acellular scaffolds in TERM include design parameters of scaffolds for therapeutic efficacy, biocompatibility, their degradation rate, and bioactivity [32].
3.2 Endogenous cell targeting
Targeting several endogenous cell moieties has also been explored to treat a variety of disease conditions, including degenerative disorders, without the necessity of cell therapy [33]. There are different ways by which an endogenous cell moiety can be targeted based on the disease condition.
These include:
(3.2.1) Small molecules.
(3.2.2) RNA therapeutic strategies.
(3.2.3) Direct reprogramming.
(3.2.4) Growth factors and proteins.
3.2.1 Small molecules
Over the past few decades, there has been tremendous focus on utilizing the small molecule (targeting a specific cell moiety) to enhance the repair process after injury or with disease [34]. For example, prostaglandin E2 is known to have a regenerative role in ischemic myocardium post myocardial infarction (MI). It activates and mobilizes the endogenous cardiac stem cell (Sca-1+ cells) population to the infarct border zone and regulates their differentiation into cardiomyocytes [35]. Single or combinations of small molecules can also be utilized to solve a disease condition [36]. The employment of small molecules has certain advantages as therapeutics in some aspects, which are:
Because of their small size, these can be effectively used to target extracellular proteins or intracellular receptors due to their ability to pass through the cell membranes easily [37].
Can be easily manufactured by chemical synthesis, and therefore these are often inexpensive [38].
Easy to store and transport.
Patient compliance is majorly due to higher oral bioavailability.
Extremely stable.
Variety of formulations available.
Driver of innovation in research and development [39].
However, small molecule drugs still face some challenges, majorly of which are drug resistance and low response rate.
3.2.2 RNA therapeutic strategies
RNA therapies generally work by manipulating gene expression or producing therapeutic proteins. There are a variety of RNA therapeutic strategies that can be utilized for pathologies with distinct genetic targets, including infectious diseases, cancer, or immune diseases [40]. It also finds its use in regenerative medicine as well [41].
3.2.2.1 Modified messenger RNA (ModRNA)
ModRNA is a type of synthetic messenger RNA (mRNA) containing non-standard residues. This is an improved version of the current mRNA therapy, which faces certain challenges that limit the success rate of this therapy in clinical settings [42]. These include:
Short half-life of mRNA, due to the presence of ubiquitous enzymes (ribonucleases (RNAses)) that degrade the unprotected mRNA.
Inflammatory nature of exogenous RNA, i.e., in vitro transcribed foreign RNA, can be recognized by certain innate immune system receptors, which ultimately decreases the target protein synthesis in the cell. This recognition leads to the release of various inflammatory cytokines, which, when it exceeds the limit, results in programmed cell death (PCD) [43].
Contrary to that, the inclusion of the modified nucleosides, such as uridine, pseudouridine, or 5-methylcytosine, in this mRNA can prevent the inflammatory reaction by escaping the recognition from the innate immune system while still translating this ModRNA into the target protein effectively in the cell [44]. ModRNA also finds its application in regenerative medicine, for example, repairing the damaged heart muscle tissue post myocardial infarction [45].
3.2.2.2 MicroRNA targeting
MicroRNAs, or miRs, are a kind of small, endogenous, non-coding RNAs of about 18–22 nucleotides in length that regulate coding gene expression and ultimately mediate changes in protein synthesis. miRNAs target mRNAs by binding to them (complementary binding) in order to exert their function. A single miRNA can have multiple mRNA targets and thus have the ability to influence many related genes, which results in multifaceted effects on cell phenotype [46]. There are two major strategies by which miR activity can be modulated based on the disease condition: (1) Overexpression, and (2) down-regulation of miRNAs. Mounting evidence has shown that modulating miRNAs provided promising results in bone regeneration, wound healing, skeletal muscle and cardiac regeneration, angiogenesis, and neurogenesis, majorly by promoting endogenous cell proliferation after injury [47, 48, 49, 50, 51].
Despite the great success of miRNA therapeutics in preclinical studies, very few clinical trials are being conducted currently. This is due to certain limitations associated with this kind of therapy, which include:
Efficient delivery of miRNA to the target site: (a) Chemical modifications of miRNA mimetics (to avoid its degradation from nucleases) prevent its recognition and loading onto argonaute and RNA-induced silencing complex (RISC). This will lead to a lesser therapeutic effect, as loading of miRNA into Argonaute and RISC is important to exert its function [52, 53]. (b) Liposome nanoparticles, one of the delivery strategies of miRNA, are relatively large, which makes it difficult to gain entry into the extracellular spaces, especially of solid tumors, to exert its effect [54].
