Mesenchymal Precursor Cells as Adjunctive Therapy in Recipients of Contemporary LVADs Deborah D. Ascheim, Annetine C. Gelijns, Daniel Goldstein, Lemuel A. Moye, Nicholas Smedira, Sangjin Lee, Charles T. Klodell, Anita Szady, Michael K. Parides, Neal Jeffries, Donna Skerrett, Doris A. Taylor, J. Eduardo Rame, Carmelo Milano, Joseph G. Rogers, Janine Lynch, Todd Dewey, Eric Eichhorn, Benjamin Sun, David Feldman, Robert Simari, Patrick T. O'Gara, Wendy Taddei-Peters, Marissa A. Miller, Yoshifumi Naka, Emilia Bagiella, Eric A. Rose and Y. Joseph Woo Circulation. published online March 28, 2014; Circulation is published by the American Heart Association, 7272 Greenville Avenue, Dallas, TX 75231 Copyright © 2014 American Heart Association, Inc. All rights reserved. Print ISSN: 0009-7322. Online ISSN: 1524-4539 The online version of this article, along with updated information and services, is located on the World Wide Web at: http://circ.ahajournals.org/content/early/2014/03/28/CIRCULATIONAHA.113.007412 Data Supplement (unedited) at: http://circ.ahajournals.org/content/suppl/2014/03/28/CIRCULATIONAHA.113.007412.DC1.html Permissions: Requests for permissions to reproduce figures, tables, or portions of articles originally published in Circulation can be obtained via RightsLink, a service of the Copyright Clearance Center, not the Editorial Office. Once the online version of the published article for which permission is being requested is located, click Request Permissions in the middle column of the Web page under Services. Further information about this process is available in the Permissions and Rights Question and Answer document. Reprints: Information about reprints can be found online at: http://www.lww.com/reprints Subscriptions: Information about subscribing to Circulation is online at: http://circ.ahajournals.org//subscriptions/ Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Mesenchymal Precursor Cells as Adjunctive Therapy in Recipients of Contemporary LVADs Running title: Ascheim et al.; Mesenchymal Precursor Cell trial in LVAD patients Deborah D. Ascheim, MD1; Annetine C. Gelijns, PhD1; Daniel Goldstein, MD2; Lemuel A. Moye, MD3; Nicholas Smedira, MD4; Sangjin Lee, MD4; Charles T. Klodell, MD5; Anita Szady, MD5; Michael K. Parides, PhD1; Neal Jeffries, PhD6; Donna Skerrett, MD7; Doris A. Taylor, PhD8; J. Eduardo Rame, MD9; Carmelo Milano, MD10; Joseph G. Rogers, MD10; Janine Lynch, MPH1; Todd Dewey, MD11; Eric Eichhorn, MD11; Benjamin Sun, MD12; David Feldman,, MD12; Robert Simari,, MD13; Patrick T. O’Gara,, MD14; Wendy y Taddei-Peters,, 6 6 15 PhD ; Marissa A. Miller, DVM, MPH ; Yoshifumi Naka, MD ; Emilia Bag Bagiella, ag gie iell lla, ll a, P PhD hD1; Eric A. Rose, MD1; Y. Joseph Woo, MD9 1 Icahn School of Medicine att Mount Sinai, New York, NY; 2Montefiore-Einstein Heart Center, Bronx, Brron nx, x, NY; NY; 3Th The he University U iversity of Texas, Houston, Un Houstonn, TX TX;; 4Cleveland Clin T Clinic nic F Foundation, oundation, Cleveland, 5 6 OH OH; H; T The hee University Univer Un ersi sity ty ooff Fl Flor Florida, o id da, Gai Gainesville, a neesvville, FL FL; N National ation onal al Hea Heart artt L Lung ungg aand un nd Bl Bloo Blood ood In Institute, nst s it i ut u e, NIH NIH, B Bethesda e a, MD; MD 7Me Mesoblast Meso soobl blas astt Inc., as Inc. In c , New c. New York, York Yo rk, NY; rk NY Y; 8Te Texas Texa xass Heart xa H ar He art In Inst Institute, sttit itut ute, H Houston, oust ou ston st on,, TX; on TX; 9Th Thee Bethesda, University Univ Un iversity of Pe Pennsylvania, ennsyylvaaniaa, Phil P Philadelphia, hi ad del elp phia, PA phi PA; 10Du Dukee University, Uni nive versitty, Durham, ve Durrhaam, NC; NC; C 1111Bay Baylor B ayloor H Health eaalth Care Care System, Sys y te tem, m D m, Dallas, alllaas, T TX; X; 1122Minn Minneapolis M in ea eapo po oliis H Heart eaart art In Institute nstit ituute ute Fo Foundation, ounda undaatiion on,, Mi Minn Minneapolis, nnnea eapo p li po l s, M MN; N; 13Ma N; Mayo May yo 14 15 C Cl Clinic, inic in ic,, Rochester, ic Roch Ro ches esste terr, r, MN; MN; Br Brigham rig igha ham ha m an andd Wo Wome Women’s men’ me n’ss Ho n’ Hosp Hospital, sppit ital al,, Bo al Bost Boston, stton on, MA MA;; Co Columbia Colu lumb lu mbia mb iaa U University nive ni vers ve rsit rs ityy it Medi Me Medical dica di call Ce ca Cent Center, nter nt er, New er New Yo York York, rk, NY rk Address for Correspondence: Deborah D. Ascheim, MD InCHOIR, Icahn School of Medicine at Mount Sinai One Gustave L. Levy