Regenerative Medicine: Transforming the Drug Discovery and ...perspectivesinmedicine.cshlp.org/content/4/8/a014084.full.pdf · Regenerative Medicine: Transforming the Drug Discovery

Regenerative Medicine: Transforming the DrugDiscovery and Development Paradigm

Sotirios K. Karathanasis

Cardiovascular & Metabolic Diseases, Gaithersburg, Maryland 20878

Correspondence: [email protected]

Despite the explosion of knowledge in basic biological processes controlling tissue regen-eration and the growing interest in repairing/replacing diseased tissues and organs throughvarious approaches (e.g., small and large molecule therapeutics, stem cell injection, tissueengineering), the pharmaceutical industry (pharma) has been reluctant to fully adopt thesetechnologies into the traditional drug discovery and research and development (R&D)process. In this article, I discuss knowledge-base gaps and other possible factors that maydelay full incorporation of these innovations in pharma R&D. I hope that this discussion willilluminate key issues that currently limit synergistic relationships between pharma and ac-ademic institutions and may even stimulate initiation of such collaborative research.

Breakthroughs in stem cell biology and theclinical evaluation of regenerative therapeu-

tics offer an unprecedented opportunity totransform traditional pharma research and de-velopment (R&D) and revolutionize futuremedical practice. Capitalizing on these opportu-nities will require a significant transformation inthe pharma R&D model, including a shift in theway pharma and academic institutions interact.

Although scientific breakthroughs are driv-en mostly by academic institutions, conversionof novel biologic insights into successful medi-cal products is dependent on the drug devel-opment and distribution infrastructure avail-able in traditional pharma. In this context,successful, purpose-driven innovation at the ac-ademia–pharma interface necessitates mutualunderstanding of the several factors including:general philosophy, long-term vision, opera-

tional and cultural paradigms, and key driversand stressors across both of these enterprises.

To describe how emerging technologies inregenerative medicine could transform pharmaR&D and revitalize this entire business sector, Iwill first outline the current pharma R&D busi-ness environment and the manner in which thisenvironment is creating the need for change inthe prevailing business model.

It has been argued that the current phar-maceutical business model is not sustainablebecause of the productivity crisis in pharmaR&D (Pammolli et al. 2011). Many factors areresponsible including escalating R&D costs, in-creased pressures for improved performancedrugs, increased payer pressures, excessive reg-ulatory stringency, increased focus on high-riskresearch involving complex therapeutic targets,and an over-reliance on “molecular reduction-

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Copyright # 2014 Cold Spring Harbor Laboratory Press; all rights reserved; doi: 10.1101/cshperspect.a014084

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ism” to deliver solutions to complex biologicalproblems (Scannell et al. 2012). In addition,limitations in human capital with a robustknowledge base in both translational medicineand therapeutics further intensifies the severityof the current predicament (FitzGerald 2010).

Initiatives to “industrialize” and “stream-line” the discovery process (Bhattacharya et al.2005) and improve process efficiency (e.g., the“lean-six-sigma” methodology) (Sewing et al.2008), are based on last century’s Frederick Tay-lor’s “scientific management” paradigm (Taylor1911) and have been largely unsuccessful in im-proving R&D effectiveness. In fact, they mayhave even contributed to an inward-focusedculture lacking sufficient flexibility to internal-ize novel therapeutic concepts emerging fromacademia. Indeed, the rate of advance in stemcell biology, regenerative medicine, and nano-sciences dramatically outpaces the rate of theirincorporation into pharma R&D.

Taken together, these factors have fueledknowledge-base and innovation deficits inpharma, which will continue to grow unless sys-temic change is made (Drews and Ryser 1996;U.S. Department of Health and Human Servic-es, Food and Drug Administration 2004; Pam-molli et al. 2011).

It is within this contextual landscape thatrecent breakthroughs in developmental biology,stem cell biology, and regenerative medicine areemerging as disruptive technologies with thepotential to revolutionize R&D and radicallytransform future medical practice.