Another challenge associated with miRNA therapeutics is the activation of the innate immune system, which leads to target cell death upon recognition of exogenous miRNA molecules [55].
Off-target effects, due to the ability of a single miRNA to target several mRNAs (otherwise known as “too many targets for miRNA effect”), this non-specific interaction could be either beneficial or trigger unknown consequences that can lessen its therapeutic effect [55]. Another off-target effect could be seen when the therapeutic miRNA is not tissue/cell-type specific. For example, obtaining a therapeutic benefit by modulating target gene expression in target cell types (say, abnormal cells) can be accompanied by modulating this same gene expression in non-target cell types (say, normal cells) if miRNA selection is not tissue/cell-type specific. Such an interaction can lead to toxicity [56].
3.2.3 Direct reprogramming
Direct epigenetic reprogramming is a very useful technique and a novel approach that converts an adult differentiated cell into another differentiated cell while bypassing the pluripotent stem cell fate (and the potential risk of tumor formation) [57]. This technique is mostly explored in the field of regenerative medicine, for example, generating functional cardiomyocytes from endogenous fibroblasts post myocardial infarction [58]. Recent research has indicated that there are five different approaches that can directly reprogram an adult differentiated cell into another phenotypic cell. These include: (a) transcription factors [59], (b) epigenetic regulators [60], (c) miRNAs, (d) small molecules and cell-penetrating peptides, and (e) pluripotency factors for indirect reprogramming [60].
Transfecting the host cell that allows for the expression of exogenous DNA or RNA drives the phenotypic changes via the activation of certain genes involved in reprogramming [61]. Cell transfection can be either transient or stable, with the former being advantageous in that it allows for the expression of the exogenous DNA or RNA without the need to be integrated into the host cell genome. This can be achieved using non-viral techniques such as liposome- and non-liposome-mediated transfection, dendrimer-based transfection and electroporation, and non-integrating viral systems such as adenovirus and adeno-associated virus (AAV) can also be used to transiently express the exogenous DNA or RNA. Unlike transient transfection, stable transfection allows the exogenous DNA or RNA to be integrated into the host cell genome permanently [62]. To achieve this, techniques such as microinjection and integrating viral systems (lentiviral and γ-retroviral vectors) can be utilized. Small molecules and cell-penetrating peptides have also been shown to regulate the process of transdifferentiating, either alone or with a combination of exogenous DNA or RNA, with the aim of increasing the efficiency of transdifferentiation [63, 64].
3.2.4 Growth factors and proteins
Growth factors (VEGF, SDF-1, IGF-1, FGF-1, HGF, PDGF, BMP family (BMP-2 and BMP-7), to name a few) are proteins that are known to influence cell growth, proliferation, motility, survival, adhesion, and differentiation and also regulate angiogenesis via the activation of specific cellular signal transduction pathways [65]. Growth factors exert their action by interacting with their receptor either in a diffusible manner (e.g., by endocrine, paracrine, autocrine, and intracrine pathways) or in a non-diffusible manner (e.g., by juxtacrine and matricrine pathways) or both [66].
These growth factors are majorly utilized for regenerative medicine applications. For instance, BMP-2 and BMP-7 (FDA approved) are involved in bone regeneration [67]. Similarly, growth factors such as PDGF-BB, IGF-1, and FGF-7 (KGF) (all FDA approved) are used to treat ankle fusion, hindfoot, primary IGF-1 deficiency, spinal fusion, and gastrointestinal injury [65]. Like other therapeutic agents, growth factor-based therapies are also associated with certain limitations, which hinders their translation into clinical applications. These include:
Protein stability and half-life.
Low yield of recombinant protein.
High costs and possibilities of side effects, due to multiple administrations in order to attain optimal concentration at the target site.
Off-target effects, i.e., if organ-specific growth factors for achieving therapeutic benefit are not selected, then the modulation expected by the desired growth factor can also involve other organs apart from the target organ, which can hence result in undesirable effect [68].
It was due to these limitations that led to advanced research that focuses on various approaches to enhance the therapeutic efficacy of exogenous growth factors. Briefly, these approaches include:
Introduction of stabilizing mutations in order to increase the half-life of the protein [70].
Identification of protein variants (e.g., by yeast and phage-based library display platforms) with improved stability and expression yield [71].
