Biologics and their delivery systems: Trends in myocardial infarction

Abstract

Cardiovascular disease is the leading cause of death around the world, in which myocardial infarction (MI) is a precipitating event. However, current therapies do not adequately address the multiple dysregulated systems following MI. Consequently, recent studies have developed novel biologic delivery systems to more effectively address these maladies. This review utilizes a scientometric summary of the recent literature to identify trends among biologic delivery systems designed to treat MI. Emphasis is placed on sustained or targeted release of biologics (e.g. growth factors, nucleic acids, stem cells, chemokines) from common delivery systems (e.g. microparticles, nanocarriers, injectable hydrogels, implantable patches). We also evaluate biologic delivery system trends in the entire regenerative medicine field to identify emerging approaches that may translate to the treatment of MI. Future developments include immune system targeting through soluble factor or chemokine delivery, and the development of advanced delivery systems that facilitate the synergistic delivery of biologics.

Keywords: Biologics; Chemokines; Controlled release; Drug delivery; Extracellular vesicle; Growth factors; Myocardial infarction; Nucleic acids; Scientometric review; Stem cells.

Figure 1 –. Pathophysiology of myocardial infarction and cardiac remodeling

(1) Localized hypoxia in the left ventricle due to disrupted blood flow causes cardiomyocytes (CM) necrosis (grey, top) and apoptosis (brown, bottom). (2) Upon reperfusion, inflammatory cells migrate to the tissue and become activated and differentiate into inflammatory cells by necrotic cell debris, damage associated molecular patterns, and inflammatory cytokines. This exacerbates CM necrosis and apoptosis. (3) Local inflammatory cells release soluble factors that also activate quiescent myofibroblasts to differentiate and release pro-fibrotic factors. (4) As a consequence, the infarcted myocardium replaces necrotic or damaged tissue (grey) with a fibrous scar (light blue) consisting of cross-linked collagen (blue lines). As remodeling progresses, more collagen is deposited and cross-linked, stiffening not only the infarct but also surrounding, healthy cardiac tissue referred to as the infarct border zone. (5) This occurs because the initial scar is not mechanically strong enough to overcome deforming forces during ventricle contraction (a) causing myofibroblasts to become or continue to be activated and secrete more fibrotic factors (b). This continues until the infarcted and surrounding tissue stiffens resulting in reduced contractility of the ventricle (c). Under these conditions, the renin-angiotensin-aldosterone system (RAAS) becomes activated and signals to the heart to increase contractile force and vasoconstriction in an attempt to increase blood flow and blood pressure (d). The increased contractile force is now able to displace the infarcted wall, maintaining this fibrotic cycle. Graphics were created with BioRender.com

Figure 2 –. Biologics employed to treat myocardial infarction

(A) Growth factors are employed to induce angiogenesis along a concentration gradient. (B) Similarly, chemokines attract various cell types from one organ in the body (Spleen, bone marrow) to traffic in a concentration dependent manner to a depot of chemokine being continuously released from a delivery system. (C) Various nucleic acids can be delivered to manipulate transcription and expression of mRNA or proteins. (D) Pluripotent stem cells can improve the infarct through replication to replace lost tissue and reduce collagen deposition or by releasing paracrine factors that can induce angiogenesis, prevent/reduce CM apoptosis, or modulate the immune response. Graphics were created with BioRender.com, Fig 2.1D was also created with Blender 3D modelling & rendering package [357].

Figure 3 –. Delivery devices for cardiac delivery of biologics

(A) Inorganic nanocarriers – quantum dots, gold nanoparticles, mesoporous silica nanoparticles, polymeric micelle (left to right) – and organic nanocarriers – viral, dendrimer, liposome, lipid micelle (left to right) – with representative images of systemic distribution of a nanocarrier loaded with GFP 3 hours after administration (top) and 6 hours after administration (bottom) following MI induction. (B) Representative histology images of microparticle retention in MI denoted by white arrows (top) and characteristic SEM images (bottom). (C) Methods of hydrogel formation and responsive gelation (top), sequestration of biologics (pink sphere) can occur within the matrix, on the hydrogel backbone, or within matrix connections (bottom-left). Upon degradation or through passive diffusion, biologics are released (bottom-right). Representative histology showing hydrogel applied to an infarct to increase ventricle wall thickness, enhanced zoom is representative of the boxed region and arrows indicate hydrogel (black) and myocardium staining positive for α-smc (blue). (D) Implantable cardiac patches applied over the infarcted region of the heart include dense polymeric structures (top) or porous meshes (bottom). Representative H&E stained histology of an implanted delivery system in an infarcted heart shows the ability to increase the wall thickness (top thickness compared to mid-left thickness). Graphics were created with BioRender.com and Blender 3D modelling & rendering package [357]. Fig. 2.2A reprinted from [70]. Copyright (2018) Nature. Fig. 2.2B reprinted from [56] & [59]. Copyright (2016) with permission from Elsevier and (2015) American Physiological Society, respectively. Fig. 2.2C reprinted from [358]. Copyright (2009) with permission from Elsevier. Fig. 2.2D reprinted from [56] & [61]. Copyright (2015) with and (2016), respectively, with permission from Elsevier.