PHARMA’S INTEREST IN STEM CELLBIOLOGY AND REGENERATIVE MEDICINE

Although pharma’s initial reaction to the realmof regenerative medicine has been cautious, it isgenerally recognized that these technologiesrepresent an opportunity for substantial marketexpansion. Indeed, because of the higher bur-den of chronic disease driven by the aging worldpopulation, healthcare costs are expected to in-crease dramatically over the next 20 years. It isestimated that by 2030 the elderly populationin the United States alone will increase by nearly32 million (Werner et al. 2011). This will sub-

stantially increase the need for more cost-effec-tive therapies for degenerative conditions com-mon in the elderly, such as heart disease, cancer,stroke, pulmonary disease, and diabetes, all ofwhich are amenable to regenerative medicineapproaches.

These future demographic trends, medicalneeds, and financial realities are very visible inpharma’s strategic planning horizon and theyare a major driver in pharma’s emerging inter-est in stem cell and regenerative technologies.In this context, pharma’s interests can be broad-ly placed in two categories: (1) drug-screeningtools, and (2) regenerative therapies. It shouldalso be noted that there is a growing interestin applying these technologies, particularly pa-tient-derived induced pluripotent stem cells(iPSCs), to risk stratify individual patients, ul-timately enabling personalized health care, an-other clearly articulated ambition of pharma.

DRUG-SCREENING TOOLS

Drug screening involves a series of decision-making steps designed to filter out irrelevantcompounds and focus on a few promising com-pounds with therapeutic potential. The coreelements of these drug-screening programs typ-ically include three sequential compound-filter-ing steps: in vitro screening, in-cells screening,and in-animal screening. Compounds meetingprespecified criteria in the in vitro screen-ing assays are progressed into the cell-screeningassays, and, if they meet the cell-screening assay,prespecified criteria are further progressed intoanimal models. Compounds emerging fromthis sequence of events are further studied inanimals to determine their pharmacokineticand pharmacodynamic (PKPD) properties anddose-dependent separation of molecular targetengagement from adverse effects because ofnontarget activity. The so-called margins ofsafety (MoS) must be defined before lead com-pounds are tested in humans.

Although there is logic to this drug-screen-ing paradigm, the persistent and growing attri-tion of experimental therapies at all stages ofclinical testing, including postapproval, pointto inherent flaws regarding translatability to hu-

S.K. Karathanasis

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mans (Kola and Landis 2004). By necessity, mostscreening cascades employ nonhuman species,raising the concern that animal disease modelscurrently in use do not recapitulate human dis-ease and, therefore, are of limited translationalpredictive value.

Similar concerns also apply to the variouscell lines used for “compound-filtering.” Al-though engineered cell lines typically used in theearly stages of drug-screening cascades helpselect compounds that trigger pathways mech-anistically linked to molecular targets, in gen-eral, these cells provide very little informationon drug-induced disease-related physiologicalchanges. Efforts to use human-derived cell linesare also plagued with inherent difficulties as cellline behavior may deviate significantly fromthe parental phenotype, and primary cells havelimited expansion potential, undergoing phe-notypic drift under prolonged culturing.

The pressing need for reliable models ofhuman disease has fueled a growing interestin pharma to leverage recent developments instem cell technologies, including stem cell–gen-erated tissues, organs, and chimeric animals,to improve predictivity of drug-screening cas-cades. The critical importance in this area is alsoemphasized by the recent initiative by Nation-al Institutes of Health (NIH), which providesfunding opportunities for research in this area(NIH 2012).

Human Disease-Relevant Cell-Based Models

Human embryonic stem cells (hESCs) were firstisolated 14 years ago (Thomson et al. 1998) andwere subsequently differentiated to diverse hu-man cell types (e.g., cardiomyocytes, hepato-cytes, and neurons). Pharma has exhibitedinterest in using hESCs in drug-screening cas-cades; however, practical implementation hasbeen severely curtailed by a number of prob-lems, including ethical issues of hESCs, limitedscalability of hESC culture in vitro, and the lackof hESCs derived from patients with diseases ofinterest (Grskovic et al. 2011). Yamanaka andcoworkers recently reported a method by whichan adult terminally differentiated cell can betransformed in vitro to a phenotype similar to

hESCs. This method involves transient expres-sion of a cocktail of transcription factors inpatient-derived cells. The resultant cells, iPSCs,can subsequently be differentiated into variouscell types, thus facilitating study of medical con-ditions present in the initial patient (Takahashiand Yamanaka 2006). This technique eliminat-ed ethical concerns associated with ESCs andprovided a renewable source of patient-specificpluripotent cells, rejuvenating pharma’s interestin the development of cellular models of humandisease. This interest has been further fueled bya continuously increasing number of disease-specific iPSC lines, produced primarily withinacademic institutions (Grskovic et al. 2011;Robinton and Daley 2012).