PEGylation to enhance the half-life of proteins, which can lead to reduced dosing frequency [72].
Use of library screening methods to isolate variants that have (a) increased receptor binding affinity and (b) decreased receptor internalization [73].
Engineering growth factors in such a way that allows them to also engage in alternative signaling pathways for the purpose of enhanced therapeutic effect.
Strategies involving protein engineering within the microenvironment in order to achieve maximum therapeutic efficacy for regenerative medicine applications [65].
Use of appropriate delivery systems that are safe and cost-effective, such as growth factor delivery through (a) decellularized extracellular matrix (ECM) and (b) exogenous engineered biomatrices, to name a few [74].
4. Mesenchymal stem cells (MSCs) as a source of therapeutic molecules for cell-free therapy
MSCs are the most commonly used cells for cell-based therapy, as they confer low immunogenicity due to relatively low levels of major histocompatibility complex class I molecules and lack expression of major histocompatibility complex class II and co-stimulatory molecules (e.g., CD40, CD80, and CD86), making them an ideal candidate for both allogenic and autologous therapeutics. MSCs are known for their marked potential in regenerative medicine due to their strong immunosuppressive and regenerative abilities [75]. The major therapeutic properties, like anti-inflammatory, anti-fibrotic, anti-oxidant, and angiogenic effects of MSCs, are due to their secretion of biologically active soluble products, including chemokines, cytokines, trophic factors, extracellular matrix (ECM) proteins, and extracellular vesicles, particularly exosomes, which can be used as “next generation” therapeutic and diagnostic [76]. Using cell-free products secreted by stem and progenitor cells allows one to significantly reduce the risks associated with a direct cell injection while maintaining efficacy similar to cellular therapy. Therefore, the application of “cell-free therapeutics” secreted by MSCs represents a promising approach in regenerative medicine [77].
Extracellular vesicles are cell-derived membrane particles ranging from 30 to 5000 nm in size, including exosomes, microvesicles, and apoptotic bodies [78]. They are released under physiological conditions, but also in response to various other stimuli. They play an important role in intercellular communication. Their release may also maintain cellular integrity by ridding the cell of damaging substances.
Exosomes are the smallest extracellular vesicles (30–100 nm) formed by the fusion of multivesicular bodies (MVBs) (containing intraluminal vesicles) with the plasma membrane. Because exosomes are formed by budding from early endosomes, they have a lipid bilayer membrane, which protects the resident genetic material (DNA, mRNA, miRNA, pre-miRNA, and other non-coding RNAs), lipids, and proteins during transportation to target cells. The most common exosome surface proteins are members of the tetraspanin family, a group of scaffold membrane proteins including CD63, CD81, and CD9 [79]. Exosomes can be used for disease diagnosis, drug delivery, and as therapeutic agents [80]. Exosomes engage in specific interactions with the recipient cells and influence various cellular processes in the recipient cells upon the uptake of exosomes. These include gene expression regulation, cell proliferation and survival, cell differentiation, immune modulation, and tissue repair and regeneration (Figure 3).
Figure 3.
Therapeutic effects of MSC-derived exosomes (Source: BioRender).
Like stem cells, exosomes exhibit many biological activities and have shown therapeutic potential in several disease contexts. For example, exosomes may protect against cisplatin-induced renal oxidative stress and renal cell apoptosis, enhance myocardial viability and prevent adverse remodeling after ischemic injury, promote angiogenesis in the setting of myocardial infarction, protect the intestines from enterocolitis, improve hypoxia-induced pulmonary hypertension, and promote functional recovery after stroke [81]. Additionally, exosomes exhibit immunomodulatory (largely anti-inflammatory) effects either by inhibiting activated effector T cells or by enriching the population of regulatory T cells [82]. The efficacy of exosomes is similar to cell-based therapy with fewer limitations, indicating that exosomes have potential as next-generation (i.e., cell-free) therapy.
Although exosomes showed promising results, there are certain concerns linked to MSC-Exosomes, which include (a) lack of standardization of molecular characteristics and its reproducibility in the context of a specific disease, (b) difficulty in obtaining high-purity exosomes of defined size, (c) bioavailability and toxicity of MSC-Exosomes after injection, and (d) unclear mode of action of MSC-Exosomes in the recipient cells due to various types of biologically active molecules present within the exosomes [83].