Place, Box 1077 New York, NY 10029 Tel: 212-659-9567 Fax: 212-423-2998 E-mail: [email protected] Journal Subject Codes: Heart failure:[110] Congestive, Heart failure:[11] Other heart failure, Treatment:[27] Other treatment, Cardiovascular (CV) surgery:[37] CV surgery: transplantation, ventricular assistance, cardiomyopathy 1 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Abstract Background—Allogeneic mesenchymal precursor cells (MPC) injected during left ventricular assist device (LVAD) implantation may contribute to myocardial recovery. This trial explores the safety and efficacy of this strategy. Methods and Results—In this multi-center, double-blind, sham-procedure controlled trial, 30 patients were randomized (2:1) to intramyocardial injection of 25M MPCs or medium during LVAD implantation. The primary safety endpoint was incidence of infectious myocarditis, myocardial rupture, neoplasm, hypersensitivity reaction, and immune sensitization (90 days postrandomization). Key efficacy endpoints were functional status and ventricular function, while temporarily weaned from LVAD support (90 days post-randomization). Patients were followed until transplant p or 12 months post-randomization, p , whichever came first. Mean age g was 57.4 (±13.6) ±13.6) years, mean LVEF 18.1%, and 66.7% were destination therapy LVADs.. No saf safety afet af etyy et events were observed. Successful temporary LVAD weaning was achieved in 50% of MPC and 20% of control patients patients at 90 days (p=0.24); the pposterior osterior probability that MPCs increased the likelihood ike keli liihood ho d ooff succ successful cces essf s ul w weaning e niing is 93 ea 93%. 3%.. At 90 ddays, ays, 3 deaths ay dea eath t s occurred occcu c rrred in in control co onttro rol andd none non in MPC MP C patients. Mean M an LVEF Me LVE VEF F following folllow fo llow win ng successful suucceesssful f wean weann was was 24.0% 24.0% 24.0 % (MPC=10) (MPC (M PC=1 PC =1 10) and andd 22.5% 22. 2.5% 5% (Control=2) Conntr t ol o =2)) (p (p=0 (p=0.56). =0 0.556) 6).. A Att 122 mo mont months, nths hs,, 30% hs 30% of of M MPC PC C aand nd d 440% 0% % ooff con ccontrol ontroll ppatients ati tien e ts en t w were ere su ere successfully ucces esssfu sfully temporarily emp mpor oraril ilyy weaned wean we a ed from from m LVAD LVAD support support rt (p=0.69) (p= p=0. 0 69 69) and nd 6 deaths dea eath ths occurred occu oc curr rred in in MPC MPC patients. paati tien nts ts. Donor-specific Donor-sp pecciffic H HLA LA ssensitization en nsi siti tiza ti zatiionn ddeveloped za evel elop el oped ed iin n 2 MPC MPC and andd 3 control an co ro contro roll patients pati pa tien ti ents ts and and resolved res esoolved byy 122 months. Conclusions—In this preliminary trial, administration of MPCs appeared to be safe and there was a potential signal of efficacy. Future studies will evaluate the potential for higher or additional doses to enhance the ability to wean LVAD recipients off support. Clinical Trial Registration Information—ClinicalTrials.gov; Identifier: NCT01442129. Key words: left ventricular assist device, heart failure, stem cell, randomized controlled trial, Mesenchymal Precursor Cell, Placebo 2 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Left ventricular assist devices (LVADs) have well-documented survival and quality of life benefits in patients with advanced heart failure both as a bridge to cardiac transplantation and as a long-term therapy in patients who are not transplant candidates.1-4 Reports of improved myocardial function have motivated investigation of the use of LVADs as a bridge to recovery. While most LVAD recipients do show some indications of reverse remodeling of the left ventricle as evidenced by salutary changes in ventricular structure, myocyte contractile strength5, normalization of extracellular matrix and tissue and circulating neurohormones6, and programs of gene expression7-10 these improvements are rarely sufficient to allow removal of the device.11 The disconnect between reverse remodeling and recovery of cardiac function have prompted efforts to investigate adjunctive d therapies to LVAD support, including novel pharmacotherapies12 and stem cells as potential interventions to augment ventricular recovery. Recent Rece Re c nt ppre-clinical ce re-c re -clinical and clinical evidencee ssuggests uggests that myo myocardial yocard yo rd dia iall transplantation of allogeneic al llo oge g neic mesenchymal mes esen en nchym mall stem ste tem m cells, cell ce llls, in