Figure 4 –. Biologic delivery trends in myocardial infarction research

Chord diagrams displaying the relationship between therapeutic targets and delivery systems (A), delivery systems and biologics (B), and therapeutic targets and biologics (C) were constructed according the literary search detailed in the methods section. Connecting ribbons display the strength of the correlation (thickness) and percentages listed along the grid for specific classes of targets, delivery systems, or biologics correspond to their contribution to their respective classifier as it relates to the other. For example, Figure 4C shows 35% of all biologics utilized among the therapeutic targets in MI that were considered are Growth Factors, of which the majority (nearly half) are employed to induce angiogenesis. Represents data for references [52-76, 78, 81-109, 195, 237, 238] obtained on Elsevier’s Scopus search engine for dates 1/1/2015 – 5/14/2020 inclusive. Chord diagrams were generated in RStudio version 3.6.1 using the circlize package [356].

Figure 5 –. Drug delivery system types and attributes

Delivery systems shown in Figure 2 (Nanocarriers, Hydrogels, Implants, and Microparticles) are further described using a Sankey Diagram created via sankeymatic.com beta software [359]. Data shown is the number of publications identified by the searching method – for brevity, only the delivery systems have the publication number shown – the individual data and reference number can be found in the supplementary data for this publication (Table 1 & 2). The distribution of delivery systems directly corresponds to Figure 2 data obtained from the coincidence search while subsequent classification was performed manually, in absence of appropriate keywords. Only nanocarriers were quantified as being locally or systemically administered because the other DDS lack the ability to be systemically administered. In some cases, applications could not be adequately described and so they were omitted (e.g. one non-viral application was in vitro only and so was not locally or systemically administered to treat MI). Data was obtained on Elsevier’s Scopus search engine for dates 1/1/2015 – 5/14/2020, inclusive. Represents data for references [52-76, 78, 81-109, 128, 143, 170, 195, 223-238, 250, 251] obtained on Elsevier’s Scopus search engine for dates 1/1/2015 – 5/14/2020, inclusive.

Figure 6 –. Synergistic delivery enhances angiogenesis

IHC staining of pericytes (α-SMA+, red) and endothelial cells (CD31+, green) 21 days following injection of saline, sulfonated reverse thermal gel (SRTG), SRTG + VEGF, SRTG + VEGF + PDGF, SRTG + VEGF + PDGF-loaded micelles, or SRTG + VEGF + IL-10 + PDGF-loaded micelles. Scale bar = 100 μm. Reprinted from [65]. Copyright (2020) American Chemical Society.

Figure 7 –. In vitro seeding of CSCs expressing cardiomyocyte markers.

Microphotographs of PLLA mesh [24] populated with CSCs after 7 days. The cells formed a tissue-like structure and expressed canonical CM markers – cTnI, Mhc, and α-SA. Scale bar = 50 μm. Reprinted from [61]. Copyright (2015) with permission from Elsevier.

Figure 8 –. Biologic types and attributes

Biologics shown in Figure 2 (Growth Factors, Nucleic Acids, Chemokines, and Stem Cells) are further described using a Sankey Diagram created via sankeymatic.com beta software [359]. Data shown is the number of publications identified by the searching method – for brevity, only the Biologics have the publication number shown – the individual data and reference number can be found in the supplementary data for this publication (Supplementary Table 1 & 2). The distribution of biologics directly corresponds to Figure 2 data obtained from the coincidence search while subsequent classification was performed manually, in absence of appropriate keywords. Because of multi-faceted therapy strategies (e.g. delivery of multiple growth factors or nucleic acids), which caused an increase in the number of examples where specific types of biologics were administered causing the diagram to appear unbalanced (e.g. Growth Factors, Nucleic Acids). Represents data for references [52-76, 78, 81-109, 128, 143, 170, 195, 223-238, 250, 251] obtained on Elsevier’s Scopus search engine for dates 1/1/2015 – 5/14/2020 inclusive

Figure 9 –. IMT-VEGF infarct targeting

Dylight 488 fluorophore (left), complexed to VEGF (middle), or complexed to IMT-VEGF (right) was injected intravenously following MI induction and a reperfusion duration of 10 minutes. One hour following injection, the heart was washed with PBS, removed, and sectioned. Reprinted from [54]. Copyright (2015) with permission from Elsevier.

