Allogeneic cardiac stem cell administration for acute myocardial infarction.

Review

Expert Review of Cardiovascular Therapy Downloaded from informahealthcare.com by Nyu Medical Center on 02/11/15 For personal use only.

Allogeneic cardiac stem cell administration for acute myocardial infarction Expert Rev. Cardiovasc. Ther. Early online, 1–15 (2015)

Veronica Crisostomo*, Javier G Casado, Claudia Baez-Diaz, Rebeca Blazquez and Francisco M Sanchez-Margallo Jesu´s Uson Minimally Invasive Surgery Centre, Carretera N-521, km 41.8, 10071 – Caćeres, Spain *Author for correspondence: Tel.: +34 927 181 032 Fax: +34 927 181 033 [email protected]

Myocardial infarction, even after reperfusion, leads to significant loss of cardiomyocytes and to a maladaptive remodeling process. A possibility gaining attention as an ancillary therapy is the use of cardiac-derived cell products, with early stage clinical trials reporting highly promising results with autologous cells. However, an autologous therapy presents limitations, such as timeframe of therapy, cell processing and culture costs, risks posed to the patient by the tissue harvesting, etc. Allogeneic cells may represent an answer, providing an off-the-shelf product that could be used in the acute stage, before the myocardial damage is irrevocable. To date, allogeneic cardiac-derived cell products are being tested extensively, but the questions of their immunogenicity (and therefore safety), efficacy, cost–effectiveness, etc. are only partially elucidated. Small Phase I/II clinical trials (ALLSTAR, CAREMI) have started and their results will shed the much needed light on the feasibility and safety of a much needed therapy. KEYWORDS: allogeneic . animal models . cardiac stem cells . experimental therapy . myocardial infarction . myocardial regeneration

Cardiovascular diseases remain a major cause of death and disability in developed countries, with coronary heart disease being responsible for almost 1.8 million deaths per year in Europe (20% of all deaths) [1] and accounting for over 379,000 deaths in the USA in 2010 [2]. Adult stem cells and their possible application for regenerative medicine after myocardial infarction are emerging as a therapeutic option, with most animal and human studies performed with the different available cell types reporting a modest improvement in cardiac function after cell transplantation [3,4]. Clinical and preclinical studies have shown that stem cells may improve cardiac function after a myocardial infarction, but the optimal type of stem cells to be used for this purpose remains to be established [5]. Nonetheless, cell-based therapies aiming at myocardial regeneration have the potential to transform the clinical management and prognosis of myocardial infarction and heart failure. Among the cell types that could be used in an allogeneic setting currently under evaluation, mesenchymal stem cells (MSCs) have been studied extensively. Originally isolated from the bone

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marrow, they have now been obtained from a variety of tissues. These cells have been shown to have multilineage potential, immunomodulatory properties and are able to secrete different cardioprotective growth factors. In the POSEIDON trial [6], Hare et al. compared the effects of increasing doses of autologous or allogeneic MSCs (20, 100 and 200 106 cells) in ischemic cardiomyopathy patients, reporting a similar safety profile and an improvement in functional capacity, ameliorated ventricular remodeling and improved quality of life following MSC administration, independently of their allogeneic or autologous nature. Contrary to other works [7], they report a better effect of the lower dose, highlighting the importance of dose-finding studies. Historically, the heart has been considered as a terminally differentiated organ, with no self-renewal ability. Since the intrinsic regenerative capacity of cardiac tissue started to be explored at the early 90s, multiple research groups have independently shown the presence of a regenerative mechanism in the adult mammalian heart [8], thus paving the way for

2015 Informa UK Ltd

ISSN 1477-9072

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Crisostomo, Casado, Baez-Diaz, Blazquez & Sanchez-Margallo

a new approach. In recent years, different groups have shown the presence of different cell populations in the adult heart that share some characteristics with stem cells. These cardiac-derived cellular products are receiving increasing interest as they are considered to posses the necessary properties to achieve cardiac regeneration via paracrine activation and trilineage differentiation [9], and have been proved in numerous preclinical studies to improve left ventricular function and attenuate remodeling after myocardial infarction [10–18]. Based on these experimental works, human clinical trials have been conducted, and highly promising results have recently been reported from the first clinical trials to use autologous cardiac cells in the setting of heart failure, either after coronary artery bypass grafting [19,20] (SCIPIO trial, [21]) or after coronary stenting [22,23] (CADUCEUS trial, [24]). However, the autologous approach presents obvious drawbacks in that the time needed for cell expansion conditions the time frame for treatment. Moreover, the regenerative potential of cells obtained from cardiac patients has been called into question [25], although other groups have reported improved potential when injecting cells of cardiac origin derived from heart failure patients in acutely infarcted mice [26]. Allogeneic cells emerge as a promising option, and several studies using this strategy have been reported [13,27–31]. The advantages of using an allogeneic product are varied. They offer the possibility of administering the cells very early after the ischemic event as an ‘off-the-shelf’ product that, at the same time, can be subjected to higher quality controls than autologous products [32], circumventing the possibility of failed culture, which is a risk inherent to all cell processing protocols. Moreover, this therapy obviates the need for endomyocardial biopsies and their potential risks for the already sick patients, while the intracoronary infusion of these cells can be performed easily in any interventional catheterization laboratory worldwide. Several groups [3,9,32,33] have advocated the earliest possible administration time based on the experimental data documenting a beneficial effect on the cells at risk by the paracrine action of the exogenous cells, which would only be possible when administering an allogeneic product. In the subject of cell therapy for the damaged heart, clinical translation of preclinical studies has been extremely fast, forcing the field to a certain amount of caution and warranting a ‘back to the bench’ trend [34], which nonetheless needs to be balanced with a rapid translation to the patient, given the astonishing potential these cardiac-derived products have. This is even more important when considering allogeneic therapy, where the potential for rejection or allosensitization needs to be carefully explored prior to proceeding to clinical practice. This review aims to provide an overview of the current state of cardiac-derived cells for cardiac regeneration, focused on their possible allogeneic use; trying to encompass the work performed in small and large animal models as well as the little clinical data available to date. While we aim to give an overview of the whole picture, we do not pretend to be comprehensive, since the work done in the field is extensive. We have therefore attempted to exclude from this review works that doi: 10.1586/14779072.2015.1011621

have no direct bearing in the development of an allogeneic therapy. The interested reader is referred to the reviews published recently regarding cardiospheres (CS) and cardiospherederived cells (CDCs) [35], c-kit+/Lin- cardiac stem cells (CSCs) [36] and, more globally, the use of cell therapy for heart failure [5]. Stem cell types obtained from the heart

Different groups have shown the presence of different cell populations in the adult heart that share some characteristics with stem cells. These cells, called CSCs, would be involved in the replacement of senescent or apoptotic cardiac cells, allowing the maintenance of cardiac homeostasis. They are self-renewing, clonogenic and multipotent, being able to differentiate into cardiomyocytes, smooth muscle cells and endothelial cells under in vitro stimulation. CSCs would also contribute to coronary vessels regeneration, in addition to cardiomyocytes [37]. During ischemic processes, several paracrine signals stimulate these CSCs to divide. However, these divisions are insufficient to replace the cell loss caused by a myocardial infarction [38]. In several studies, CSCs have been isolated and expanded from human myocardium biopsies, offering a new source of stem cells for cardiovascular regenerative medicine [39]. The different populations of CSCs have been classified mainly according to the expression of different surface markers. These heart-derived cell populations have shown great promise in the first Phase I clinical trials reported [19,20,22,23], with potentially very exciting results up to 2 years after treatment. There appears to be an inherent heterogeneity of heart-derived cellular products, with varied endogenous populations having been described. The relationship between the different populations described to date awaits clarification through comprehensive characterization, and whether one or more markers may be expressed in the same cell could help explain the controversy regarding the number of different CSCs [40]. In this review, some of the most relevant populations are shown in FIGURE 1. c-kit+ progenitor cells

Beltrami et al. first reported the existence of a resident population of CSCs in the rat heart. They described a c-kit+/Linpopulation which is self-renewing, clonogenic, multipotent and able to differentiate into the three cardiogenic cell lineages (myocytes, smooth muscle cells and endothelial cells) [41]. First described by Yarden et al. in 1987 as a proto-oncogene [42], c-kit is a tyrosine kinase receptor expressed at high levels in hematopoietic stem cells, multipotent progenitors, common myeloid progenitors and less abundantly in other cell types. The ligand for this receptor has been described as stem cell factor (a substance that causes certain types of cells to grow), also known as steel factor or c-kit ligand. The signaling pathways downstream of the c-kit receptor are essential for regulating proliferation, survival and other vital functions in early hematopoietic cells [43]. The c-kit+ CSCs were originally considered a population apart from other progenitor cells as they did not appear to Expert Rev. Cardiovasc. Ther.


Allogeneic CSC administration for acute myocardial infarction

express endothelial or bone marrow progenitor markers, such as CD34 or CD45, expressed in hematopoietic stem cells. They were also negative for the transcription factors and cytoplasmic proteins found in mature cardiomyocytes, endothelial cells and smooth muscle cells, including Isl1 [44]. However, other studies have documented the presence of different subpopulation of c-kit+ CSCs, some of them expressing mesenchymal and endothelial markers [45,46].