In conclusion, both stem/non-stem cell-based and cell-free therapies have made significant progress in the treatment of various diseases. Although cell-based therapies show promise, the challenges associated with these therapies hinder their use in clinical settings. On the other hand, cell-free therapies overcome these challenges, particularly those related to transplantation and immune rejection, thereby offering a unique alternative for treating a variety of diseases. Among cell-free products, MSC-derived extracellular vesicles, particularly exosomes, are considered especially promising for the treatment of diseases due to their high concentration of biologically active molecules. However, this field is still emerging, and challenges such as the standardization and purification of clinical-grade exosomes remain significant concerns. Therefore, further research is focused on addressing these challenges to make MSC-derived exosomes an ideal candidate for cell-free therapy in the treatment of various diseases.
We acknowledge the knowledge databases used in framing this chapter. Additionally, we would like to acknowledge BioRender for enabling us to create our illustrations.
1.Kim IS. A brief overview of cell therapy and its product. Journal of the Korean Association of Oral and Maxillofacial Surgeons. 2022;39(5):201-202. DOI: 10.5125/jkaoms.2013.39.5.201
2.Han Y, Yang J, Fang J, Zhou Y, Candi E, Wang J, et al. The secretion profile of mesenchymal stem cells and potential applications in treating human diseases. Signal Transduction and Targeted Therapy. 2021;7(1):1-19. DOI: 10.1038/s41392-022-00932-0
3.Gang EJ, Jeong JA, Hong SH, Hwang SH, Kim SW, Yang IH, et al. Skeletal myogenic differentiation of mesenchymal stem cells isolated from human umbilical cord blood. Stem Cells (Dayton, Ohio). 2002;22(4):617-624. DOI: 10.1634/stemcells.22-4-617
4.Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini FC, Krause DS, et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy. 2006;8(4):315-317. DOI: 10.1080/14653240600855905
5.Galderisi U, Peluso G, Bernardo D, Giovanni. Clinical trials based on mesenchymal stromal cells are exponentially increasing: Where are we in recent years? Stem Cell Reviews and Reports. 2022;18(1):23-36. DOI: 10.1007/s12015-021-10231-w
6.Zhou T, Yuan Z, Weng J, Pei D, Du X, He C, et al. Challenges and advances in clinical applications of mesenchymal stromal cells. Journal of Hematology & Oncology. 2021;14(1):24. DOI: 10.1186/s13045-021-01037-x
7.Herberts CA, Kwa MSG, Hermsen HPH. Risk factors in the development of stem cell therapy. Journal of Translational Medicine. 2011;9(1):29. DOI: 10.1186/1479-5876-9-29
8.Boltze J, Arnold A, Walczak P, Jolkkonen J, Cui L, Wagner D-C. The dark side of the force – Constraints and complications of cell therapies for stroke. Frontiers in Neurology. 2015;6:6. DOI: 10.3389/fneur.2015.00155
9.Kim HJ, Park J-S. Usage of human mesenchymal stem cells in cell-based therapy: Advantages and disadvantages. Development & Reproduction. 2017;21(1):1-10. DOI: 10.12717/DR.2017.21.1.001
10.Aly RM. Current state of stem cell-based therapies: An overview. Stem Cell Investigation. 2020;7:1-10. DOI: 10.21037/sci-2020-001
11.El-Kadiry AE-H, Rafei M, Shammaa R. Cell therapy: Types, regulation, and clinical benefits. Frontiers in Medicine. 2021;8:8. DOI: 10.3389/fmed.2021.756029
12.Mount NM, Ward SJ, Kefalas P, Hyllner J. Cell-based therapy technology classifications and translational challenges. Philosophical Transactions of the Royal Society B: Biological Sciences. 2015;370(1680):20150017. DOI: 10.1098/rstb.2015.0017