Consistent generation and establishmentof disease- and patient-specific human cell sys-tems with well-defined phenotypes linked tokey aspects of human disease has captured theimagination of both academic and pharma sci-entists. Clearly, incorporation of such models indrug-screening cascades will improve humantranslatability and open the possibility of drug“tailoring” for disease subsegments with similarunderlying pathoetiologies, ultimately contrib-uting to the other aspiration of pharma: person-alized medicine.

Furthermore, the availability of cell systemsthat closely recapitulate human disease patho-etiology has the potential to usher in a new eraof phenotypic screening, whereby compoundsare evaluated for their effects on cellular pheno-types rather than prespecified molecular targets,thus addressing the issue of molecular reduc-tionism discussed above. The concept of pheno-typic screening is not new to pharma (Swinneyand Anthony 2011), but, until recently, cell sys-tems that faithfully recapitulated human diseasewere not available and previous efforts in thisarea using alternative systems were largely un-successful.

Finally, there are even ambitions to develop“in vitro clinical trials” and “patients in a dish,”whereby patient-derived differentiated iPSCsare tested for drug response before the initia-tion of formal clinical studies. Accurate iden-tification of responders before the initiation ofclinical trials could have a marked impact on

Innovation at the Academia–Pharma Interface

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sample size and could provide informationabout drug effects in off-target tissues (Dimoset al. 2011; Grskovic et al. 2011).

Although in theory iPSCs can give rise toall cell types in an adult organism, consistentdifferentiation of these cells into mature celltypes remains a technical challenge. Commit-ment and differentiation of pluripotent stemcells into specific lineages is determined by theexact concentration and timing of the necessaryparacrine factors and related signaling path-ways. In principle, successful differentiationprotocols attempt to recapitulate normal hu-man embryonic development, a still-evolvingarea of research. Nevertheless, in vitro differen-tiation protocols have been developed for a fewcell types such as cardiomyocytes, neurons, andhepatocytes (Dimos et al. 2011; Grskovic et al.2011).

A notable recent accomplishment in thisarea is the consistent generation of human whiteand brown adipocytes from iPSCs and the reca-pitulation of a human disease-relevant pheno-type (insulin resistance) after exposure of thesecells to free fatty acids (FFAs), known to be keydisease drivers in humans (Ahfeldt et al. 2012).

In addition to the difficulties of consistentdifferentiation of iPSCs into tissue-specific celltypes, there is the issue of differentiated cell ma-turity. Frequently, differentiated iPSCs exhibitan immature fetal phenotype in vitro, only as-suming a mature adult phenotype after graftingin animals (Dimos et al. 2011). Clearly, under-standing how the state of maturity of these cellsaffects the relationship between cell phenotypeand disease relevance needs to be further de-fined before the introduction of the cells intodrug-screening cascades.

Finally, the issue of epigenetic somaticmemory of iPSCs needs to be addressed. Recentdata suggest that epigenetic controls present inthe parental cells are preserved in the derivediPSCs and influence their differentiation capac-ity in favor of lineages closely related to the pa-rental cell (Robinton and Daley 2012). Al-though such limitations can be overcome bytreating with DNA demethylating agents or ex-posure to specific cytokines, it is important thatthese issues are addressed up front to avoid po-

tential effects of epigenetic factors from irrele-vant tissues influencing the phenotype in theultimate disease model. It should be also notedthat any epigenetic contributions to disease ex-pression in disease-relevant tissues are likely tobe lost in iPSCs derived from tissues and cellsnot directly involved in the disease process.However, this could be overcome if iPSCs arederived from tissues either participating in oraffected by the disease.

Clearly, robust cell phenotypes closelylinked to human disease are essential beforethe incorporation of iPSC-derived diseasemodels into screening cascades. Although thishas been relatively straightforward for mono-genic cell autonomous diseases, such as in thecase of certain cardiac arrhythmogenic diseases(Grskovic et al. 2011; Oh et al. 2012), it is morechallenging with complex multifactorial diseas-es. For example, diseases such as type 2 diabetesmellitus or atherosclerosis are multifactorial,multiorgan disorders involving a variety of tis-sues and cell types. In these cases, the relativecontribution of each specific cell type to diseaseexpression and a linked disease-relevant cellphenotype needs to be clearly defined beforethe adoption into drug-screening cascades.