Review

c-kit + CPCs

SP CPCs

Sca-1 + CPCs

c-kit Sca-1 Isl-1

Sca-1+ progenitor cells

CD90 In 2003, Oh et al. reported the isolation of a ‘myocyte depleted’ fraction of adult carCD105 diac cells expressing Sca-1 and CD31, but Abcg2 negative for blood cell lineage markers Mef2c (CD34, CD8, B220; Gr-1, Mac-1 and Nkx2.5 TER119). These cells were also positive Isl1 + CPCs CS/CDCs for c-kit, Flt-1, Flk-1, vascular endothelialcadherin, von Willebrand factor and Figure 1. Distribution of the main surface markers in the different CPC populations. The expression of the different surface markers, detailed in the figure leghematopoietic stem cell markers end, is represented for each of the cardiac progenitor cell population referred in the text. CD45 and CD34. They showed telomeCDCs: Cardiosphere-derived cells; CPCs: Cardiac progenitor cells; CS: Cardiospheres; rase activity and were able to differentiate SP: Side population. into mature cardiomyocytes when cultured in the presence of 5-azacytidine [47]. Sca-1 was first reported as a cell surface marker of hemato- potential than Sca+/CD31- cells. In vitro, SP cells can differenpoietic stem cells belonging to the Ly-6 antigen family [48]. tiate into cardiomyocytes, smooth muscle cells and endothelial Recently, many reports have demonstrated that multipotent cells, and also into glial cells and neurons, suggesting that the stem cells derived from bone marrow and skeletal muscle also origin of these cells could be in the neural crest [54,56]. More than 93% of adult cardiac SP cells are express Sca-1. These cells are able to differentiate in vitro into Sca-1+CD45-CD34-, exhibit levels of telomerase expression simdifferent lineages (hepatocytes, cardiomyocytes, skeletal muscle, ilar to neonatal myocardium and express cardiogenic transcripneural cells, endothelial cells, adipocytes and myocytes) if they tion factors. Moreover, these cells do not express cardiac are submitted to the appropriate stimuli [49]. Sca-1 plays a role structural genes until stimulated to differentiate in vitro [50]. in myoblast differentiation, proliferation, fusion and cell-cycle However, these cells are relatively rare in the adult tissue, and exit. It appears to downregulate muscle cell proliferation, their ability to contribute to the regeneration and functional thereby maintaining a pool of functional progenitor cells for repair of damaged cardiac muscle remains to be determined [40]. muscle homeostasis and repair [50].

Side population cells

Isl1+ progenitor cells

Side population (SP) cells were first described by Goodell et al. as a subpopulation of murine bone marrow cells able to block the Hoechst 33342 dye efflux activity [51]. Some years later, the existence of a cardiac SP was reported by Pfister et al. These cells were CD31-/Sca-1+ and they were capable of cardiomyogenic differentiation into mature cardiomyocytes through a process mediated by cellular coupling with adult cardiomyocytes [52] or in the presence of cardiomyogenic agents such as 5-azacytidine [53], oxytocin or trichostatin A [54]. This was the first population isolated directly from digested heart tissue. Cardiac SP cells are highly enriched for Sca-1, suggesting a possible relationship between Sca-1+ cells and SP cells in the heart [55]. It has also been described as a Sca+/CD31+ subset in the isolated SP cells that seem to have lower cardiogenic

Isl1 is a LIM-homeodomain transcription factor that controls cardiomyocyte specification and differentiation. Isl1 is a marker of myocardial lineage during mammalian cardiogenesis and identifies a common population of progenitor cells in the heart that can differentiate into cardiomyocytes, smooth muscle and endothelial cells. Two-thirds of the cells within the entire heart originate from Isl1-positive progenitor cells [57]. This transcription factor promotes postnatal angiogenesis and vasculogenesis by improving the angiogenic properties of endothelial cells and MSCs [58]. Isl1 is a characteristic marker of an abundant population in the embryonic heart. Some of these cells remain undifferentiated even after the complete heart formation but its number decreases gradually [44]. In the adult heart, some


doi: 10.1586/14779072.2015.1011621

Review


Isl1+ cardioblasts can be found in atria and ventricles as isolated cells. They also express early cardiac differentiation markers such as Nkx2.5 and GATA4 but lack mature cardiomyocytes transcription factors, Sca-1 and c-kit [37]. These cells, however, have only been described to date in neonatal specimens, so their application for cardiac regeneration may be limited [59].


CS & CDCs

In 2004, Messina et al. described a new isolation technique for cardiac resident progenitor cells from murine hearts as well as from human atrium and ventricle subcultures. After enzymatic digestion of the tissue and seeding in culture flasks, a monolayer of adherent cells with fibroblastic morphology grew in the flasks. Some days later, some small, round and bright cells started to migrate over this monolayer, being in suspension. When these cells were seeded in poly-D-lysine-coated flasks, they gave rise to cluster formations that were called CS. These cells were clonogenic and expressed endothelial progenitors and pluripotency markers, showing the same properties as cardiac progenitor cells (CPCs). They were able to self-renew and differentiate into cardiac specialized cells as cardiomyocytes and vascular cells, either in vivo and in vitro [60]. These CSCs growing as CS represent per se an actual cardiac microtissue, where the most differentiated cells are localized in the outer layers, while those with a higher differentiation potential and expression of stem cell and angiogenesis markers are in the inner core [61,62]. Three years after their first report, Smith et al. used CS to obtain CDCs from human and porcine tissue. They compared both phenotypes, finding several similarities and slight differences [63]. Human CS are c-kit+ and CD105+ (one of the regulators for the TGF-b receptor, important for angiogenesis and hematopoiesis processes). CS also show a high expression of connexin 43, a component of the tight junctions between cells. CDCs also express connexin 43, which suggests an electric coupling potential between these cells as well as between them and cardiac myocytes. CDCs are also positive for CD90, CD34 and CD31 and negative for CD133, CD45 and blood antigens [60,63]. Other considerations

Even though the main subpopulations are characterized by the expression of c-kit, Sca-1 or Isl1 separately, Di Felice et al. first reported the concomitant expression of these three markers in a population of CPCs isolated without any sorting or selection. They suggested for the first time that there could be a unique population of cells with several subpopulations and that the several markers identified could be only the effect of different cultivation conditions in the different laboratories [64,65]. Also, the origin of CPCs has been widely discussed, as the involvement of the bone marrow in human cardiac chimerism has been described and the implication of extrinsic cellular regulatory processes have been proposed to participate in cardiac repair after injury, despite the fact that, in the absence of myocardial damage, circulating bone marrow progenitors cells do doi: 10.1586/14779072.2015.1011621

not home to the heart [45]. According to that, although in the past years the characterization and description of different subpopulations of CSCs have been the aim of several studies, it is necessary to respond to a major question that many authors are nowadays trying to solve: Are they real subpopulations or could they be just phenotypic variations of only one cell type? If so, a better knowledge of the different markers expressed in each variation will be needed to select the best therapeutic option for cardiac repair [50]. Finally, other selection methods apart from cell surface markers should also be considered. In this sense, a new CSC subpopulation has been recently described by Koninckx et al. based on the high aldehyde dehydrogenase enzymatic activity and named cardiac atrial appendage stem cells. As they defend, the major advantage of using an enzymatic reaction for cell detection is that a more homogeneous and viable population can solely be isolated compared with isolations based on the antigen–antibody interaction [66]. Apart from endogenous CSCs, there are new emerging cellbased therapies worth noting. The first of them is the use of induced pluripotent stem cells for cardiac tissue repair. Under certain conditions, these cells can differentiate into functional cardiomyocytes. Moreover, preclinical studies in small and large animal models of myocardial infarction have reported promising results in terms of cell retention, cardiac lineage differentiation and cardiac functionality. The main disadvantage of these novel cell types is the risk of tumor formation, similar to embryonic stem cells. However, the replacement of retro- and lentiviral vectors with nonviral components may prove to be a solution to this problem [38,67]. Another emerging cell-derived therapy is the use of exosomes as these particles have been recently described as the mediators of the paracrine effect of stem cells having an immunomodulatory potential under allogeneic settings [68]. Exosomes present several advantages compared to the use of stem cells as they mitigate many of the safety concerns and limitations associated with the transplantation of viable replicating cells. For example, transferred cells may die or not fully home into the site of damaged tissue, whereas biological factors can be locally administered with a controlled dosage [69]. At present, the therapeutic potential of exosomes derived from MSCs and CDCs has been successfully applied in murine models for the treatment of cardiovascular diseases [70,71]. Immunological considerations for allogeneic cell therapy

The immunogenicity of stem cells is a serious and controversial issue which may directly affect the efficacy of stem cell-based treatment. Although the use of autologous stem cells is widely accepted from an immunological point of view, for a clinical use and to extend the therapeutic use of CSCs to a wide range of patients, the autologous administration of CSCs becomes very difficult [5]. Although new automated and high-throughput cell culture systems are being developed [72], for autologous therapies, growing the patient’s own CSCs is likely to be time Expert Rev. Cardiovasc. Ther.



consuming, expensive and require sophisticated instrumentation, and therefore it would be available to a very limited number of patients. The immunogenicity and immunomodulatory properties of MSCs have been extensively described in the past [73,74] but less information is available regarding CSCs. Since the mechanism of action of these cells is still not fully understood, it would be difficult to fully elucidate the immunogenicity of CSCs in an allogeneic setting. Nowadays, there are different hypotheses concerning the mechanisms through which CSCs could provide a clinical benefit. In the past years, the transdifferentiation hypothesis attempted to explain the beneficial effect of CSC transplantation [75]. However, the in vivo differentiation of adult stem cells into cardiac myocytes has not been demonstrated conclusively and, if it occurs, its frequency would be very low [76]. From an immunological perspective, the transdifferentiation of stem cells into cardiac or vascular cells does have an import on the immunogenicity of the implanted cells. In vitro experiments using MSCs have demonstrated that undifferentiated cells expressed very low levels of MHC class I and class II molecules. After differentiation toward myocytes, smooth muscle or endothelial cells, these cells acquire an increased expression of MHC class Ia and II together with an increased susceptibility to allogeneic cytotoxic T cells [77]. The in vivo experiments using allogeneic MSCs in a rat model have clearly demonstrated that differentiated allogeneic cells are eliminated from the host tissue more rapidly than undifferentiated cells. Finally, it is important to note that very similar results have been found in allogeneic and xenogeneic settings where the transplanted cells were lost from the infarcted rat heart within 5 weeks [77] and 6 weeks [78], respectively. For these reasons, it is now accepted that transdifferentiation cannot be the main mechanism to explain improvements in cardiac function after stem cell therapy. The paracrine hypothesis is gaining adepts. Published evidence demonstrates that stem cells release soluble factors and extracellular vesicles with analogous effects in MSCs [70,79] and CSCs [71]. The transplanted allogeneic CSCs have been proved to release paracrine factors that stimulate endogenous repair mechanisms, and experimental evidences in the infarcted rat model have shown that allogeneic cells survive in the myocardium only for a short period [13]. According to the paper published by Marban’s laboratory in 2012 using mismatched CDCs, this short period is long enough to stimulate regenerative pathways [13]. The administration of rat-derived CDCs in allogeneic recipients has demonstrated that these cells are hypoimmunogenic and did not induce a humoral response. However, it is important to note that, in these in vivo experiments, a cellular immune response was reported and alloreactive lymphocytes exhibited an enhanced proliferation against CSCs. If CSCs truly induce memory T cells for clinical trials, the fact that repeated administrations of these cells may induce a robust secondary immune response should be taken into account, since this will result in a rapid clearance of administered cells. informahealthcare.com