13.De Pieri A, Rochev Y, Zeugolis DI. Scaffold-free cell-based tissue engineering therapies: Advances, shortfalls and forecast. npj Regenerative Medicine. 2021;6(1):1-15. DOI: 10.1038/s41536-021-00133-3
14.Sánchez A, Schimmang T, García-Sancho J. Cell and tissue therapy in regenerative medicine. Stem Cell Transplantation. 2012;741:89-102. DOI: 10.1007/978-1-4614-2098-9_7
15.June CH, O’Connor RS, Kawalekar OU, Ghassemi S, Milone MC. CAR T cell immunotherapy for human cancer. Science. 2018;359(6382):1361-1365. DOI: 10.1126/science.aar6711
16.Vitale C, Strati P. CAR T-cell therapy for B-cell non-Hodgkin lymphoma and chronic lymphocytic leukemia: Clinical trials and real-world experiences. Frontiers in Oncology. 2020;10:1-17. DOI: 10.3389/fonc.2020.00849
17.Sivaraj D, Chen K, Chattopadhyay A, Henn D, Wu W, Noishiki C, et al. Hydrogel scaffolds to deliver cell therapies for wound healing. Frontiers in Bioengineering and Biotechnology. 2021;9:1-17. DOI: 10.3389/fbioe.2021.660145
18.Foo JB, Looi QH, Chong PP, Hassan NH, Yeo GEC, Ng CY, et al. Comparing the therapeutic potential of stem cells and their secretory products in regenerative medicine. Stem Cells International. 2021;2021:1-30. DOI: 10.1155/2021/2616807
19.Ma Z-J, Yang J-J, Lu Y-B, Liu Z-Y, Wang X-X. Mesenchymal stem cell-derived exosomes: Toward cell-free therapeutic strategies in regenerative medicine. World Journal of Stem Cells. 2020;12(8):814-840. DOI: 10.4252/wjsc.v12.i8.814
20.Weng Z, Zhang B, Wu C, Yu F, Han B, Li B, et al. Therapeutic roles of mesenchymal stem cell-derived extracellular vesicles in cancer. Journal of Hematology & Oncology. 2021;14(1):136. DOI: 10.1186/s13045-021-01141-y
21.Maron-Gutierrez T, Rocco PRM. Cell-free therapies: Novel approaches for COVID-19. Frontiers in Immunology. 2020;11:1-6. DOI: 10.3389/fimmu.2020.583017
22.Hu C, Zhao L, Zhang L, Bao Q, Li L. Mesenchymal stem cell-based cell-free strategies: Safe and effective treatments for liver injury. Stem Cell Research & Therapy. 2020;11(1):377. DOI: 10.1186/s13287-020-01895-1
23.Tan MI, Alfarafisa NM, Septiani P, Barlian A, Firmansyah M, Faizal A, et al. Potential cell-based and cell-free therapy for patients with COVID-19. Cells. 2022;11(15):2319. DOI: 10.3390/cells11152319
24.Chou Y, Alfarafisa NM, Ikezawa M, Khairani AF. Progress in the development of stem cell-derived cell-free therapies for skin aging. Clinical, Cosmetic and Investigational Dermatology. 2023;16:3383-3406. DOI: 10.2147/CCID.S434439
25.Burdick JA, Mauck RL, Gorman JH, Gorman RC. Acellular biomaterials: An evolving alternative to cell-based therapies. Science Translational Medicine. 2013;5(176):1-9. DOI: 10.1126/scitranslmed.3003997
26.Vo TN, Kasper FK, Mikos AG. Strategies for controlled delivery of growth factors and cells for bone regeneration. Advanced Drug Delivery Reviews. Targeted Delivery of Therapeutics to Bone and Connective Tissues. 2012;64(12):1292-1309. DOI: 10.1016/j.addr.2012.01.016
27.Yang Z, Li H, Yuan Z, Fu L, Jiang S, Gao C, et al. Endogenous cell recruitment strategy for articular cartilage regeneration. Acta Biomaterialia. 2020;114:31-52. DOI: 10.1016/j.actbio.2020.07.008
28.Wang B, Wang G, To, Filip, Butler JR, Claude A, McLaughlin RM, et al. Myocardial scaffold-based cardiac tissue engineering: Application of coordinated mechanical and electrical stimulations. Langmuir. 2013;29(35):11109-11117. DOI: 10.1021/la401702w
29.Amirazad H, Dadashpour M, Zarghami N. Application of decellularized bone matrix as a bioscaffold in bone tissue engineering. Journal of Biological Engineering. 2022;16(1). DOI: 10.1186/s13036-021-00282-5. ISSN 1754-1611
