Theoretical basis, state and challenges of living cell-based drug delivery systems

Abstract

The therapeutic efficacy of drugs is determined, to a certain extent, by the efficiency of drug delivery. The low efficiency of drug delivery systems (DDSs) is frequently associated with serious toxic side effects and can even prove fatal in certain cases. With the rapid development of technology, drug delivery has evolved from using traditional frameworks to using nano DDSs (NDDSs), endogenous biomaterials DDSs (EBDDSs), and living cell DDSs (LCDDSs). LCDDSs are receiving widespread attention from researchers at present owing to the unique advantages of living cells in targeted drug delivery, including their excellent biocompatibility properties, low immunogenicity, unique biological properties and functions, and role in the treatment of diseases. However, the theoretical basis and techniques involved in the application of LCDDSs have not been extensively summarized to date. Therefore, this review comprehensively summarizes the properties and applications of living cells, elaborates the various drug loading approaches and controlled drug release, and discusses the results of clinical trials. The review also discusses the current shortcomings and prospects for the future development of LCDDSs, which will serve as highly valuable insights for the development and clinical transformation of LCDDSs in the future.

Keywords: Clinical transformation; Controlled drug release; Drug loading approaches; Living cells; Targeted drug delivery.

Figure 1

Living cell drug delivery system, from application to loading techniques to controlled release (Figure was created with BioRender.com).

Figure 2

The properties of various living cell carriers (Figure was created with BioRender.com).

Figure 3

Drug loading strategies-based living cells, encompassing extracellular and intracellular drug loading approaches, as well as genetically engineered (Figure was created with BioRender.com).

Figure 4

Relevant illustration of drug loading techniques involving physical binding with living cells. (A) T-TMP was synthesized through self-assembling TMP and FPR targeting peptide with PLGA-TK-PEG-pep, followed by specific targeting neutrophils through the FRP receptor. (B) Confocal images exhibit neutrophils have a enhanced red fluorescence of indocyanine green Cy5-labeled T-TMP and NT-TMP. (C) Flowcytometry results shows the uptake of T-TMP by neutrophils. (A-C Reproduced with permission Copyright 2023, Wiley-VCH GmbH). (D) (left)Three layers cellular backpack assembly; and (right) Confocal micrographs of cells (nucleus, blue; membrane, green; backpacks: red). (Left, created with BioRender.com; Right, Reproduced with permission copyright 2020, The American Association for the Advancement of Science). (E) The scheme of NEs@STING-Mal-NP were synthesized by the covalent binding between Mal on the surface of STING-Mal-NP and the thiols on the surface of neutrophils after reduction reaction by TCEP. (F) Confocal microscopy image of neutrophils with or without TCEP reduction after incubation with STING-Mal-NP-Rhodamine (red). Nucleus was stained by Hoechst (blue) (E-F Reproduced with permission copyright 2023, American Chemical Society). (G) The tannic acid encapsulates granzyme B and perforin formed nanocomplex, and the electrostatic interaction or hydrophobic interaction between the polyphenol moieties of nanocomplexes and platelet surface. (H) The SEM images of platelets and PDCs, which demonstrated that PDCs had a rougher surface structure compared with unmodified platelets. (G-H Reproduced with permission copyright 2023, Elsevier).

Figure 5

Relevant illustration of drug loading techniques involving chemical binding with living cells. (A) Schematic representation of the host-guest complex formed by β-CD and ferrocene. β-CD, β-cyclodextrin. (B) SEM and (C) CLSM images are obtained for red blood cells (RBCs) incubated sequentially with DSPE-PEG-CD and ferrocene-NP (RBC-NP), as well as RBCs coated with Fc-NP (without β-CD) on their surface (RBC+NP). The green fluorescence indicated the presence of DIO-loaded ferrocene-NP. (A-C Reproduced with permission copyright 2022, Elsevier). (D) Schematic illustration of click chemistry reaction. TK-M/Lu were prepared by using solvent evaporation method and subsequently tethered onto the surface of MSCs via bioorthogonal click chemistry. (E) SEM images revealed the attachment of luteolin-loaded micelles with (MSC-DB-M/Lu) or without DBCO (MSC-TK-M/Lu), to the surface of MSCs. Respectively. (D-E Reproduced with permission copyright 2023, Elsevier). (F) Schematic representation of the fabrication procedure for macrophage-liposome complexes. The DSPE-PEG-biotin-modified liposomes are conjugated to streptavidin (STA)-modified macrophages through STA-biotin interaction. (G) CLSM images of the physical mixture of FITC-PEG-biotin and DSPE-PEG-STA-modified macrophages, while the undecorated cells showed no significant attachment. (H) Evaluation of the migratory capacity of macrophages with and without DSPE-PEG-STA modification revealed that surface modification did not significantly impact macrophage motility, as evidenced by a cell migration assay (F-H Reproduced with permission copyright 2022, American Chemical Society).