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Looking toward clinical translation, the allogeneic rejection risk of CSCs has been described in a recent paper from Al-Daccak et al. In this paper, the authors elegantly demonstrated that human CPCs shift their signaling capacities within the allogeneic setting toward signals that promote the development, maintenance and functioning of anti-inflammatory response [80]. Moreover, the authors demonstrated that PD-L1 plays a key role to identify low-risk allogeneic cardiac cells. Further studies are still needed to assure that allogeneic CSCs are safe and to explore the host response to this allogeneic therapy, but the evidence available so far is encouraging, and considering the enormous potential of cardiac-derived cellular products, Phase I clinical trials have been started using CDCs (ALLSTAR) and hCPCs (CAREMI). Animal studies

Research in animal models is mandatory to assess the safety and efficacy of any new therapy prior to clinical translation [34,81,82], and animal studies have demonstrated their usefulness in guiding the performance of clinical trials [4]. Small laboratory animals, such as rodents, have been broadly used for cardiology research, and significant insights into cardiovascular biology have been gained from such models. However, these small species are not adequate in some aspects, and many human trials extrapolating rodent data have failed to reproduce the results obtained in the laboratories [83], thus making large animal models essential [34,82,84] for bridging the gap between murine models and human disease. TABLE 1 presents an overview of experimental studies performed in the field. Proof of concept: rodent studies with allogeneic cardiac-derived products

The work performed so far with truly allogeneic cells is strictly limited since most studies in small rodents use either severe combined immunodeficient (SCID) mice and human cells [60,61,63,85–87] or syngeneic cells [18,41,88]. However, and since these studies provide the highly needed proof of concept, we have attempted to include some of them in the present review in a meaningful way. Most works describing a cellular type for the first time have also provided some rodent data to support their regenerative capabilities. Thus, the pioneering work performed by Beltrami et al. [41] describing c-kit+/Lin- cells demonstrated their capability to improve cardiac function in rats, as later corroborated by other works [89]. Similarly, the first study published using CS [60] injects human CS in the peri-infarct area in SCID mice, using echocardiography to assess functional progress and reporting a preserved infarct wall thickness and percent fractional shortening, in the absence of changes to infarct size between treated and control animals. They describe the existence of bands of regenerating myocytes and neovascularization. Further down the road, works from the same laboratory [63] describe the first preclinical use of CDCs in a similar study, also using human cells in SCID mice. In this case, echocardiographic follow-up showed an improvement in global doi: 10.1586/14779072.2015.1011621

Review



Table 1. Animal studies using nonautologous cardiac-derived cell products for myocardial regeneration during the acute/subacute phase of myocardial infarction. Study (year)

Species

Cell type

Remarks

Ref.

Beltrami et al. (2003)

Rat

Lin-, c-kit+

Original description. IM administration at the border zone on day 0 (105 cells)

[41]

Oh et al. (2003)

Mouse

Sca-1+

Original description. IV administration on day 0 (106 cells)

[47]

Messina et al. (2004)

SCID mouse

CS

Original description. IM administration at the border zone on day 0 (10 CS/animal)

[60]

Dawn et al. (2005)

Rat

c-kit+

IC administration on day 0 (106 cells)

[18]

Smith et al. (2007)

SCID mouse

hCDCs

Original description. IM administration at the border zone on day 0 (105 cells)

[63]

Bearzi et al. (2007)

SCID mouse, rat

c-kit+

IM administration at the border zone on day 0 resulting in chimeric hearts

[89]

Rota et al. (2008)

Rat

c-kit+

IM administration at the border zone on day 20 (40,000 cells)

[91]

Takehara et al. (2008)

Immunosuppressed swine

hCDCs + bFGF releasing hydrogel

IM administration at the border zone at 4 weeks (2 107 cells)

[96]

Li et al. (2009)

Mouse

Sca-1+

IM administration at the border zone on day 0 (5 105 cells)

[88]

Cheng et al. (2010)

Rat

CDCs


[103]

Chimenti et al. (2010)

SCID mouse

hCDCs and hCS

IM administration at the border zone on day 0 (105 cells)

[61]

6

Davis et al. (2010)

Rat

CDCs and outgrowth cells

IM administration at the border zone on day 0 (10 cells)

[90]

Carr et al. (2011)

Rat

CDCs

IM administration at the border zone on day 0 (2 106 cells) followed by IV on day 2 (4 106 cells)

[92]

Malliaras et al. (2012)

Rat

CDCs


[13]

Oskouei et al. (2012)

SCID mouse

Adult and fetal ckit+ and BM-MSCs

IM administration on day 0 (36,000 CSCs, 36,000 and 1 106 BM-MSCs)

[87]

Li et al. (2012)

SCID mouse

Rat and human CDCs, BM-MSCs, AD-MSCs and BMMNCs

IM administration at the border zone on day 0 (5 105 all cell types except 106 BM-MNCs)

[85]

Malliaras et al. (2013)

Swine

CDCs

IC administration 3 weeks after infarction (12.5 106 cells)

[12]

Williams et al. (2013)

Swine

hCSCs, hMSCs and combination hCSCs/hMSCs

IM administration at the border zone on day 14 (1 106 hCSCs, 200 106 hMSCs)

Tseliu et al. (2013)

Rat

CS

IM administration at the border zone on day 0 (40,000 CS)

[17]

Cheng et al. (2014)

SCID mouse

hCDCs from healthy, infarcted and advanced heart failure patients

IM administration at the border zone on day 0 (100,000 hCDCs)

[26]

Yee et al. (2014)

Swine

CDCs and CS

Transendocardial NOGA-guided injection on day 28 (escalating doses from 5 to 100–200 CS)

[31]

[102]

Meeting abstracts have been excluded. AD-MSCs: Adipose-derived mesenchymal stem cells; BM-MNC: Bone marrow-derived mononuclear cells; bFGF: Basic fibroblast growth factor; BM-MSCs: Bone marrowderived mesenchymal stem cells; CS: Cardiosphere; CSCs: Cardiac stem cells; hCDCs: Human cardiosphere-derived cells; IC: Intracoronary; IM: Intramyocardial; IV: Intravenous; SCID: Severe combined immunodeficiency.

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Expert Rev. Cardiovasc. Ther.



left ventricular ejection fraction (LVEF) in CDC-treated animals versus controls, and while no differences in infarct size between groups were seen either, there were regions of viable myocardiocytes (stained red with Masson’s Trichrome) within the infarct area in the treated hearts but not in the control animals. In an attempt to decrease the time needed for cell culture, cells directly grown from cardiac explants (outgrowth cells), that represent an intermediate step in the generation of CS, were compared to CDCs [90]. After a thorough characterization including in vitro differentiation, secretome and phenotyping, infarcted rats were injected with CDCs, outgrowth cells, fibroblasts or phosphate-buffered saline, and followed up using MRI, which is the gold standard for functional cardiac evaluation. These cells possessed genetic and surface markers of CPCs, were able to differentiate into a cardiac phenotype, secreted cardioprotective and angiogenic cytokines (including VEGF and IGF-1, but not HGF or PDGF) and could influence cardiac function, with LVEF 3 weeks after infarction being significantly higher in CDC- or outgrowth cells-injected animals compared to the phosphatebuffered saline- or fibroblast-injected groups. Moreover, a trend toward decreased infarct size compared to controls was also evidenced in treated rats. Since no clear difference was seen between the two treatment groups, outgrowth cells may represent an alternative requiring less time-consuming processing prior to administration. Studies performed in small animals have also been used to determine the capabilities of cardiac-derived cellular products prior to advancing to more clinically relevant species. For example, the ability of CSCs to cross the vessel wall and reconstitute all cardiac tissues when delivered in a clinically relevant manner was demonstrated by Dawn et al. [18]. In this work, c-kit+/Lin- CSCs expressing enhanced green fluorescent protein (EGFP) were administered in infarcted rats via the coronary tree, and echocardiographic follow-up revealed a preservation of cardiac anatomy and function in CSC-treated animals compared to controls, where a clear deterioration of functional parameters was seen. EGFP+ cells were found in clusters of small cells of an immature, neonatal appearance within the infarcted myocardium, whereas in noninfarcted areas they were dispersed and of a much greater size (and mature phenotype). Independently of the different degree of maturation, which is a concern that still needs to be addressed in the field, this work proved that CSCs administered into the coronary tree could home to the myocardium. Another interesting study performed with the same type of cells [91] looked into the differential effect of the activation of endogenous CPCs with growth factors (GFs) or the direct implantation of CPCs, finding that both therapies had a salubrious effect on the scar, with the infarcted wall being significantly (p > 0.001) thicker and showing both scar and viable myocytes in the treated groups compared to controls. They conclude that both treatments are able to salvage up to 45% of the infarcted tissue and replace it with functional myocardium. An interesting part of this study is that they also evaluated the ability of the injected or activated CPCs to migrate to the scar, informahealthcare.com