30.Ikada Y. Challenges in tissue engineering. Journal of the Royal Society Interface. 2006;3(10):589-601. DOI: 10.1098/rsif.2006.0124
31.Chan BP, Leong KW. Scaffolding in tissue engineering: General approaches and tissue-specific considerations. European Spine Journal. 2008;17(4):467-479. DOI: 10.1007/s00586-008-0745-3
32.Molina E, Maria I, Malollari KG, Komvopoulos K. Design challenges in polymeric scaffolds for tissue engineering. Frontiers in Bioengineering and Biotechnology. 2021;9:9. DOI: 10.3389/fbioe.2021.617141
33.Parayath NN, Amiji MM. Therapeutic targeting strategies using endogenous cells and proteins. Journal of Controlled Release: Official Journal of the Controlled Release Society. 2017;258:81-94. DOI: 10.1016/j.jconrel.2017.05.004
34.Maniam S, Maniam S. Small molecules targeting programmed cell death in breast cancer cells. International Journal of Molecular Sciences. 2021;22(18):9722. DOI: 10.3390/ijms22189722
35.Cheng H, Huang H, Guo Z, Chang Y, Li Z. Role of prostaglandin E2 in tissue repair and regeneration. Theranostics. 2021;11(18):8836-8854. DOI: 10.7150/thno.63396
36.Wishart DS. Chapter 3: Small molecules and disease. PLoS Computational Biology. 2012;8(12):e1002805. DOI: 10.1371/journal.pcbi.1002805
37.Yang NJ, Hinner MJ. Getting across the cell membrane: An overview for small molecules, peptides, and proteins. In: Methods in Molecular Biology. Vol. 1266. Clifton, N.J: Springer Nature; 2015. pp. 29-53. DOI: 10.1007/978-1-4939-2272-7_3
38.Fu H, Yao J, Zhang M, Xue L, Zhou Q, Li S, et al. Low-cost synthesis of small molecule acceptors makes polymer solar cells commercially viable. Nature Communications. 2022;13(1):3687. DOI: 10.1038/s41467-022-31389-y
39.Gurevich EV, Gurevich VV. Therapeutic potential of small molecules and engineered proteins. Handbook of Experimental Pharmacology. 2014;219:1-12. DOI: 10.1007/978-3-642-41199-1_1
40.Kim Y-K. RNA therapy: Current status and future potential. Chonnam Medical Journal. 2020;56(2):87. DOI: 10.4068/cmj.2020.56.2.87
41.Chabanovska O, Galow A-M, David R, Lemcke H. mRNA—A game changer in regenerative medicine, cell-based therapy and reprogramming strategies. Advanced Drug Delivery Reviews. 2021;179:114002. DOI: 10.1016/j.addr.2021.114002
42.Granados-Riveron JT, Aquino-Jarquin G. Engineering of the current nucleoside-modified mRNA-LNP vaccines against SARS-CoV-2. Biomedicine & Pharmacotherapy. 2021;142:111953. DOI: 10.1016/j.biopha.2021.111953
43.Uehata T, Takeuchi O. RNA recognition and immunity-innate immune sensing and its posttranscriptional regulation mechanisms. Cells. 2020;9(7):1701. DOI: 10.3390/cells9071701
44.Liu A, Wang X. The pivotal role of chemical modifications in mRNA therapeutics. Frontiers in Cell and Developmental Biology. 2022;10:901510. DOI: 10.3389/fcell.2022.901510
45.Kaur K, Hadas Y, Kurian AA, Żak MM, Yoo J, Mahmood A, et al. Direct reprogramming induces vascular regeneration post muscle ischemic injury. Molecular Therapy: The Journal of the American Society of Gene Therapy. 2021;29(10):3042-3058. DOI: 10.1016/j.ymthe.2021.07.014
46.O'Brien J, Hayder H, Zayed Y, Peng C. Overview of MicroRNA biogenesis, mechanisms of actions, and circulation. Frontiers in Endocrinology. 2018;9:402. DOI: 10.3389/fendo.2018.00402
47.Yang C, Liu X, Zhao K, Zhu Y, Hu B, Zhou Y, et al. miRNA-21 promotes osteogenesis via the PTEN/PI3K/Akt/HIF-1α pathway and enhances bone regeneration in critical size defects. Stem Cell Research & Therapy. 2019;10(1):65. DOI: 10.1186/s13287-019-1168-2
48.He L, Zhu C, Jia J, Hao X-Y, Yu X-Y, Liu X-Y, et al. ADSC-exos containing MALAT1 promotes wound healing by targeting miR-124 through activating Wnt/β-catenin pathway. Bioscience Reports. 2020;40(5):BSR20192549. DOI: 10.1042/BSR20192549