in particular, partiicular, can can enhance enha en hanc ncee cardiac nc cardia iacc pe performance erf rfoorm ormanc mancce in set settings etti et tin ngs of ngs 13-15 13-1 15 acute ac cut utee an aand d ch chr chronic ronicc fu roni ffunctional nctiion onall iimpairment. mpai mp airm ai rmeent. ent. t 13Unlike Unllik ikee whole whol wh olee organ ol orga or gann transplantation ga tran tr a spla an spla l nttat atio i n or many io man ny ot other theer allogeneic cell cel elll transplants, t an tr ansp spla sp laant n s,, mesenchymal mes e en ench chym ch ymal ym a stem al ste tem m cell celll transplants tra r nsspl plan ants an ts do do not not appear appe ap pear pe a to ar to cause caus ca usee rejection us and instead may be associated with evidence of induced tolerance to the donor. 16, 17 We have, therefore, begun investigation of allogeneic mesenchymal precursor cell transplantation concomitant with LVAD placement in patients with advanced heart disease. While our ultimate goal is the achievement of robust bridging to recovery, allosensitization could adversely impact donor suitability in LVAD recipients who are transplant candidates. Accordingly, the primary goal of the initial trial reported here was exploration of the safety of intramyocardial implantation of a single low dose of allogeneic mesenchymal precursor cells (MPCs) together with assessment of left ventricular performance during short intervals of 3 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 temporary reduction of LVAD support, over 1 year of observation after the implants, to assess safety and any impact on reverse remodeling. Methods Study Design and Trial Oversight This early phase, randomized trial was designed to enroll 30 patients, and if safety would be established, a larger follow-up trial will be conducted. Patients were randomly assigned in a 2:1 ratio to 25 million MPCs (Mesoblast, Inc.) or control, comprised of cryoprotective medium alone (50% Alpha-MEM/42.5% ProFreeze NAO Freezing Medium/7.5% DMSO). Randomization was blocked to ensure equivalence of group size. All investigators and patients were masked mask ma skked to to treatment reatment intervention and overall outcomes data. Endpoints were measured monthly until 90 days, every months randomization. All da ays ys,, an andd ev eve ery 60 ddays ays thereafter until 12 month ths aafter th fter randomiz zat a io on. n A ll patients were transplantation transplant until months ffollowed olllowed untill cardiac carddiac ac tra rans ra nspl ns plaanta pl antati t on ti on ((for for bbridge riddge to tra anspllan antt [BTT]) [B BTT]) TT ) oorr un ntil til 12 m o t hs on following randomization BTT Destination Therapy whichever first. fo oll llow owin ow i g ra in rand ndom om mizat izatiionn (f (forr B TT aand nd dD esttina es nati na t on ti on T hera hera rapy py [[DT]), DT]) DT ]),, wh whic che hevver ver ca came me fir irrstt. The trial tria tr iall was ia was conducted c nd co n uc ucte tedd in 11 te 11 U.S. U S. centers U. cen nte ters r with rs wit i h a Data Dat ataa and an Clinical Clin Cl inic in ical ic al Coordinating Coo oord rdin rd in nat a ing Center (DCC; International Center for Health Outcomes and Innovation Research [InCHOIR], Icahn School of Medicine at Mount Sinai) under an investigational new drug application. Interagency Registry for Mechanically Assisted Circulatory Support (INTERMACS) definitions were utilized for all relevant adverse events; bleeding events were defined by transfusion of 4 units of packed cells within any 24 hour period during the first 7 days post LVAD implantation, and any transfusion of packed cells within any 24 hour period thereafter. An independent Clinical Events Committee adjudicated adverse events and causes of death. An NIH-appointed protocol review committee (PRC) reviewed the trial design and data and safety monitoring board 4 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 (DSMB) reviewed the trial progress. Institutional review boards of participating centers and the DCC approved the protocol, and all patients provided written informed consent. Patients and Interventions The target population was adults with end-stage heart failure, of either ischemic or non-ischemic etiology, who had a planned, clinically indicated LVAD implantation for BTT or DT. Assist devices were required to be FDA approved, contemporary, implantable, continuous flow LVADs; selection of the specific device was left to the discretion of the surgeon. Patients were ineligible