It is possible to induce disease-relevant phe-notypes by exposing cells to pathoetiologic fac-tors such as exposure of adipocytes to FFAs toinduce adipose tissue insulin resistance (Ahfeldtet al. 2012). Differentiation of iPSCs from asingle affected patient into multiple tissue typesfollowed by exposure of the differentiated cellsto the same causal stimulus may clarify the rel-ative contribution of different tissue types to adisease state. One such example would includecomparison of myocyte and adipocyte responseto FFAs in culture to compare their relative con-tribution to systemic insulin resistance. Glu-cose-stimulated insulin secretion (GSIS) ofpancreatic b cells produced from the same pa-tient, in the presence and absence of similardisease-inducing factors, would provide infor-mation on the contribution of b-cell dysfunc-tion. Undoubtedly, the data produced by thistype of assay could have major implicationsfor patient-specific drug targeting and person-alized healthcare.

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This concept could be expanded further byleveraging the existing knowledge base in theliterature on physiological and pathophysiolog-ical stimuli known to alter cell behavior in mul-tiple disease states. For example, to study kidneydisease, patient-derived podocytes could betreated with glucose or suPAR to induce pheno-typic changes in podocyte foot morphology,thought to be a surrogate of the disease pheno-type known as podocyte effacement (Wei et al.2011). Such an assay would not only facilitatetranslation of drug candidates, but would alsoenable phenotypic screening to discover noveldrugs for kidney diseases. Similarly, exposureof iPSC-derived intestinal L cells (from multi-ple patients) to drugs designed to increaseGLP-1 secretion could delineate patient popu-lations responsive to specific candidate drugsand could also facilitate phenotypic screeningfor novel drugs capable of inducing intestinalGLP-1 release. This assay could also be used asthe starting point for the study of genetic mark-ers associated with drug responsiveness. Fur-thermore, and particularly relevant to the focusof this collection, a recently reported phenotyp-ic screen for compounds that promote cardio-genesis of ESCs led to discovery of novel com-pounds that block TGF-b signaling by inducingTGF-bR2 degradation, thereby selectively di-recting uncommitted mesodermal cells towardcardiac lineages (Willems et al. 2012). Becauseof the importance of TGF-b signaling in endo-thelial to mesenchymal transitions (EndMT)and the role of this process in postmyocardialinfarction (MI) cardiac fibrosis (von Gise andPu 2012), it will be very exciting to test thesecompounds for their effects in in vivo MI mod-els. There are many more such examples includ-ing patient-derived macrophages for reversecholesterol transport (RCT) and endothelialcell activation and cholesterol uptake assaysfor vascular therapies to name only a few(Adams and Garcıa-Cardena 2012).

Another exciting development is the incor-poration of human iPSC-derived tissues and or-gans in whole animals, creating human–animalchimeras. For example, a mouse chimeric modelwith human iPSC-derived human adipose tis-sue (Ahfeldt et al. 2012) and a similar model

with hESC-derived pancreatic progenitor cells(Schulz et al. 2012) were recently reported. Al-though such models offer an unprecedented op-portunity to study human tissue function in thecontext of whole organism physiology, long-term preservation of human tissues in suchmodels is dependent on continuous suppres-sion of the recipient animal’s immune system.Use of immunosuppressed mouse strains canfacilitate this type of experiment, but new solu-tions may be required to make the chimericsystem more widely applicable.

Drug Toxicology and Safety Assays

Testing candidate drug toxicity in animals be-fore human clinical trials is unreliable. It is oftenfound that compounds that appear safe in ani-mals are toxic in humans. Predictive toxicologyusing human iPSC-derived tissue cells couldenable attrition of compounds with unaccept-able safety profiles early in the evaluation pro-cess and will also decrease animal use (Wobusand Loser 2011). This is a rapidly advancingarea with the European Union 7th FrameworkProgramme “Innovative Medicine Initiative”supporting further research (Innovative Medi-cine Initiative 2011), and several cell-based pre-dictive toxicity assays are now commerciallyavailable. Remaining challenges in this area in-clude scalability of cell culture and productionof homogeneous and mature cell populations(Wobus and Loser 2011).