Figure 6

Relevant illustration of drug intracellular loading techniques with living cells. (A) Neutrophils sense, capture, and engulf pathogens by recognizing the PAMPs with TLRs (left), Preparation of NPNs by coating OMVs on NPs, which inherit PAMPs from the OMVs (right). (B) Treatment-induced cell death created an inflammatory environment of the residual tumor and induced the production of G-CSF, GM-CSF, and chemokines CXCL1 and MIP-2. #1a The released G-CSF and GM-CSF increased neutrophil production from bone marrow. #1b The released CXCL1 and MIP-2 broadcasted the location of the inflamed tumor. #2 Neutrophils entered the blood circulation and encountered the injected NPNs. #3 Neutrophils sensed NPNs with the recognition of LPS and lipoprotein by TLRs and subsequently engulfed them. #4 Neutrophils laden with NPNs were recruited into the tumor site in response to the chemokine gradient through the following cascade: adhesion, crawling and transmigration. #5 NPNs were released from neutrophils to kill tumor cells along with the formation of NETs in the inflamed tumor (A-B Reproduced with permission copyright 2020, Springer Nature). (C) Schematic of the erythrocyte shape change in the drug-loading process: tsw, tst, tpc, and tep denote the swelling time, stretching time, pore opening-closing time, and loading time, respectively (Reproduced with permission copyright 2018, Springer Nature). (D) Rearrangement of the cell membrane at the molecular level (left) and atomic level (right) under an electric field. Intact bilayer. Intact bilayer (top). Process of water molecules penetrating the bilayer (middle). Reorientation of lipids (bottom) (Reproduced with permission copyright 2022, Royal Society of Chemistry). (E) Schematic diagram of ultrasound-guided drug delivery. Ultrasound triggers microbubble oscillation (expansion and shrinkage) and collapse, causing vessel deformation, rupture and permeability change, which allows efficient drug delivery in spatiotemporally controlled way. (Reproduced with permission copyright 2020, Elsevier).

Figure 7

Relevant illustration of drug loading techniques involving genetically engineered. (A) Human pluripotent stem cells were engineered with CARs and differentiated into CAR-neutrophils that are loaded with rough silica nanoparticles containing hypoxia-targeting tirapazamine or other drugs, as a dual immunochemotherapy. (B-C) The neutrophils loaded with smooth and rough SiO₂-TPZ NPs, and detected significant cellular uptake of rough SiO2-TPZ NPs than smooth SiO₂-TPZ NPs via fluorescence microscope and flow cytometry analysis. (A-C Reproduced with permission copyright 2023, Springer Nature) (D) Schematic of workflow for therapeutic uses of bioengineered enucleated cells (Cargocytes). (E) Fluorescent confocal images show that the hT-MSC-derived Cargocytes (“Cargocytes”) maintain well-organized cytoskeletal structure. The hT-MSCs/Cargocytes stained with rhodamine phalloidin for F-actin cytoskeleton (left), or anti-α-Tubulin antibody for microtubule network (right), and Hoechst for nucleus. (F) Graph shows cell surface expression of CXCR4 by flow cytometry. MSC ^CXCR4, CXCR4 lentivirus-engineered hat-MSC; 2hr/24hr/48hr Cargocytes, MSC^CXCR4-derived Cargocytes analyzed at indicated time points post-enucleation. (G) MSCs/Cargocytes migrated in Boyden chambers towards the indicated concentrations of SDF-1α for 2hr. Bar graph represents the cell migration index (migrated MSCs/Cargocytes versus loading control) (D-G Reproduced with permission copyright 2023, Springer Nature).

Figure 8

The mechanisms underlying drug controlled release and their in vivo fate (Figure was created with BioRender.com).

Figure 9

Relevant illustration of drug controlled release through NETs, PMPs, and damage to cell membrane structures. (A) The progression for a neutrophil from apoptosis to NETosis. When a healthy neutrophil receives apoptotic, necroptotic, or pyroptotic stimuli, the cell will open a pore in the membrane (GSDME, MLKL, or GSDMD, respectively). These pores permit calcium influx, causing PAD4 activation, which converts positively charged arginine residues in histone to neutrally charged citrulline. This causes DNA to unwind from histones. Given enough time, PAD4 activity causes the neutrophil to extrude its DNA in the process of NETosis, forming NETs. Created with BioRender.com. (B) Resting neutrophils are round and devoid of fibers (left), The activated cells in (middle) have many pseudopods and show NETs (arrow). Analysis of cross sections of the NETs by transmission electron microscopy (TEM) revealed they were not surrounded by membranes (right). (Reproduced with permission copyright 2004, The American Association for the Advancement of Science) (C) Schematic indicate PMPs released from the PDCs to activated-PLTs. Created with BioRender.com. (D) TEM images of (i) platelet-drug conjugates, (ii) activated PDCs, (iii) PMP generation, and (iv) PMPs from PDCs. Red arrows indicate PMPs (above). Confocal fluorescence images of Cy3-labeled platelet-drug conjugates pre-and post-activation (Reproduced with permission copyright 2023, Elsevier). (E) The formation of the hydrogel in which laser radiation triggers the release of insulin (INS). (F) The in vitro release profile of INS when the laser was cyclically varied between on and off for several repetitions. The concentration of INS increased as the laser was turned on, while no more INS was released from INS-ICG@RBC hydrogel when the laser was turned off . (G) The morphological changes of INS-ICG@ER during the “on-off” INS release (E-G Reproduced with permission copyright 2023, Elsevier).

Figure 10

Conclusion and perspective of LCDDS (Figure was created with BioRender.com).