if percutaneous LVAD or biventricular mechanical support was anticipated, if they had cardiothoracic surgery or myocardial infarction within 30 days prior to randomization, had were undergone prior cardiac transplantation, LV reduction surgery, or cardiomyoplasty, cardiomyoplas astty, ty, orr w eree er considered to have an acute reversible cause of heart failure. Other selected exclusion criteria included antibody known ncllud uded ed tthe he ppresence reese sennce nc of >10% anti-HLA antibo ody dy ttiters iters with kno own w sspecificity peci pe c ficity to MPC donor HLA antigens, infection within prior H LA A antigens s, aactive ctive ssystemic yste ys temi te micc in mi infe f ctio fe ct on wit thiin 488 hhours ouurs urs pr rio or to o rrandomization, ando an dom do miza zaation tion,, hi hhistory s orry of st of cancer within prior ca anc ncer er prior pri rior or to to screening sccreeen eniingg (excluding (exc (e xcclu udi ding ng basal basal asal cell celll ca ccarcinoma), rciinooma ma), ), oorr st sstroke roke ke w ithhin hin 30 ddays ayss pr ay rior ior to randomization. Patients were ineligible received andomizatio on. n P atie at ieent ntss we ere ine neli ne ligi li giibl b e iff tthey h y ha he hadd re rece ceiv ived iv ed pprior rior ri or sstem tem te m ce cell ll ttherapy heeraapy ffor o cardiac or repair, or any investigational cell based therapy within 6-months prior to randomization. (Supplemental Materials include complete eligibility criteria). The investigational agent was allogeneic mesenchymal precursor cells (MPCs), which are a STRO-3 immuno-selected, culture-expanded, immature sub-fraction of adult bone marrowderived mononuclear cells.18 The allogeneic MPCs are formulated and cryopreserved in 7.5% DMSO/50% Alpha Modified Eagle’s Medium (MEM) and 42.5% ProFreeze® and stored in the vapor phase of liquid nitrogen until use. Cell procurement, processing, cryopreservation, and storage procedures were performed by a contract manufacturing facility under cGMP conditions. 5 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Donor and process testing were conducted for transmissible infectious diseases, karyotype, tumorigenicity, sterility, endotoxins, and mycoplasma. The product is characterized by cell count, viability, surface antigen expression of STRO-1, CC-9, and HLA class I and II. Cryopreserved products were shipped to sites for local storage, and cells were thawed and injected according to study procedures. Intramyocardial injections of MPCs or control were performed at the time of LVAD implantation. Injection procedures were protocol-defined, providing standardization of the intervention across sites and designed to maximize injections across as much of the left ventricular (LV) myocardium as possible. LVAD implantation and management were performed inn accordance with the Directions for Use, and the protocol provided guidance with wit ithh respect reesppec ectt to t long-term ong-term management, including optimization of hemodynamic off-loading of the LV, reduction mitral mean edu uct ctio ionn of m io ittra rall rregurgitation egurgitation when present, aand n optimization of nd o me ean blood pressure. LVAD Dw weaning eaaniing w was as ddefined as efin efin ned aass a tra transient ansientt rreduction eduuctio ctionn in i ppump um mp sspeed peed eed to m minimize inim in im miz ize fo forw forward rw ward flow fl low w tthrough h ou hr ough gh tthe he ppump umpp in um in oorder rd der tto o as aassess sse seesss nnative at ve m ative myocardial yoocaard dia iall fu func function. ncti nc tion on.. P on Protocol-specified rootoc otoccol ol-s -spe peccifie pe ci ed guidelines for orr weaning weaani ning ng were werre adopted a op ad opte teed from from the the Harefield Hare Ha refi re f el fi eldd Hospital Hosp Ho spit sp ital it a protocol, al pro roto toco to col, co l, and and included inc nclu lude lu d d guidance for antithrombotic regimens, incremental speed reductions to a “low speed” target of 6,000 rpm, and monitoring of patients during the wean. The 6000 rpm target was chosen as this is the minimum speed necessary to prevent retrograde flow through the pump into the LV.19 Weans were performed in patients deemed to be clinically stable by the clinical team and were terminated if patients developed signs or symptoms of low output or vascular congestion, such as light headedness, dyspnea, fatigue, chest pain or pulmonary edema. Patients who completed weaning but developed transient symptoms at any point during “low speed” were categorized as “wean failures” for that particular wean. A six minute walk (6MW) was performed after 20 6 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 minutes of “low speed”. A comprehensive echocardiogram was performed at full support prior to the wean, at 15 minutes of “low speed” and again following the 6MW. The LVAD was reprogrammed to full support thereafter. Endpoints The primary endpoint was safety, defined by the incidence of potential study intervention-related adverse events within 90 days post-randomization, including infectious myocarditis, myocardial rupture, neoplasm, hypersensitivity reaction, and immune sensitization (defined as a clinical syndrome accompanied by detection of donor specific antibodies within 30 days of onset of the syndrome). The key efficacy endpoints were functional status, defined by the ability to tolerate wean from LVAD support for 30 minutes without signs or symptoms of hypoperfusion, hypoper erfu fusi s onn, and si and left le ventricular ejection fraction (LVEF) while weaned from LVAD support at 90 days postanddom mizat izat atio ionn. Ventricular io Ventricular en function was quantifi fied fi ed by LVEF ass ses e sedd by transthoracic randomization. quantified assessed ec choocardiogram am (T TTE) TTE) E w hile hi lee w eaane nedd fr fro om LVAD LVAD D support, supppo port rt,, in n tthose hose hos se ppatients atieent ntss able able tto o be b w ean ea ned ned echocardiogram (TTE) while weaned from weaned for fo or 30 minutes. min inut utees. ut es Second nd dar a y en endp d oi dp o nt ntss fo fforr pa ppatients tien ti en nts w ho ttolerated o er ol eraate tedd we wean anin an in ng in iincluded clud cl uded ud ed eechocardiographic c oc ch ocar ardi ar diog di ographic og Secondary endpoints who weaning assessments of myocardial size and function, 6MW, and duration of wean, all assessed while weaned from support, at multiple time points (30, 60 and 90 days post-randomization, and every 60 days thereafter until cardiac transplantation or 12 months, whichever comes first) over the 12 months of the trial follow-up. Additional endpoints included the incidence of serious adverse events, anti-HLA antibody sensitization, neurocognitive outcomes, survival to transplantation, and exploratory mechanistic assessments including histological assessments of myocardium at explant (at cardiac transplantation, LVAD replacement or autopsy), peripheral blood cell phenotypic and functional analyses, and plasma chemo/cytokine analyses at multiple time points 7 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 over the 12 months of trial follow-up. Statistical Analysis Safety A safety monitoring plan was based on pre-specified rare events and mortality. Enrollment would be halted if any of the pre-specified events associated with experimental treatment (infectious myocarditis, myocardial rupture, neoplasm, hypersensitivity reaction, or immune sensitization) were observed within 90 days post-randomization. Similarly, randomization would be halted if, after 10 patients were randomized, the posterior probability that mortality on active therapy was increased compared to control exceeded 80%. Rates of adverse events were compared using Poisson regression. Efficacy Superiority MPC assessed Bayesian Su upe peri rioorit ri orit ityy of o M PC compared to control was ass ssses e sed using a Ba aye y siian approach. approach. The posterior probability successes (ability LVAD wean minutes prob bability that at tthe hee pproportion ropo port rtio rt ionn of su io succ cces essses (a (ab bilitty ty to ttolerate olerat oler a e LV at VAD AD w ean fo ean forr 300 m i ut in utees es aatt 990 0 days group greater the da ys ppost-randomization) o tos t-ra rand ra ndom om mizat atiionn) n) iin n th tthee MP MPC C gr grou oupp wa ou wass gr gre eate t r th te than an n tthe hee pproportion ropo ro po orttio ionn of ssuccesses ucccess uc sssess iin n th he wass ca calculated observed successes control groupp wa w calc lccul u atted bbased ased as e oon ed n th thee ob obse serv se rved rv ed d pproportions ro opo port r io rt ions ns ooff su succ cces cc esse es sess in tthe se he ttwo w groups. wo A non-informative prior distribution, beta (1,1), was assumed for the true success probabilities in the two treatment groups. Categorical variables were summarized as frequencies, continuous variables as means and standard deviations. As per the protocol, LVEF was evaluated only in patients who tolerated the LVAD