In summary, although there has been re-markable progress over the last few years in es-tablishing human iPSC-derived models of dis-ease, several remaining gaps, such as consistentgeneration of fully differentiated cells, establish-ment and validation of disease-relevant pheno-typic read-outs, and development of disease-relevant chimeric human–animal constructs,need be addressed before such technologiesare fully integrated within pharma R&D. Devel-opment of human multicellular models that canreplicate aspects of human organ physiology,disease pathogenesis, cell-type diversity, and ge-nomic complexity, as envisioned by the NHIRFA mentioned above, has the potential to rev-olutionize future drug discovery screening.



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REGENERATIVE THERAPIES

The concept of tissue regeneration has beenpart of human civilization for millennia. In-deed, the ancient Greek legend of Prometheusincludes mention of liver regeneration. Accord-ing to the legend, Prometheus, a deity whotook pity on mortals, stole fire from Zeus tobring them light and comfort. Zeus, angered bythis gesture, punished Prometheus by tether-ing him to a rock and sending an eagle to eathis liver each day. Overnight, the liver wouldregrow for the eagle’s return. Regenerativemedicine in humans, in the form of organtransplantation and hematopoietic stem celltransplantation has been performed since the1950s (Maienschein 2011). More recently, withthe re-emergent interest in regenerative thera-peutics, there has been a flurry of preclinicaland clinical activity in this space. In general,these efforts involve approaches such as celltransplantation or paracrine factor–inducedactivation of endogenous stem cells using smallor large molecules and bioactive materials (bio-polymers, nanofibers). In the following para-graphs, I discuss opportunities and gaps thatneed to be filled before pharma is ready toengage further.

Mesenchymal Stem Cell Autologousand Allogeneic Cell Therapies

Transplantation of bone marrow (BM)–de-rived hematopoietic stem cells is now a standardclinical procedure for the treatment of certainhematological conditions. Mesenchymal stemcells (MSCs), a fibroblast-like subpopulationfound in BM and other tissues, have been shownto differentiate into a variety of adult tissuesand, after systemic administration, have beenshown to be safe regarding tumorigenesis andother adverse side effects (Jung et al. 2012). In-deed, MSC regenerative therapies are being cur-rently tested in a variety of diseases includ-ing immune modulation, bone and cartilagerepair, cardiac regeneration, and wound repair(Sensebe et al. 2010; Culme-Seymour et al.2012). It is also noteworthy that such adult tis-sue–derived stem cell regenerative cell therapies

are advancing rapidly in veterinary medicinewhere there is less regulatory stringency. Forexample, adult stem cell therapies for equinearticular cartilage restoration and chondrogen-esis have been shown to improve clinical symp-toms (Borjesson and Peroni 2011).

Despite these developments and the in-creased enthusiasm for such approaches to tis-sue regeneration, there are clear scientific, clin-ical, regulatory, and financial risks (McKernanet al. 2010; Tozer 2010; Jung et al. 2012). Scien-tific and clinical risks include choice of cell do-nor. Although autologous cells would be pref-erable, donor age appears to affect the efficacy ofthe isolated cells. It has been shown that agingsignificantly reduces survival and differentia-tion potential of BM-MSC (Roobrouck et al.2008). This age-dependent regenerative declineof endogenous stem cells, referred to as “stemcell aging,” is thought to be driven by severalfactors in the tissue niche from which the adultcells are derived, including inflammation andexcessive oxidative stress present in the diseasecontext. These age-related changes in stem cellphenotype and behavior may significantly im-pact the regenerative potential of cell-replace-ment therapies for tissue regeneration (Jasperand Kennedy 2012).

Reliable scale production of the desired phe-notype is necessary, but not sufficient, for asuccessful cell-based therapeutic strategy. Issuespertaining to delivery of the therapeutic cell tothe target organ would need to be solved. Re-ported attempts to deliver putative therapeuticcells to the heart exemplify the challenges asso-ciated with cell delivery (Ptaszek et al. 2012).Even if a cell can be delivered to the target organin a safe manner, which ensures optimal reten-tion at the site, survival and maturation of thiscell in situ are not guaranteed (Le Huu et al.2012).