Review

demonstrating that the cells migrate via the expression of different MMPs, that have a marked catalytic effect easing the migration of CPCs from the border toward the center of the scar. Moreover, rodent studies are pivotal in discriminating the relative roles of different strategies or mechanisms of action, as proven by Chimenti et al. [61]. This group injected human CDCs in the peri-infarct area of SCID mice after permanent coronary ligation to assess the direct and paracrine contributions of cardiac-derived cell therapy to the regeneration obtained after administration in an infarcted heart and quantify the relative contributions of each mechanism to the beneficial effects observed after therapy. They demonstrated that human CS and CDCs have the ability to secrete IGF-1, HGF and VEGF in vitro and in vivo. Since the cells are implanted in a mice model, and the secreted GFs come from human cells, they were able to prove the existence of in vivo secretion by finding the factors secreted by human cells at 1 and 3 weeks after administration. In terms of engraftment, they showed that at 1 week only about 30% of the signal intensity from luciferase-labeled CDCs remained and no cells could be detected by this nonquantitative method at 3 weeks. To calculate the relative contribution of paracrine and direct action of CDCs, they identified the new cells at both vascular structures and the myocardium, reporting that only about 20% of the new capillaries were human in origin. Since the paracrine effects can be exerted both on the myocardium and on endogenous stem cells, they studied the possible activation of endogenous CSCs by the injected hCDCs, finding an increase in endogenous c-kit positive cells around the human CDCs. This thorough study concludes that the mechanism of regeneration is threefold (direct differentiation, indirect paracrine effect on the myocardium and indirect paracrine effects on the endogenous CSCs), but the major contributors are represented by paracrine effects, in their two dimensions. A recent study aimed to explore repeated administration of CDCs. Considering that the inflammatory milieu of the heart immediately after reperfusion could be detrimental for cell engraftment, Carr et al. [92] administered CDCs labeled with GFP and micron-sized particles of iron oxide intramyocardially immediately after reperfusion and systemically via the tail vein at 2 days after the infarction, and the evolution of cardiac function demonstrated improved functionality in CDC-treated animals. An interesting finding is that cells positive for GFP and micron-sized particles of iron oxide were found in these animals at 16 weeks after injection, thus pointing to greatly improved cell retention and survival with this administration regimen. To assess the feasibility and safety of an allogeneic therapy, Malliaras et al. [13] aimed to establish the safety of allogeneic CDC administration in the absence of immunosuppression. Considering the evidence pointing toward a mainly paracrine effect, these authors consider that rejection may not be an issue for this therapy, since if the cells are cleared after exerting their beneficial paracrine effects their disappearance would not doi: 10.1586/14779072.2015.1011621


Review


present a problem. However, the potential limitations of allogeneic therapy are not only related to their effectiveness but also to safety concerns. Therefore, they characterized the in vitro immunological properties of CDCs, monitoring the signs of immunorejection (such as inflammatory cytokine secretion, leukocyte infiltration and development of cellular or humoral memory response) and the survival of the cells as well as their functional effect. They injected CDCs of syngeneic, allogeneic and xenogeneic origin, as well as a control group, in the periinfarct area in rats, quantifying the levels of circulating inflammatory cytokines to assess immunogenicity of the cells as well as performing a post-mortem pathological evaluation of the immune response. The baseline immunophenotype of the studied cells suggests that they may be useful as an allogeneic product: expression of MHC class I antigens confers the cells with protection against NK cell-mediated deletion, while the absence of MHC class II antigens allows CDCs to escape direct recognition from CD4+ T helper cells. They found that during the first 3 weeks after injection, the clearance of syngeneic cells was slower than allogeneic cells, while cells of xenogeneic origin were rejected within 1 week. However, allogeneic and syngeneic transplantations exerted comparable and sustained beneficial effects on infarcted heart structure and function as evidenced by decreased infarct size, increased infarcted wall thickness and limited remodeling when compared to xenogeneic and control animals. These benefits persisted over time, up to 6 months post treatment, which supports the paracrine activation of endogenous cells as the mechanism of action. In terms of immunogenicity, there was a slight transient lymphohistiocytic infiltration at 3 weeks in the allogeneic group that was much lower than that observed with xenotransplantation and that had completely subsided at 6 months. To further explore the effect of allotransplantation, researchers from the same laboratory [17] performed intramyocardial injection of CS (not CDCs) in infarcted rats. First, they completed a characterization of the immunological profile of the CS, and, as happened with CDCs, showed that the outer cell layers expressed intermediate levels of MHC I in their surface, but no detectable MHC II. Interestingly, effects were very similar between allogeneic and syngeneic transplantation, with improvements in LVEF and ventricular volumes present as soon as 1 week and sustained for up to 6 months. Cell engraftment was low, as expected, with no cells being detected in the allogeneic transplantation group after 1 week. Despite this low engraftment, benefits were sustained over time. Pathology showed no immune rejection signs in syngeneic or allogeneic groups, but a robust cell-mediated response was seen in the xenogeneic group. Curiously, inflammatory cytokines at day 7 were lower in the syngeneic and allogeneic groups (when compared with the control group), which points to an immunomodulatory action, consistent with reports from other groups [80] that could be responsible, at least in part, for the sustained effect seen over time. Considering the multitude of cell types currently under evaluation for cardiac therapy, Li et al. [85] performed a direct head-to-head comparison of various stem cell types of human doi: 10.1586/14779072.2015.1011621

origin, namely, CDCs, bone marrow-derived MSCs, adiposetissue-derived MSCs and bone marrow-derived mononuclear cells. These authors showed that cardiac-derived cells showed the greatest in vitro potential for myogenic differentiation and angiogenesis induction, along with a balanced and relatively high production of angiogenic and antiapoptotic factors. Similarly, cardiac-derived cells provided the greatest functional benefit in an SCID mice model of experimental infarction, enhancing cell engraftment and myogenic differentiation rates with the lowest number of apoptotic cells and the best preserved/repaired heart architecture. In a further study, they compared the effects of CDCs with that of a subpopulation of c-kit+ cells purified from CDCs and reported that CDCs performed better than c-kit+ in terms of functional benefit, which could be attributed to the fact that CDCs secreted higher amounts of paracrine factors than c-kit+ cells, which could account for the better results with these cells. In other direct comparison, Oskouei et al. [87] studied the relative therapeutic and biological efficacy of human c-kit+ CSCs of fetal and adult origin (36,000 CSCs of each kind per animal, respectively) versus two doses of bone marrow-derived MSCs (36,000 and 1 106 cells) in infarcted SCID mice. Eight weeks after administration, they reported a greater effect of fetal CSCs on preserving ventricular volumes and LVEF and decreasing infarct size. While adults CSCs performed on a level with the high-dose MSCs administered, the amount of cells needed for a similar effect was 30-times greater, which points to a greater potency of these cells. In terms of engraftment, as identified using human-specific DNA probes, CSC-treated hearts showed significantly higher incorporation of human stem cells, which was much lower in the high-dose MSC group and almost negligible in the low-dose MSCs (they report the appearance of only a few stained cells). Moreover, evidence of myocyte differentiation was seen in CSC-treated animals but not in MSC-treated animals with either dose. Their results indicate a greater potential of human CSCs compared to MSCs. However, not all rodent studies have reported good results. In an independent study, Li et al. [88] used a transgenic mouse strain expressing firefly luciferase and EGFP as CSCs donor and followed up the fate of the cells after intramyocardial administration in an infarction model using bioluminescence imaging. Two days after injection, a robust signal was detected. However, this signal decreased dramatically over time, to 6.7% of baseline at day 7, and to just 0.4% after 8 weeks, indicating poor cell retention. Along these lines, cardiac function, as monitored by multiple techniques (they used echocardiography, MRI, pressure–volume loops analysis, PET scan and pathological determination of the infarct size) deteriorated similarly in treated and control animals, while it remained stable in sham mice. These results, strikingly different from those reported by other groups with cardiac-derived cell products, could be attributed to differences in isolation techniques, donor cell sources, animal model (reperfused vs nonreperfused), etc. Similarly, van Berlo et al. [93] have recently reported that the contribution of Expert Rev. Cardiovasc. Ther.



c-kit+ cells to the heart’s cardiomyocytes is minimal, on the order of 0.03% or less, while they mostly gave raise to cardiac endothelial cells. For this study, they used a mouse model in which they used the Kit locus for tracing the cells and assess the frequency with which c-kit+ cells generate cardiomyocytes in vivo. This work, however, refers to the in situ c-kit+ cell performance, which may not reflect the regenerative potency of ex vivo proliferated cells. Along these lines, another group [94] has reported that these cells are necessary and sufficient for cardiac regeneration. After ablating the proliferating endogenous CSCs and their progeny with 5-fluorouracil, no regeneration or functional recovery was forthcoming in an isoproterenolinduced damage in the murine model, while those animals that had only received isoproterenol, but not the 5-fluorouracil, slowly recovered cardiac function over 28 days, as expected from this model. Taken together, these conflicting results advise caution with the use of exogenous cardiac-derived cells in clinical trials. The amount of mechanistic studies available to elucidate the mechanisms by which this therapeutics could improve cardiac function is still scarce. Large animal studies