49.Pinheiro A, Naya FJ. The key Lnc (RNA)s in cardiac and skeletal muscle development, regeneration, and disease. Journal of Cardiovascular Development and Disease. 2021;8(8):84. DOI: 10.3390/jcdd8080084
50.Tiwari A, Mukherjee B, Dixit M. MicroRNA key to angiogenesis regulation: MiRNA biology and therapy. Current Cancer Drug Targets. 2018;18(3):266-277. DOI: 10.2174/1568009617666170630142725
51.Fan C, Li Y, Lan T, Wang W, Long Y, Yu SY. Microglia secrete miR-146a-5p-containing exosomes to regulate neurogenesis in depression. Molecular Therapy: The Journal of the American Society of Gene Therapy. 2022;30(3):1300-1314. DOI: 10.1016/j.ymthe.2021.11.006
52.Gasparello J, Lomazzi M, Papi C, D'Aversa E, Sansone F, Casnati A, et al. Efficient delivery of MicroRNA and AntimiRNA molecules using an Argininocalix [4] arene macrocycle. Molecular Therapy. Nucleic Acids. 2019;18:748-763. DOI: 10.1016/j.omtn.2019.09.029
53.Wahid F, Shehzad A, Khan T, Kim YY. MicroRNAs: Synthesis, mechanism, function, and recent clinical trials. Biochimica et Biophysica Acta (BBA) - Molecular Cell Research. 2010;1803(11):1231-1243. DOI: 10.1016/j.bbamcr.2010.06.013
54.Lee SWL, Paoletti C, Campisi M, Osaki T, Adriani G, Kamm RD, et al. MicroRNA delivery through nanoparticles. Journal of Controlled Release: Official Journal of the Controlled Release Society. 2019;313:80-95. DOI: 10.1016/j.jconrel.2019.10.007
55.Zhang S, Cheng Z, Wang Y, Han T. The risks of miRNA therapeutics: In a drug target perspective. Drug Design, Development and Therapy. 2021;15:721-733. DOI: 10.2147/DDDT.S288859
56.Segal M, Slack FJ. Challenges identifying efficacious miRNA therapeutics for cancer. Expert Opinion on Drug Discovery. 2020;15(9):987-991. DOI: 10.1080/17460441.2020.1765770
57.Grath A, Dai G. Direct cell reprogramming for tissue engineering and regenerative medicine. Journal of Biological Engineering. 2019;13:14. DOI: 10.1186/s13036-019-0144-9
58.Yi BA, Mummery CL, Chien KR. Direct cardiomyocyte reprogramming: A new direction for cardiovascular regenerative medicine. Cold Spring Harbor Perspectives in Medicine. 2013;3(9):a014050. DOI: 10.1101/cshperspect.a014050
59.Sindhu C, Samavarchi-Tehrani P, Meissner A. Transcription factor-mediated epigenetic reprogramming. Journal of Biological Chemistry. 2012;287(37):30922-30931. DOI: 10.1074/jbc.R111.319046
61.Wang J, Place RF, Portnoy V, Huang V, Kang MR, Ho MKC, et al. Inducing gene expression by targeting promoter sequences using small activating RNAs. Journal of Biological Methods. 2015;2(1):e14. DOI: 10.14440/jbm.2015.39
62.Chong ZX, Yeap SK, Ho WY. Transfection types, methods and strategies: A technical review. PeerJ. 2021;9:e11165. DOI: 10.7717/peerj.11165
63.Seo B, Hong Y, Do J. Cellular reprogramming using protein and cell-penetrating peptides. International Journal of Molecular Sciences. 2017;18(3):552. DOI: 10.3390/ijms18030552
64.Kaitsuka T, Tomizawa K. Cell-penetrating peptide as a means of directing the differentiation of induced-pluripotent stem cells. International Journal of Molecular Sciences. 2015;16(11):26667-26676. DOI: 10.3390/ijms161125986
65.Ren X, Zhao M, Lash B, Martino MM, Julier Z. Growth factor engineering strategies for regenerative medicine applications. Frontiers in Bioengineering and Biotechnology. 2020;7:1-9. DOI: 10.3389/fbioe.2019.00469
66.Tada S, Kitajima T, Ito Y. Design and synthesis of binding growth factors. International Journal of Molecular Sciences. 2012;13(5):6053-6072. DOI: 10.3390/ijms13056053
67.Gillman CE, Jayasuriya AC. FDA-approved bone grafts and bone graft substitute devices in bone regeneration. Materials Science and Engineering: C. 2021;130:112466. DOI: 10.1016/j.msec.2021.112466