It is also worth noting that the discovery ofiPSCs generated a rush of enthusiasm aboutusing patient-derived iPS cells for cell trans-plantation. However, a number of concernssuch as risk for tumorgenicity, epigenetic mem-ory, and differentiated cell purity need to beaddressed before this vision becomes reality(Vitale et al. 2011).

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The mechanism by which transplanted cellsinfluence cardiac physiology is strongly debated,with a numberof leading investigators in the fieldarguing that most of the beneficial effects are aresult of paracrine effects rather than direct par-ticipation of the transplanted cells in tissue re-generation (Penn et al. 2011). Some investigatorsfurther argue that stem cells are not even neces-sary as secreted factors may be entirely responsi-ble for the benefits observed clinically. This, to-gether with the need to improve the endogenousmicroenvironment, suggests that alternative ap-proaches involving delivery of paracrine andother trophic factors may hold promise.

From the regulatory perspective, relevant is-sues include: (1) cell characterization and qual-ity control (identity, purity, potency, tumor-genicity, chromosomal stability, and processvalidation), (2) biodistribution (ability to trackcells into different microenvironments and tis-sues), (3) disease relevance of animal models(large animal models reflecting the disease in-dication are preferable for surgical implantationand follow-up of cell products), (4) in vivo dif-ferentiation (desired mode of action), (5) im-mune rejection and persistence, (6) clinicalpharmacodynamic biomarkers (to follow thedifferentiation status of the cells during treat-ment), (7) clinical pharmacokinetic markers (tofollow the administered cells during the study),(8) clinical dose finding, (9) efficacy on clinical-ly meaningful endpoints/overall efficiency, and(10) safety regarding teratomas and pathologi-cal self-renewal (Halme and Kessler 2006; Euro-pean Medicines Agency 2011).

From the business perspective, relevant is-sues include: (1) manufacturing (regulationscovering product manufacturing, packaging,facility validation, consistency and safety of ma-terials, sterility of the product, and, where ap-propriate, sterilization and viral inactivationprocedures), (2) logistics (living cell productsrequire precisely controlled conditions andmay require cryopreservation and implementa-tion of cold chain logistics and distributioncapabilities), (3) reimbursement, and (4) salesand marketing (awareness and understandingof consumers and health care providers of re-generative medicines therapies) (Tozer 2010).

In summary, although a recently publishedcomprehensive review on ongoing cell therapytrials ends with the optimistic note that “overall,these data are highly encouraging for the emerg-ing cell therapy industry, given the early stage ofthe technology platform in its life cycle, totalnumber of trials, the spread of clinical trialphases and the range of medical indications”(Culme-Seymour et al. 2012), there remains sig-nificant challenges before this becomes a rou-tine approach to discovering and developingnew medicines.

Paracrine Factors and DevelopmentalPathway Modulators

Use of paracrine factors as cardiac regenerativetherapeutics is based on the premise that theheart has intrinsic regenerative capacity andthat progenitor cells in the adult heart, if ade-quately stimulated, could influence the heart’sresponse to injury. It is now largely accepted thatthe beneficial effects observed in trials associat-ed with putative cardiac stem cells are the resultof paracrine factor release from these cells, rath-er than residence of delivered cells in the heart(Penn et al. 2011).

Although amphibians and certain fish areknown to maintain a robust ability for cardiacregeneration throughout life, until recently, thiswas thought to be absent in mammals. In fact,many of the earliest cardiac regenerative thera-pies were designed based on the assumption thatcardiac cells possessed no regenerative capacity.However, recent work has shown that cardiacregenerative activity does exist in mice, but ex-tinguishes very rapidly after birth (Porrello et al.2011). An evolving body of work using mousemodels has revealed the presence of a progenitorpopulation that persists on the epicardial sur-face of the adult heart. These cells, referred to asepicardial progenitor-derived cells (EPDCs), arestimulated to divide after cardiac injury (e.g.,myocardial infarction) but are not independentlycapable of reversing injury. This cell populationis, therefore, a compelling target for paracrinefactor-based therapeutics (Smart et al. 2012).