As previously noted, rodent studies fail short on their predictive capability, and clinical trials should not proceed relying only on results obtained from a heart that is three orders of magnitude smaller than the human heart, and that therefore cannot answer pivotal questions such as doses, delivery method, etc. [34]. On the other hand, a meta-analysis of 52 studies using large animal models of ischemic heart disease to evaluate the human relevance of such studies of stem cell therapy for cardiac disease confirmed that large animal experiments may predict the outcomes of a clinical trial, but of course some caution is to be expected when interpreting data obtained during a limited follow-up time in otherwise healthy animals with a view to translate the works to aged and generally ill patients. Despite these limitations, the validity of large animal models was established as well as the fact that cardiac cell therapy is safe and leads to an improvement in cardiac function. The use of clinically compatible methods and techniques that can be carried out in large species ensures comparability with other works and a rapid translation of preclinical studies [4]. In a visionary paper published in 2005, Linke et al. [95] reported the existence of c-kit, MDR1 or Sca-1 positive, Lincells in canine hearts. Subsequently, they proved that the periinfarction injection of HGF and IGF-1 after permanent left anterior descending coronary artery occlusion resulted in an 11-fold increase in the number of progenitors per square centimeter within the infarct core and in a 16-fold increase in the border zone. In terms of regeneration, they identified new myocytes well in excess of the lost amount of cells, but these newly formed cells were markedly small, thus reconstituting only 17% of the dead myocyte mass. The advantage of GFs has been explored in other works. Of note is the study carried out by Takehara et al. [96], who assessed for the first time a informahealthcare.com

Review

hybrid approach with very good results, that have led to a clinical trial of combination therapy (ALCADIA, [97]) [98]. They explore a xenogeneic approach, since the ALCADIA trial is designed for autologous therapy, and thus have immunosuppression given to all animals through the course of the studies. They used controlled delivery of bFGF to enhance CDC engraftment in infarcted pigs, reporting a marked improvement in cardiac function and hCDC engraftment (as shown by superparamagnetic iron oxide labeling and MR tracking [99]). They also evaluated arteriolar density using a smooth muscle positive immunohistochemistry and described functionally stable microvascular networks that support engraftment and differentiation. In the available literature, truly allogeneic studies in large animal models are mostly reported only in the abstract form so far, since the possibility of using allogeneic cardiac-derived cellular products as therapeutics in themselves is relatively recent. To date, allogeneic CSCs have been tested in swine 8 weeks [31], 2–3 weeks [12], 1 week [28] and immediately [27,29,30,100] after infarction. Among those, Marban’s laboratory has reported extensive experience with this therapy, both in small [13,17] and large models [12,27,30,31] and using different administration routes to take advantage of their two cellular products, CS and CDCs. Since the size of CS precludes their administration via the coronary artery, they have been delivered intramyocardially [101] or transendocardially [31]. In a prior work with autologous cells, Lee et al. [101] compared the intramyocardial injection of CDCs and CS in swine subjected to a 2.5 h occlusion of the mid LAD. They report an interesting superiority of CS in terms of improving hemodynamics and regional function and in attenuating ventricular remodeling. They found that an injection of 0.5 106 cells per injection site resulted in the greatest cell retention at 24 h, compared with higher doses. Although cell treatment did not increase LVEF relative to baseline, the deterioration of this parameter in the control group meant that the final LVEF was significantly better in cell-treated groups. This preservation of global LVEF in cell-treated animals compared to placebo was not observed with intracoronary administration of CDCs in the same experimental model [11]. Thus, intramyocardial injection may represent a more effective route of administration than the intracoronary route. Yee et al. [31] administered allo-CS at 15 different sites around the scar in pigs with the NOGA system, which identifies viable myocardium and scar and allows targeting to the border zone of the infarction. After 8 weeks, they report no deaths, no immunorejection seen with either histology or donor-specific antibodies as well as a reduced scar size, increased viable mass and preserved cardiac morphology in CS-injected pigs compared to placebo. Intramyocardial delivery via thoracoscopy has also been described successfully [102], representing a clinically translatable minimally invasive option for cell delivery. However, the intracoronary route is, from a clinical point of view, the most practical method for cell delivery to the heart, as it is widely available worldwide, less invasive than other administration routes and the cells can be administered to the entire doi: 10.1586/14779072.2015.1011621


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myocardium at risk [10,16]. Kanazawa et al. injected different doses of CDCs immediately after reperfusion in the swine LAD occlusion model, concluding that the optimum dose for these studies is 7.5 106 CDCs [30], and subsequently reporting a cardioprotective effect related to CDC administration, with a significant decrease in the area at risk evaluated 48 h after administration [27]. When performing the administration 2–3 weeks after the ischemic event, rather than immediately, this group has described a higher dose of 12.5 106 allo-CDCs [12], finding only a slight, marginally significant (compared to placebo), focal lymphoplasmacytic infiltration corresponding to grade 1R of the International Society for Heart and Lung Transplantation in the absence of foci of myocyte damage or circulating antidonor antibodies. In terms of efficacy, MRI demonstrated improved function (evidenced by the preservation of LVEF in treated animals), limited remodeling (as seen by the evolution of ventricular volumes), decreased infarct size and increased viable myocardium when compared to placebo at 2 months post intervention (scar size does not change in controls and decreases by 3.6% in treated animals, along with increased viable myocardial mass). Regarding engraftment, 4.3 ± 2.2% of injected allo-CDCs were still present at 24 h after administration, but no evidence of the cells could be found at 2 months. Their results are comparable to those of autologous therapy with the same cells [11], but with a greater import on LVEF that they consider could be attributed to an earlier administration, a benefit that, if consistently reported in other studies, would be inherent to allogeneic therapy. Another study injected allogeneic, nonmatched, cloned male EGFP-transduced porcine c-kit+ CSCs intracoronary in infarcted pigs [100]. While >95% of injected CSCs were found in the damaged pig myocardium at 30 min through to 1 day, no evidence of the injected cell could be found at 3 weeks after infarction. There was, however, a significant increase in the number of autologous c-kit+ CSCs in the border and infarct regions, which determined a preservation of the myocardial architecture, thus pointing to an improvement in myocardial cell survival and physiologically meaningful regeneration brought about by the activation of the endogenous CSC compartment. Our group has recently reported the experience with intracoronary administration of allogeneic porcine CSCs either at different timepoints (immediately or after a week) [29] or with different doses (25 106 or 50 106 CSCs) [28] in a clinically relevant swine model of reperfused myocardial infarction. These preliminary studies confirmed the safety of this approach regardless of the moment of administration or the dose, suggesting a trend toward a better performance of pCSCs when administered 1 week after the ischemic insult. While cardiac-derived cells appear to be more potent than other cell types studied so far, attempts have been made to enhance their therapeutic efficacy, including combination therapy with other cell types [102], concurrent administration with GFs [96] or magnetic targeting [103]. Recently, Williams et al. [102] reported the enhanced cardioregenerative potential of the combination of human c-kit+ cells with bone marrow-derived doi: 10.1586/14779072.2015.1011621

MSCs compared to each cell type administered alone in a porcine myocardial infarction model. Since MSCs can regulate CSCs niches, possess immunomodulatory properties and secrete numerous growth factors, the authors hypothesize that cell-tocell interactions could be exploited for therapeutic purposes and that the combination of both cell types would increase therapeutic efficacy. Moreover, not only was cardiac function improved twofold in the combination group compared to each cell alone, but cell engraftment was sevenfold greater 4 weeks after treatment. As an example of GF administration to improve cell retention, the successful use of controlled delivery of bFGF to enhance CDC engraftment in infarcted pigs has been described above [96]. In a different and intriguing approach, Cheng et al. [103] report enhanced engraftment and functional benefit of iron-labeled cells subjected to a 1.3T magnetic field during intramyocardial injection in the infarcted rat heart that was maintained for 10 min thereafter. After ensuring that the use of superparamagnetic microspheres did not affect cell viability, these authors compared the 24 h and 3 weeks retention and effects of labeled CDCs subjected or not to the magnet, reporting that retention and 3 weeks engraftment were increased threefold compared to nontargeted cells, and that it translated to improved cardiac architecture and function at 3 weeks, with a strong correlation between the LVEF and cell retention as measured at this timepoint. Other approaches that could be useful, but, to our knowledge, have not been applied with cardiac-derived cellular products so far, are cell encapsulation [104,105], promotion of cell survival, for example, by the concurrent administration of simvastatin [106] or by activation of Akt, a serine-threonine kinase [107], the use of a vascular adipose flap as cell donor [108], etc. Clinical trials

The different studies have proven that CSCs are present in the heart of mammal species, including man, and can be used to help regenerate the infarcted myocardium in different ways. Human studies have been performed and even completed in some cases, with CSCs from autologous origin, with extremely good results. In the two clinical trials using autologous cells of cardiac origin whose results have been made available to date, SCIPIO investigators reported a 12.3% increase in LVEF at 1 year after treatment with autologous CSCs (c-kit+) in ischemic heart failure [19,20], whereas the patients enrolled in the CADUCEUS trial did not show any changes in this parameter compared to the control group after 1 year, despite demonstrating a 11.1% reduction in scar size (compared to unchanged scar size in control patients) and an increase in viable myocardium by 22.6 g in patients treated with CDCs versus 1.8 g in controls [22,23]. Building on the preclinical experience discussed above, and considering the highly encouraging results of the autologous therapy trials, two clinical trials are underway with allogeneic heart-derived cellular products (ALLSTAR, CAREMI) for acute myocardial infarction. The first results from an allogeneic cardiac-derived stem cell product trial were advanced in the Expert Rev. Cardiovasc. Ther.