Figure 10 –. Novel approaches to target delivery to the infarct

(1.) Synthetic modifications of polypeptide molecules to include a peptide binding domain specific to proteins upregulated in the infarct. (2.) Electrostatic coating (top) or synthetic modification of polypeptide molecules to include a binding domain specific for ECM proteins upregulated following MI. (3.) Steric hindrance of viral spike proteins prevents cell transduction. (4.) Enzymatic cleavage of the peptide inhibiting spike protein binding facilitates localized cell transduction. Graphics were created with BioRender.com

Figure 11 –. TANNylation of GFP

(1.) Mixing green fluorescent protein (GFP) with tannic acid (TA) results in the formation of aggregated nanoparticles that (2.) have the ability to bind to the ECM without binding to glycocalyx located in systemic vasculature. Reprinted from [70]. Copyright (2018) Nature.

Figure 12 –. Encapsulation and release of VEGF and PDGF from a coacervate & hydrogel system

(A) Conceptual diagram detailing how PDGF and VEGF were encapsulated in the delivery system. (B) Cumulative release of VEGF and PDGF, in which the loading efficiency was 87% and 97%, respectively, for 100 ng attempted encapsulation. Reprinted from [64]. Copyright (2015) with permission from Elsevier.

Figure 13 –. Electrostatic loading of both siRNA and pDNA using an amphiphilic cationic polymer.

Reprinted from [53]. Copyright (2016) with permission from Elsevier.

Figure 14 –. EV delivery of miRNA-181a improves heart function by increasing Foxp3+ Treg cells

Reprinted from [104]. Copyright (2019) with permission from Elsevier.

Figure 15 –. Modification of siRNA to include a cholesterol molecule can facilitate increased transduction and encapsulation.

An MMP-degradable, hydrazide-modified HA macromer (HA-MMP-HYD) and a cyclodextrin and aldehyde-modified HA macromer (CD-ALD-HA) were combined to form an MMP-2 responsive hydrogel. The CD group provided a binding site for cholesterol, facilitating loading of cholesterol-modified siRNA. The inclusion of this cholesterol group can facilitate improved cell transduction of the nucleic acid. Reprinted from [107]. Copyright (2018) with permission from Elsevier.

Figure 16.. Characterization of LBL-coated MSC function.

(A) Representative SEM images of MSCs (top) and coated MSCs (bottom) for day 1, 3, and 7 of culture. (B). Quantification of cell spreading indicates that LBL MSCs have lagged spreading rate relative to non-coated. (C). LBL coated MSC recruitment in a scratch assay is also lagged relative to non-coated in a scratch test assay. Reprinted from [52]. Copyright (2017) with permission from Elsevier.

Figure 17 –. 5-year changes in biologic delivery research:

Chord diagrams displaying the individual changes (ribbons) and their net impact (outer grid) for research conducted in 2015-2019 as compared to the previous 5 year period, 2010-2014. Chords represent the relationship between therapeutic targets and delivery systems (A), delivery systems and biologics (B), and therapeutic target and biologics (C). Individual changes are quantified in the table, in which the cell color, green denotes a gain and red denotes a loss, matches the ribbon color in A-C. (D). Delivery system use has shown a prominent increase in hydrogel usage, more therapies/studies have targeted/explored immunomodulation, and nominal reductions in studies employing nucleic acids has translated to increases in chemokine, growth factor, and stem cell delivery. Data was obtained on Elsevier’s Scopus search engine for dates 1/1/2015 – 5/14/2020, inclusive. Chord diagrams were generated in RStudio version 3.6.1 using the circlize package [356].

Figure 18 –. Compressed microsphere cardiac patch

(Top) Conceptual diagram showing the application of a compressed microsphere patch secured by a chitosan sheet. (Bottom) Representative image of a microsphere patch implanted onto a rat heart at day 0 (left) and MRI at day 56 (right). Arrows denote the edges of the patch. Reprinted from [56]. Copyright (2016) with permission from Elsevier.

Figure 19 –. CSC-loaded microneedle array patch

An example microneedle array (MNA) swells to form a hydrogel facilitating paracrine factor release to the infarcted tissue and CSC migration. The MNA also acts as a scaffold for the implanted cells. Reprinted from [198]. Copyright (2018) AAAS.