The growing understanding of the networksof signaling events that control early cardiac



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progenitor cell specification and differentiation,such as the sequential activation of the Notch,Wnt/b-catenin, and BMP (Klaus et al. 2012),and the knowledge that some of these systemsregulate stem cell proliferation and differentia-tion in the adult heart, raise the possibility thatsmall molecule modulators of these signalingsystems could redirect stem cell fate in the dis-eased adult heart leading to clinically effectivecardiac regeneration. For example, it was recent-ly shown that canonical Wnt signaling limitscell renewal of an adult stem cell populationknown as cardiac side population (CSP), andblocks their ability to contribute to endogenouscardiac regeneration. These data suggest thatWnt signaling inhibitors exert potent cardio-protective effects after myocardial injury (Oiko-nomopoulos et al. 2011).

Inspired by such a possibility, several inves-tigators have employed phenotypic screens toidentify molecules with cardiac regenerationpotential. A recent screen in zebrafish usingheart size as a screening end point yielded cer-tain Wnt inhibitors (cardionogens) shown toinduce cardiogenesis during and after gastrula-tion (Ni et al. 2011). Similarly, a mouse ESCphenotypic screen identified a 1,4-dihydropy-ridine selective TGF-b inhibitor that promotescardiogenesis and beating cell clusters in mESCwhen added between days three and five of dif-ferentiation (Willems et al. 2012).

Furthermore, modulation of the paracrinefactor milieu via delivery of specific growth fac-tors and chemokines to the myocardial tissuehas been reported to lead to effective cardiacregeneration. Cardiac regeneration has been ob-served with TB4 (Smart et al. 2011), VEGF (Linet al. 2012), SCF (Yaniz-Galende et al. 2012),and SDF-1, which has recently shown promisein improving heart function in patients withsevere heart failure (Penn et al. 2012).

It should be also mentioned that an alterna-tive approach to cardiac regeneration involvingdirect reprogramming of endogenous fibro-blasts into functional cardiomyocytes has alsobeen recently reported. Although this offers theexciting possibility to regenerate the myocardi-um, reduce scar size, and improve cardiac func-tion, these benefits need to be confirmed in

large animal models with thicker myocardiumsimilar to that of humans (Bruneau 2012).

Although this paracrine therapy approach isconceptually appealing and consistent with thecurrent pharma business model for moleculeoptimization and delivery to the marketplace,there are questions that need to be addressed.

For example, if the replicative and chrono-logical aging as well as the scar tissue and otherpathogenic factors affect the functional capacityand regenerative potential of endogenous stemcells in heart disease patients, as has been sug-gested, the therapeutic effectiveness of paracrinetherapies could be hampered (Mohsin et al.2011). Encouragingly, however, a recent com-parison of the postinfarction myogenic poten-tial of neonatal and adult hearts revealed thatthe failure of adult postinfarct myogenesis is notbecause of context-dependent restriction ofprecursor differentiation, but rather because ofthe distinct developmental potential of adultand neonatal heart progenitor cells (Jesty et al.2012). Elucidation of key mechanisms and dis-covery of critical molecular cues that controlstem cell development may reveal approachesfor resetting the developmental potential ofstem cells in adult heart disease patients, therebyallowing recapitulation of neonatal heart regen-eration and improved cardiovascular function.

Avery recent example along these lines is thecardiac regeneration and improved heart func-tion observed after adenoviral gene transfer ofthe c-kit ligand stem cell factor (SCF) into theinfarcted myocardium. The experimental evi-dence suggests that transient overexpression ofSCF in the heart increases recruitment of c-kitþ

cardiac cells and leads to regeneration via acti-vation of the Wnt signaling (Yaniz-Galendeet al. 2012). Similarly, elucidation of mechano-chemical pathways responsible for transmittingtissue rigidity signals that control cardiac stemcell proliferation and cell-fate choices may allowchemical reprogramming of the patient cardiacmechanical environment in line with the regen-erative strategy therapeutic objectives (Kshitizet al. 2012).

It should also be noted that, although up tonow only individual paracrine factors have beenexplored, robust and effective cardiac regenera-

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tion may require a combination of paracrinefactors. Therefore, a challenge for the future isto define such paracrine factor cocktails andtailor them toward indication-specific cardiacpathobiologies as the inherent paracrine factorenvironment may differ significantly betweenacute myocardial infarction and chronic heartfailure (Penn et al. 2011).