63rd American College of Cardiology Meeting held in March 2014. In this meeting, the preliminary results (with a follow-up of at least 3 months after injection) from the Phase I ALLSTAR trial were reported. This trial aims to determine the safety profile of allogeneic CDCs administered via the infarctrelated coronary artery between 4 weeks and 12 months after infarction. Phase I was an open-label, dose-escalating study, and its results are being highly encouraging, meeting the first safety endpoint at 1 month after infusion: there were no acute myocarditis attributable to the infusion, no deaths due to ventricular tachycardia or fibrillation, no sudden deaths and no major adverse cardiac events. Phase II has been approved by the data safety monitoring board and enrollment is reported to be ongoing for the double-blind, randomized, placebocontrolled Phase II [109]. On the other hand, the CAREMI trial has recently started its dose-escalating phase. Considering the preclinical and in vitro results reported by this Consortium [28,29,80], the results from this trial will be awaited eagerly. Expert commentary

The very fact that there are numerous stem cells currently under investigation for cardiac regeneration shows that the optimal ancillary cell therapy for the treatment of myocardial infarction has not been found yet. Cardiac-derived cellular products appear to be highly promising in both experimental and clinical settings. Results with autologous cells using these therapeutics are, so far, the most promising results obtained from human trials with cell therapy for heart failure. If we could harness the potential of allogeneic cell therapy using these cells, which appears feasible so far, we may be witness to a turning point in the history of postinfarction therapy. It has been proven that the administration of CSCs puts in motion a paracrine system that activates survival pathways on the cells at risk and activates the endogenous stem cell compartment [110]. Once these events are triggered, the presence of the cells is not

Review

needed to maintain the benefit, and once the allogeneic cells are cleared we would be obtaining an autologous tissue from an exogenous product, a definitely promising and intriguing possibility. The field is, however, still in its infancy, and there are several questions that need to be addressed in experimental studies, and that could have different answers with the different cell products available, such as administration time and route, dose, possibility of allosensitization, utility of hybrid or combination therapies, etc. Five-year view

As discussed earlier, the possibilities inherent to allogeneic CSC administration are huge, and the answer from the clinical trials already started or planned will definitely tell if we see the beginning of the much-heralded cardiac regenerative therapy. This will entail the use of generic off-the-shelf therapies which will be ready to be applied at any time wherever there are the technical means and professional expertise needed to treat acute myocardial infarctions, as is the case for most large medical centers. Moreover, and because of their generic nature, these therapies will be available to the majority of patients. The possibilities are staggering, but a word of caution will be necessary, and a much needed return to the bench-side will help us determine and fully understand the mechanisms of action, allogeneic risks and to define the particularities of the therapy. Financial & competing interests disclosure

This paper has been funded via the European FP7-HEALTH-2009-1.4-3, Grant Agreement 242038. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties. No writing assistance was utilized in the production of this manuscript.

Key issues .

Cell-based therapies aiming at myocardial regeneration have the potential to transform the clinical management and prognosis of myocardial infarction and heart failure.

.

Highly promising results have recently been reported from the first clinical trials to use autologous cardiac cells in the setting of heart failure, either after coronary artery bypass grafting [19,20] (SCIPIO trial, [21]) or after coronary stenting [22,23] (CADUCEUS trial, [24]).

.

Allogeneic cells emerge as a promising option and several studies using this strategy have been reported [13,27–30]. The advantages of using an allogeneic product are varied. They offer the possibility of administering the cells very early after the ischemic event, as an ‘offthe-shelf’ product that, at the same time, can be subjected to higher quality controls than autologous products.

.

There are several heart-derived cell populations described to date, expressing different characteristics and/or surface markers: c-kit+, Sca-1+, side population cells, Isl1+, cardiospheres and cardiosphere-derived cells.

.

Heart-derived products are suspected to act via a paracrine activation of endogenous cardiac stem cell immunomodulation.

.

Rodent studies with different heart-derived cell types have confirmed their safety and potential (data).

.

Swine studies support these results and help in defining dose, administration route and similar questions.

.

Early clinical trials have been started. Preliminary safety results are available in the ALLSTAR trial, and Phase II has been started in view of the said results.


doi: 10.1586/14779072.2015.1011621

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cells in porcine ischemic cardiomyopathy. Circulation 2009;120:1075-83

References


Papers of special note have been highlighted as: . of interest .. of considerable interest 1.

Nichols M, Townsend N, Scarborough P, Rayner M. Cardiovascular disease in Europe: epidemiological update. Eur Heart J 2013;34:3028-34

2.

Go AS, Mozaffarian D, Roger VL, et al. Heart disease and stroke statistics— 2014 update: a report from the american heart association. Circulation 2014;129: e28-e292

3.

4.

5.

6.

7.

8.

9.

10.

11.

Dimmeler S, Burchfield J, Zeiher AM. Cell-based therapy of myocardial infarction. Arterioscler Thromb Vasc Biol 2008;28: 208-16 van der Spoel TIG, Jansen of Lorkeers SJ, Agostoni P, et al. Human relevance of pre-clinical studies in stem cell therapy: systematic review and meta-analysis of large animal models of ischaemic heart disease. Cardiovasc Res 2011;91:649-58 Sanganalmath SK, Bolli R. Cell therapy for heart failure: a comprehensive overview of experimental and clinical studies, current challenges, and future directions. Circ Res 2013;113:810-34 Hare JM, Fishman JE, Gerstenblith G, et al. Comparison of allogeneic vs autologous bone marrow–derived mesenchymal stem cells delivered by transendocardial injection in patients with ischemic cardiomyopathy: the poseidon randomized trial. J Am Med Assoc 2012;308:2369-79 Schuleri KH, Feigenbaum GS, Centola M, et al. Autologous mesenchymal stem cells produce reverse remodelling in chronic ischaemic cardiomyopathy. Eur Heart J 2009;30:2722-32 Koudstaal S, Jansen of Lorkeers SJ, Gaetani R, et al. Concise review: heart regeneration and the role of cardiac stem cells. Stem Cells Trans Med 2013;2:434-43 Gnecchi M, Zhang Z, Ni A, Dzau VJ. Paracrine mechanisms in adult stem cell signaling and therapy. Circ Res 2008;103: 1204-19 Ellison GM, Torella D, Dellegrottaglie S, et al. Endogenous cardiac stem cell activation by insulin-like growth factor-1/ hepatocyte growth factor intracoronary injection fosters survival and regeneration of the infarcted pig heart. J Am Coll Cardiol 2011;58:977-86 Johnston PV, Sasano T, Mills K, et al. Engraftment, differentiation, and functional benefits of autologous cardiosphere-derived

doi: 10.1586/14779072.2015.1011621

12.

..

Malliaras K, Smith RR, Kanazawa H, et al. Validation of contrast-enhanced magnetic resonance imaging to monitor regenerative efficacy after cell therapy in a porcine model of convalescent myocardial infarction. Circulation 2013;128:2764-75 The first full paper reporting the use of allogeneic cardiosphere-derived cells (CDCs) in a clinically relevant animal model (swine reperfused infarct) with clinically compatible delivery (intracoronary) and follow-up (MRI) methods.

13.

Malliaras K, Li TS, Luthringer D, et al. Safety and efficacy of allogeneic cell therapy in infarcted rats transplanted with mismatched cardiosphere-derived cells. Circulation 2012;125:100-12

14.

Welt FGP, Gallegos R, Connell J, et al. Effect of cardiac stem cells on left-ventricular remodeling in a canine model of chronic myocardial infarction. Circ Heart Fail 2013;6:99-106

15.

16.

17.

18.

Li R-K, Mickle DAG, Weisel RD, et al. Optimal time for cardiomyocyte transplantation to maximize myocardial function after left ventricular injury. Ann Thorac Surg 2001;72:1957-63 Bolli R, Tang X-L, Sanganalmath SK, et al. Intracoronary delivery of autologous cardiac stem cells improves cardiac function in a porcine model of chronic ischemic cardiomyopathy. Circulation 2013;128: 122-31 Tseliou E, Pollan S, Malliaras K, et al. Allogeneic cardiospheres safely boost cardiac function and attenuate adverse remodeling after myocardial infarction in immunologically mismatched rat strains. J Am Coll Cardiol 2013;61:1108-19 Dawn B, Stein AB, Urbanek K, et al. Cardiac stem cells delivered intravascularly traverse the vessel barrier, regenerate infarcted myocardium, and improve cardiac function. Proc Natl Acad Sci USA 2005;102:3766-71

19.

Bolli R, Chugh AR, D’Amario D, et al. Cardiac stem cells in patients with ischaemic cardiomyopathy (SCIPIO): initial results of a randomised phase 1 trial. Lancet 2011;378:1847-57

.

The first report of the Phase I SCIPIO clinical trial results using autologous c-kit+ cardiac stem cells in cardiac patients.

20.

Chugh AR, Beache GM, Loughran JH, et al. Administration of cardiac stem cells in patients with ischemic cardiomyopathy: the

SCIPIO Trial: surgical aspects and interim analysis of myocardial function and viability by magnetic resonance. Circulation 2012;126:S54-64 .

Updated and expanded information on the results obtained with the administration of autologous c-kit+ cardiac stem cells during the Phase I SCIPIO clinical trial.

21.

Cardiac stem cell infusion in patients with ischemic cardiomyopathy (SCIPIO). Available from: https://clinicaltrials.gov/ct2/ show/NCT00474461

22.

Malliaras K, Makkar RR, Smith RR, et al. Intracoronary cardiosphere-derived cells after myocardial infarction: evidence for therapeutic regeneration in the final 1-year results of the CADUCEUS trial. J Am Coll Cardiol 2014;63:110-22

.

The final results obtained 1 year after autologous CDC administration in the CADUCEUS trial.

23.

Makkar RR, Smith RR, Cheng K, et al. Intracoronary cardiosphere-derived cells for heart regeneration after myocardial infarction (CADUCEUS): a prospective, randomised phase 1 trial. Lancet 2012;379: 895-904

.

The first results obtained in the CADUCEUS trial using autologous CDCs.

24.

CArdiosphere-Derived aUtologous Stem CElls to Reverse ventricUlar dySfunction (CADUCEUS). Available from: https:// clinicaltrials.gov/ct2/show/NCT00893360

25.

Dimmeler S, Leri A. Aging and disease as modifiers of efficacy of cell therapy. Circ Res 2008;102:1319-30

26.

Cheng K, Malliaras K, Smith RR, et al. Human cardiosphere-derived cells from advanced heart failure patients exhibit augmented functional potency in myocardial repair. JACC Heart Fail 2014;2:49-61

27.

Kanazawa H, Malliaras K, Yee K, et al. Cardioprotective effect of allogeneic cardiosphere-derived cells: reduction of infarct size and attenuation of no-reflow when administered in the infarct-related artery after reperfusion in pigs with acute myocardial infarction. J Am Coll Cardiol 2013;61(10_S):doi:10.1016/ S0735-1097(13)61818-5

28.