68.Niu Y, Li Q, Ding Y, Dong L, Wang C. Engineered delivery strategies for enhanced control of growth factor activities in wound healing. Advanced Drug Delivery Reviews. 2019;146:190-208. DOI: 10.1016/j.addr.2018.06.002
69.White SJ, Chong JJH. Growth factor therapy for cardiac repair: An overview of recent advances and future directions. Biophysical Reviews. 2020;12(4):805-815. DOI: 10.1007/s12551-020-00734-0
70.Mitchell AC, Briquez PS, Hubbell JA, Cochran JR. Engineering growth factors for regenerative medicine applications. Acta Biomaterialia. 2016;30:1-12. DOI: 10.1016/j.actbio.2015.11.007
71.Cochran JR, Kim Y-S, Lippow SM, Rao B, Wittrup KD. Improved mutants from directed evolution are biased to orthologous substitutions. Protein Engineering, Design and Selection. 2006;19(6):245-253. DOI: 10.1093/protein/gzl006. ISSN 1741-0134
72.PEGylated Protein Drugs: Basic Science and Clinical Applications. 10.1007/978-3-7643-8679-5
73.Cochran JR, Kim Y-S, Lippow SM, Rao B, Wittrup KD. Improved mutants from directed evolution are biased to orthologous substitutions. Protein Engineering, Design & Selection: PEDS. 2006;19(6):245-253. DOI: 10.1093/protein/gzl006
74.Vo TN, Kasper F, Kurtis; Mikos, Antonios G. Strategies for controlled delivery of growth factors and cells for bone regeneration. Advanced Drug Delivery Reviews. 2012;64(12):1292-1309. DOI: 10.1016/j.addr.2012.01.016
75.Stewart MC, Stewart AA. Mesenchymal stem cells: Characteristics, sources, and mechanisms of action. Veterinary Clinics of North America: Equine Practice. Cell-based Therapies in Orthopedics. 2011;27(2):243-261. DOI: 10.1016/j.cveq.2011.06.004
76.Liang X, Ding Y, Zhang Y, Tse H-F, Lian Q. Paracrine mechanisms of mesenchymal stem cell-based therapy: Current status and perspectives. Cell Transplantation. 2014;23(9):1045-1059. DOI: 10.3727/096368913X667709
77.Vizoso FJ, Eiro N, Cid S, Schneider J, Perez-Fernandez R. Mesenchymal stem cell Secretome: Toward cell-free therapeutic strategies in regenerative medicine. International Journal of Molecular Sciences. 2017;18(9):1852. DOI: 10.3390/ijms18091852
78.Akers JC, Gonda D, Kim R, Carter BS, Chen CC. Biogenesis of extracellular vesicles (EV): Exosomes, microvesicles, retrovirus-like vesicles, and apoptotic bodies. Journal of Neuro-Oncology. 2013;113(1):1-11. DOI: 10.1007/s11060-013-1084-8
79.Théry C, Zitvogel L, Amigorena S. Exosomes: Composition, biogenesis and function. Nature Reviews Immunology. 2002;2(8):569-579. DOI: 10.1038/nri855
80.Huda MN, Nafiujjaman M, Deaguero IG, Okonkwo J, Hill ML, Kim T, et al. Potential use of exosomes as diagnostic biomarkers and in targeted drug delivery: Progress in clinical and preclinical applications. ACS Biomaterials Science & Engineering. 2021;7(6):2106-2149. DOI: 10.1021/acsbiomaterials.1c00217
81.NV Lakshmi Kavya A, Subramanian S, Ramakrishna S. Therapeutic applications of exosomes in various diseases: A review. Biomaterials Advances. 2022;134:112579. DOI: 10.1016/j.msec.2021.112579
82.Wang G-J, Liu Y, Qin A, Shah SV, Deng Z-b, Xiang X, et al. Thymus exosomes-like particles induce regulatory T cells. The Journal of Immunology. 2008;181(8):5242-5248. DOI: 10.4049/jimmunol.181.8.5242
83.Rezaie J, Feghhi M, Etemadi T. A review on exosomes application in clinical trials: Perspective, questions, and challenges. Cell Communication and Signaling. 2022;20(1):145. DOI: 10.1186/s12964-022-00959-4
Written By
Archana Sreenivas and Deepak K. Jha
Submitted: 28 August 2024Reviewed: 06 December 2024Published: 11 March 2025
Open access peer-reviewed chapter - ONLINE FIRST