It should also be emphasized that the overallstrategy to reengage powerful developmentalsystems and direct them toward new tissue gen-esis and organ re-engineering is associated withobvious safety risks. To ameliorate these con-cerns, there is a need for effective local deliverysystems. This is an important area of researchand progress is regularly reported in the litera-ture. Recently, self-assembling peptide nano-fibers were used for delivery of VEGF (Linet al. 2012) and collagen-chitosan hydrogelswere used for delivery of TB4 (Chiu et al.2012) both resulting in enhanced cardiac regen-eration.

In conclusion, this is a very exciting area ofresearch with the potential to deliver novel med-icines that will radically change current medicalpractice for the treatment of heart diseases. Inaddition, it should be reiterated that these para-crine and developmental pathway modulatorapproaches fit perfectly within the current phar-ma R&D paradigm providing a unique incen-tive for pharma to contribute their strong drugR&D experience to focused pharma/academiacollaborations aiming to explore novel thera-peutic concepts and deliver efficient clinicalproof of concepts.

CONCLUDING REMARKS

Pharma is in urgent need to turn around andrevitalize their business model. Innovative tech-nologies such those offered by stem cell biologyand regenerative medicine promise a disruptive,“step-change” transformation of the currentpharma R&D model and the opportunity todeliver medicines that will radically alter futuremedical practice (Denoon and Vollebregt 2010).Although sensationalization of the medical po-tential of regenerative medicine is unavoidableand has raised concerns about inflated expecta-

tions (Tozer 2010), the focused research activi-ties in this area by academia, government (in-cluding the NHI and IMI initiatives), and privateenterprises assure timely resolution of remain-ing gaps and actualization of the immense clin-ical value of these technologies in the near fu-ture. Pharma’s primary strengths are the processby which lead compounds are turned into amarketable drug, which is a formidable processthat includes large-scale production, distribu-tion, quality control, and interactions with reg-ulatory agencies. Also included are structuringand conducting clinical trials, as well as reim-bursement and lobbying, which are essential forthis emerging business sector to mature into asustainable business enterprise (Denoon andVollebregt 2010). Given the complementarystrengths of academic institutions and theirskills in identification and validation of noveltherapeutic targets, a collaborative approach be-tween pharma and academia is essential to bringthe exciting potential of regenerative therapeu-tics into a reality.

ACKNOWLEDGMENTS

I am truly indebted to my colleagues LaurenDrowley and Richard Lawson, AstraZeneca,and Leon Ptaszek, MGH, Harvard, for theirthoughtful reading of the manuscript and theirmany suggestions for improvement.

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2014; doi: 10.1101/cshperspect.a014084Cold Spring Harb Perspect Med Sotirios K. Karathanasis Development ParadigmRegenerative Medicine: Transforming the Drug Discovery and

Subject Collection The Biology of Heart Disease

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Blanpain, et al.Sigolène M. Meilhac, Fabienne Lescroart, Cédric

DiseasePersonalized Genomes and Cardiovascular

Kiran Musunuru Cardiovascular Biology and MedicineToward a New Technology Platform for Synthetic Chemically Modified mRNA (modRNA):

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Congenital Heart DiseaseComplex Genetics and the Etiology of Human

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Xiaojun Lian, Jiejia Xu, Jinsong Li, et al.Genetic Networks Governing Heart Development

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Monogenic DisordersonHeart Disease from Human and Murine Studies

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Sotirios K. Karathanasisfrom a Research-Based Pharmaceutical CompanyCardiovascular Drug Discovery: A Perspective

G. Gromo, J. Mann and J.D. FitzgeraldMyocardial Tissue Engineering: In Vitro Models

and Christine MummeryGordana Vunjak Novakovic, Thomas Eschenhagen

Genetics and Disease of Ventricular Muscle

G. SeidmanDiane Fatkin, Christine E. Seidman and Jonathan

DiseasePluripotent Stem Cell Models of Human Heart

Dorn, et al.Alessandra Moretti, Karl-Ludwig Laugwitz, Tatjana

Embryonic Heart Progenitors and Cardiogenesis

et al.Thomas Brade, Luna S. Pane, Alessandra Moretti,

http://perspectivesinmedicine.cshlp.org/cgi/collection/ For additional articles in this collection, see

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