Baez-Diaz C, Crisostomo V, Maestre J, et al. Safety and efficacy assesment of intracoronary delivery of porcine cardiac stem cells in a swine model of acute myocardial infarction. comparison of two different cell doses. J Am Coll Cardiol



2014;63(12_S):doi: 10.1016/ S0735-1097(14)61763-0 29.


30.

31.

32.

33.

34.

Crisostomo V, Baez-Diaz C, Maestre J, et al. Allogeneic cardiac stem cell administration for acute myocardial infarction. a timing experimental study in swine. J Am Coll Cardiol 2014; 63(12_S):doi: 10.1016/S0735-1097(14) 61756-3

39.

Kanazawa H, Malliaras K, Yee K, et al. Allogeneic cardiosphere-derived cells after reperfusion are effective in reducing infarct size and attenuatting adverse remodelling in pigs with acute myocardial infarction. J Am Coll Cardiol 2013;61(10_S):doi:10.1016/ S0735-1097(13)61833-1

40.

Yee K, Malliaras K, Kanazawa H, et al. Allogeneic cardiospheres delivered via percutaneous transendocardial injection increase viable myocardium, decrease scar size, and attenuate cardiac dilatation in porcine ischemic cardiomyopathy. PLoS One 2014;9:e113805 Houtgraaf JH, de Jong R, Kazemi K, et al. Intracoronary infusion of allogeneic mesenchymal precursor cells directly after experimental acute myocardial infarction reduces infarct size, abrogates adverse remodeling, and improves cardiac function. Circ Res 2013;113:153-66 Kubal C, Sheth K, Nadal-Ginard B, Galinãnes M. Bone marrow cells have a potent anti-ischemic effect against myocardial cell death in humans. J Thorac Cardiovasc Surg 2006;132:1112-18 Nadal-Ginard B, Torella D, Ellison G. Cardiovascular regenerative medicine at the crossroads. clinical trials of cellular therapy must now be based on reliable experimental data from animals with characteristics similar to human’s. Rev Esp Cardiol (Engl Ed) 2006;59:1175-89

35.

Kreke M, Smith RR, Marban L, Marban E. Cardiospheres and cardiosphere-derived cells as therapeutic agents following myocardial infarction. Expert Rev Cardiovasc Ther 2012;10:1185-94

36.

Hong K, Bolli R. Cardiac stem cell therapy for cardiac repair. Curr Treat Options Cardio Med 2014;16:1-19

37.

38.

therapy for cardiac disease - a review. Folia Histochem Cytobiol 2011;49:13-25

41.

Sanz-Ruiz R, Arranz AV, Iban˜es EG, et al. Randomized clinical trials in stem cell therapy for the heart - old and new types of cells for cardiovascular repair. In: Gholamrezanezhad A, editor. Stem cells in clinics and research. ISBN: 978-953-307797-0 InTech, Rijeka, Croatia; 2011 Barile L, Chimenti I, Gaetani R, et al. Cardiac stem cells: isolation, expansion and experimental use for myocardial regeneration. Nat Clin Pract Cardiovasc Med 2007;4(Suppl 1):S9-S14 Beltrami AP, Barlucchi L, Torella D, et al. Adult cardiac stem cells are multipotent and support myocardial regeneration. Cell 2003;114:763-76

..

The original description of c-kit+ Lincardiac stem cells (CSCs).

42.

Yarden Y, Kuang WJ, Yang-Feng T, et al. Human proto-oncogene c-kit: a new cell surface receptor tyrosine kinase for an unidentified ligand. EMBO J 1987;6: 3341-51

43.

Edling CE, Hallberg B. c-Kit – a hematopoietic cell essential receptor tyrosine kinase. Int J Biochem Cell Biol 2007;39: 1995-8

44.

Leri A. Human cardiac stem cells: the heart of a truth. Circulation 2009;120:2515-18

45.

Bearzi C, Leri A, Lo Monaco F, et al. Identification of a coronary vascular progenitor cell in the human heart. Proc Natl Acad Sci USA 2009;106:15885-90

46.

47.

Gambini E, Pompilio G, Biondi A, et al. C-kit+ cardiac progenitors exhibit mesenchymal markers and preferential cardiovascular commitment. Cardiovasc Res 2011;89:362-73 Oh H, Bradfute SB, Gallardo TD, et al. Cardiac progenitor cells from adult myocardium: homing, differentiation, and fusion after infarction. Proc Natl Acad Sci USA 2003;100:12313-18

Review

51.

Goodell MA, Brose K, Paradis G, et al. Isolation and functional properties of murine hematopoietic stem cells that are replicating in vivo. J Exp Med 1996;183: 1797-806

52.

Pfister O, Mouquet F, Jain M, et al. CD31but Not CD31+ cardiac side population cells exhibit functional cardiomyogenic differentiation. Circ Res 2005;97:52-61

.

The original description of cardiac side population cells.

53.

Oh H, Chi X, Bradfute SB, et al. Cardiac muscle plasticity in adult and embryo by heart-derived progenitor cells. Ann N Y Acad Sci 2004;1015:182-9

54.

Oyama T, Nagai T, Wada H, et al. Cardiac side population cells have a potential to migrate and differentiate into cardiomyocytes in vitro and in vivo. J Cell Biol 2007;176:329-41

55.

Goichberg P, Chang J, Liao R, Leri A. Cardiac stem cells: biology and clinical applications. Antioxid Redox Signal 2014; 21(14):2002-17

56.

Tomita Y, Matsumura K, Wakamatsu Y, et al. Cardiac neural crest cells contribute to the dormant multipotent stem cell in the mammalian heart. J Cell Biol 2005;170: 1135-46

57.

Fonoudi H, Yeganeh M, Fattahi F, et al. ISL1 protein transduction promotes cardiomyocyte differentiation from human embryonic stem cells. PLoS One 2013;8: e55577

58.

Barzelay A, Ben-Shoshan J, Entin-Meer M, et al. A potential role for islet-1 in post-natal angiogenesis and vasculogenesis. Thromb Haemost 2010;103:188-97

59.

Barile L, Messina E, Giacomello A, Marbań E. Endogenous Cardiac Stem Cells. Prog Cardiovasc Dis 2007;50:31-48

60.

Messina E, De Angelis L, Frati G, et al. Isolation and expansion of adult cardiac stem cells from human and murine heart. Circ Res 2004;95:911-21

.

The original description of Sca-1+ CSCs.

..

The original description of cardiospheres.

48.

61.

Vanelli A. Isolation, characterization and differentiation potential of cardiac progenitor cells in adult pigs. PhD Thesis Universita` degli Studi di Milano, Milan; 2011. Available from: http://hdl.handle.net/ 2434/169553

van de Rijn M, Heimfeld S, Spangrude GJ, Weissman IL. Mouse hematopoietic stem-cell antigen Sca-1 is a member of the Ly-6 antigen family. Proc Natl Acad Sci USA 1989;86:4634-8

Chimenti I, Smith RR, Li T-S, et al. Relative roles of direct regeneration versus paracrine effects of human cardiosphere-derived cells transplanted into infarcted mice. Circ Res 2010;106:971-80

49.

Matsuura K, Nagai T, Nishigaki N, et al. Adult cardiac Sca-1-positive cells differentiate into beating cardiomyocytes. J Biol Chem 2004;279:11384-91

..

Quantification of direct and paracrine effects after CDC transplantation.

62.

Jezierska-Wozniak K, Mystkowska D, Tutas A, Jurkowski MK. Stem cells as

50.

Gaetani R, Rizzitelli G, Chimenti I, et al. Cardiospheres and tissue engineering for myocardial regeneration: potential for clinical application. J Cell Mol Med 2010;14:1071-7


Holmes C, Stanford WL. Concise review: stem cell antigen-1: expression, function, and enigma. Stem Cell 2007;25:1339-47

doi: 10.1586/14779072.2015.1011621

Review


63.


Smith RR, Barile L, Cho HC, et al. Regenerative potential of cardiosphere-derived cells expanded from percutaneous endomyocardial biopsy specimens. Circulation 2007;115:896-908

64.

Di Felice V, De Luca A, Colorito ML, et al. Cardiac stem cell research: an elephant in the room? Anat Rec (Hoboken) 2009;292:449-54

65.

Di Felice V, Zummo G. Stem cell populations in the heart and the role of Isl1 positive cells. Eur J Histochem 2013;57:e14

66.

perspective. Mayo Clin Proc 2009;84: 876-92 77.

Huang XP, Sun Z, Miyagi Y, et al. Differentiation of allogeneic mesenchymal stem cells induces immunogenicity and limits their long-term benefits for myocardial repair. Circulation 2010;122: 2419-29

89.

Bearzi C, Rota M, Hosoda T, et al. Human cardiac stem cells. Proc Natl Acad Sci USA 2007;104:14068-73

90.

Davis DR, Kizana E, Terrovitis J, et al. Isolation and expansion of functionally-competent cardiac progenitor cells directly from heart biopsies. J Mol Cell Cardiol 2010;49:312-21

78.

Grinnemo KH, Mansson A, Dellgren G, et al. Xenoreactivity and engraftment of human mesenchymal stem cells transplanted into infarcted rat myocardium. J Thorac Cardiovasc Surg 2004;127:1293-300

91.

Rota M, Padin-Iruegas ME, Misao Y, et al. Local activation or implantation of cardiac progenitor cells rescues scarred infarcted myocardium improving cardiac function. Circ Res 2008;103:107-16

Koninckx R, Daniels A, Windmolders S, et al. The cardiac atrial appendage stem cell: a new and promising candidate for myocardial repair. Cardiovasc Res 2013;97: 413-23

79.

Timmers L, Lim SK, Hoefer IE, et al. Human mesenchymal stem cell-conditioned medium improves cardiac function following myocardial infarction. Stem Cell Res 2011;6:206-14

92.

Carr CA, Stuckey DJ, Tan JJ, et al. Cardiosphere-derived cells improve function in the infarcted rat heart for at least 16 weeks – an MRI study. PLoS One 2011;6:e25669

67.

Fuentes T, Kearns-Jonker M. Endogenous cardiac stem cells for the treatment of heart failure. Stem Cells Cloning 2013;6:1-12

80.

93.

68.

Blazquez R, Sanchez-Margallo FM, de la Rosa O, et al. Immunomodulatory potential of human adipose mesenchymal stem cells derived exosomes on in vitro stimulated T cells. Front Immunol 2014;5:556

Lauden L, Boukouaci W, Borlado LR, et al. Allogenicity of human cardiac stem/ progenitor cells orchestrated by programmed death ligand 1. Circ Res 2013;112:451-64

van Berlo JH, Kanisicak O, Maillet M, et al. c-kit+ cells minimally contribute cardiomyocytes to the heart. Nature 2014;509:337-41

..

Immunomodulatory properties of cardiac progenitor cells.

94.

81.

Monnet E, Chachques JC. Animal models of heart failure: what is new? Ann Thorac Surg 2005;79:1445-53

Ellison GM, Vicinanza C, Smith AJ, et al. Adult c-kit(pos) cardiac stem cells are necessary and sufficient for functional cardiac regeneration and repair. Cell 2013;154:827-42

82.

Suzuki Y, Yeung AC, Ikeno F. The pre-clinical animal model in the translational research of interventional cardiology. J Am Coll Cardiol Intv 2009;2: 373-83

95.

83.

Dixon JA, Spinale FG. Large animal models of heart failure. Circ Heart Fail 2009;2: 262-71

Linke A, Mu¨ller P, Nurzynska D, et al. Stem cells in the dog heart are self-renewing, clonogenic, and multipotent and regenerate infarcted myocardium, improving cardiac function. Proc Natl Acad Sci USA 2005;102:8966-71

96.

84.

Suzuki Y, Yeung AC, Ikeno F. The Representative porcine model for human cardiovascular disease. J Biomed Biotechnol 2011;2011:195483

Takehara N, Tsutsumi Y, Tateishi K, et al. Controlled delivery of basic fibroblast growth factor promotes human cardiosphere-derived cell engraftment to enhance cardiac repair for chronic myocardial infarction. J Am Coll Cardiol 2008;52:1858-65

85.

Li T-S, Cheng K, Malliaras K, et al. Direct comparison of different stem cell types and subpopulations reveals superior paracrine potency and myocardial repair efficacy with cardiosphere-derived cells. J Am Coll Cardiol 2012;59:942-53

.

A proof-of-concept study for the hybrid approach to be used in the ALCADIA clinical trial: combination of abFGF releasing hydrogel and CDCs.

97.

86.

Mohsin S, Khan M, Toko H, et al. Human cardiac progenitor cells engineered with Pim-I kinase enhance myocardial repair. J Am Coll Cardiol 2012;60:1278-87

Autologous human cardiac-derived stem cell to treat ischemic cardiomyopathy (ALCADIA). Available from: https:// clinicaltrials.gov/ct2/show/NCT00981006

98.

Takehara N NM, Ogata T, Nakamura T, et al. The ALCADIA (Autologous human cardiac-derived stem cell to treat ischemic cardiomyopathy) trial. late-breaking clinical trial abstracts. Circulation 2012;126: 2776-99

99.

Kraitchman DL, Bulte JW. Imaging of stem cells using MRI. Basic Res Cardiol 2008;103:105-13

69.

Maguire G, Friedman P, McCarthy D, et al. Stem cell released molecules and exosomes in tissue engineering. Procedia Eng 2013;59:270-8

70.

Lai RC, Chen TS, Lim SK. Mesenchymal stem cell exosome: a novel stem cell-based therapy for cardiovascular disease. Regen Med 2011;6:481-92

71.

Ibrahim AG, Cheng K, Marban E. Exosomes as critical agents of cardiac regeneration triggered by cell therapy. Stem Cell Reports 2014;2:606-19

72.

Kami D, Watakabe K, Yamazaki-Inoue M, et al. Large-scale cell production of stem cells for clinical application using the automated cell processing machine. BMC Biotechnol 2013;13:102

73.

Le Blanc K, Ringden O. Immunomodulation by mesenchymal stem cells and clinical experience. J Intern Med 2007;262:509-25

74.

75.

76.

Garcia-Castro J, Trigueros C, Madrenas J, et al. Mesenchymal stem cells and their use as cell replacement therapy and disease modelling tool. J Cell Mol Med 2008;12: 2552-65 Anversa P, Leri A, Kajstura J. Cardiac regeneration. J Am Coll Cardiol 2006;47: 1769-76 Gersh BJ, Simari RD, Behfar A, et al. Cardiac cell repair therapy: a clinical

doi: 10.1586/14779072.2015.1011621

87.

Oskouei BN, Lamirault G, Joseph C, et al. Increased potency of cardiac stem cells compared with bone marrow mesenchymal stem cells in cardiac repair. Stem Cells Transl Med 2012;1:116-24

88.

Li Z, Lee A, Huang M, et al. Imaging survival and function of transplanted cardiac resident stem cells. J Am Coll Cardiol 2009;53:1229-40



100.


101.

102.

Ellison GM, Torella D, Trigueros C, et al. Use of heterologous non-matched Cardiac Stem Cells (CSCs) without immunosuppression as an effective regenerating agent in a porcine model of acute myocardial infarction. Eur Heart J 2009;30:495 Lee S-T, White AJ, Matsushita S, et al. Intramyocardial injection of autologous cardiospheres or cardiosphere-derived cells preserves function and minimizes adverse ventricular remodeling in pigs with heart failure post-myocardial infarction. J Am Coll Cardiol 2011;57:455-65 Williams AR, Hatzistergos KE, Addicott B, et al. Enhanced effect of combining human cardiac stem cells and bone marrow mesenchymal stem cells to reduce infarct size and to restore cardiac function after myocardial infarction. Circulation 2013;127:213-23


103.

104.

105.

106.

Cheng K, Li TS, Malliaras K, et al. Magnetic targeting enhances engraftment and functional benefit of iron-labeled cardiosphere-derived cells in myocardial infarction. Circ Res 2010;106:1570-81 Gomez-Mauricio RG, Acarregui A, Sanchez-Margallo FM, et al. A preliminary approach to the repair of myocardial infarction using adipose tissue-derived stem cells encapsulated in magnetic resonance-labelled alginate microspheres in a porcine model. Eur J Pharm Biopharm 2013;84:29-39 Barnett BP, Kraitchman DL, Lauzon C, et al. Radiopaque alginate microcapsules for X-ray visualization and immunoprotection of cellular therapeutics. Mol Pharm 2006;3: 531-8 Yang Y-J, Qian H-Y, Huang J, et al. Combined therapy with simvastatin and bone marrow–derived mesenchymal stem cells increases benefits in infarcted swine

Review

hearts. Arterioscler Thromb Vasc Biol 2009;29:2076-82 107.

Gnecchi M, He H, Melo LG, et al. Early beneficial effects of bone marrow-derived mesenchymal stem cells overexpressing Akt on cardiac metabolism after myocardial infarction. Stem cells 2009;27:971-9

108.

Galvez-Monton C, Prat-Vidal C, Roura S, et al. Post-infarction scar coverage using a pericardial-derived vascular adipose flap. Pre-clinical results. Int J Cardiol 2013;166: 469-74

109.

Capricor. Available from: http://capricor. com/clinical-trials/allstar/

110.

Ellison G, Nadal-Ginard B, Torella D. Optimizing cardiac repair and regeneration through activation of the endogenous cardiac stem cell compartment. J Cardiovasc Transl Res 2012;5:667-77

doi: 10.1586/14779072.2015.1011621

Delayed administration of allogeneic cardiac stem cell therapy for acute myocardial infarction could ameliorate adverse remodeling: experimental study in swine.

Allogeneic adipose stem cell therapy in acute myocardial infarction.

The math of sisyphus: the conundrum of stem cell administration for myocardial infarction and myocardial failure.

Resveratrol activates endogenous cardiac stem cells and improves myocardial regeneration following acute myocardial infarction.

Acute myocardial infarction after ergonovine administration for uterine bleeding.

Letter: Cardiac pacing in acute myocardial infarction.

Intravenous administration of atorvastatin-pretreated mesenchymal stem cells improves cardiac performance after acute myocardial infarction: role of CXCR4.

Intracoronary Administration of Allogeneic Adipose Tissue-Derived Mesenchymal Stem Cells Improves Myocardial Perfusion But Not Left Ventricle Function, in a Translational Model of Acute Myocardial Infarction.

An isolated cardiac relapse after allogeneic hematopoietic stem cell transplantation for acute lymphoblastic leukemia.

Cardiac rehabilitation after acute myocardial infarction resuscitated from cardiac arrest.

Stem cells for myocardial infarction.

Stem Cell Therapy for Myocardial Infarction 2001-2013 Revisited.

Cardiac repair in a mouse model of acute myocardial infarction with trophoblast stem cells.

Cardiac repair in a porcine model of acute myocardial infarction with human induced pluripotent stem cell-derived cardiovascular cells.

Thrombolysis for acute myocardial infarction.

Acute intervention for myocardial infarction.

Magnesium for acute myocardial infarction?

Cardiac Stem Cell Hybrids Enhance Myocardial Repair.

Cardiac Relapse of Acute Myeloid Leukemia after Allogeneic Hematopoietic Stem Cell Transplantation.

Bronchogenic Carcinoma with Cardiac Invasion Simulating Acute Myocardial Infarction.

Diagnostics on acute myocardial infarction: Cardiac troponin biomarkers.

Effect of spironolactone on cardiac remodeling after acute myocardial infarction.

Stem cell factor gene transfer improves cardiac function after myocardial infarction in swine.

Integrin β1 Increases Stem Cell Survival and Cardiac Function after Myocardial Infarction.