wean. Fisher’s Exact Test and Wilcoxon sum-rank test were used to compare the two groups when the sample size permitted. Kaplan-Meier curves and the log-rank test were used to assess survival in the two groups. Sample size was determined based on simulations. A sample size of 30 patients was 8 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 chosen to allow us to detect an approximate tripling of the odds that active therapy is superior (i.e., from 50% probability of active therapy’s superiority, or 1:1 odds, to 75% or 3:1 odds) with probability 75% or more if the absolute probability of a successful outcome with active therapy is about 10-15% higher than for control. Results Patients Eighty one patients were screened, 47 eligible, and 30 were randomized (Figure 1); 20 patients to intramyocardial MPC administration and 10 to intramyocardial injection of cryoprotective was medium (control). Treatment intervention was withheld from one MPC patient who who wa as randomized andomized prior to obtaining core lab results off the panel of reactive antibodies (PRA) to exclude pre-existing ex xcl clud udee pr ud ree exissti tinng ng donor-specific antibodies. The The treatment trea eatm tmen tm entt groups grroups oups were wer eree similar simillar with sim witth respect reesppectt to to baseline bassel elin in ne characteristics char ch arracte teri risstic ri sticss (Table (Ta Tab ble 11). ). T he mean was years (±13.6) male. LVEF 18% me ean aage ge w as 557.4 as 7.44 ye ear arss (± (±1 13.6) 13 6) an andd 83% 83% we were re m ale l . Th Thee me mean an L V F was VE was 18 8% (± 44.3), .3)), ), 337% 7% with HeartMate had ischemicc cardiomyopathy, car a di diom omyo om yopa yo paath thy, y and and n all all l patients pattie ient ntss were nt were r implanted imp pla lant nted nt ed w ithh He Hear artM ar tMat tM atte II I ®L LVADs V Ds VA (Thoratec Corp.), 67% were implanted for DT indication. Safety and Mortality No patients developed a primary safety event within 90 days following randomization (the primary endpoint), or during the 12-month follow-up period. At 90 days, there were three deaths (30%) in the control group and none in the MPC group. In a post-hoc analysis, the posterior probability that mortality at 90 days is reduced in the MPC group exceeded 80%. Six MPC patients (30%) died over the 12 months, with no additional deaths occurring in the control group. Figure 2 depicts the Kaplan-Meier survival curves. The most frequent primary causes of death in 9 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 this patient population were LVAD failure (22.2%) and multi-system organ failure (22.2%); and the most frequent underlying causes of death were pump thrombus (33.3%) and sepsis (22.2%). Causes of death were similar between the groups, and no deaths were classified as related to the study intervention. Serious adverse event rates were similar between the two groups (Table 2). At 90 days the serious adverse event rate was 1.30 per patient-month in the treatment group and 1.16 in the control group. The overall rate was 6.95 per patient-year in the treatment group and 6.89 in the control group. The most prevalent serious adverse events over the course of the trial were major bleeding, respiratory failure, right heart failure, and localized non-device infection. The major bleeding event rate per patient-year at 12 months was 3.88 in the treatment group upp and n 33.97 nd .997 in tthe he control group; major bleeding requiring surgery or re-hospitalization is depicted in Table 2. Of note, no otee, one one trial tria tr iall patient, ia paati tieent, en who did not receive intramyocardial intram myo ocardial injections inject n tio ions aatt the th time of LVAD implantation mplantation pl (described (de d sc scribe bedd above), be abov ab ove) ov e),, had e) haad mul m multiple ultiplle bleeding bleeedi edingg eevents venntss an ve andd re rece received ceiv i ed 332 iv 2 un unit units itss of ppacked it acked acke ed red cells ce ell llss ov over er tthe he ccourse ouurse ooff tthe he ttrial. r all. Do ri Dono Donor-specific norr sp rspec eciffic H ec HLA LA ssensitization LA ensi en siti si t za zati tion onn ddeveloped ev vellop oped e pposted osst-randomization andomizatio on in two two w MPC MPC C and and in in three th hre reee control cont co n ro nt roll patients. pati pa t ent ti nts. s. By By one one year, yeear ar,, one one sensitized s ns se nsit itiz it ized iz ed ppatient atient in each arm died and all donor-specific antibodies had resolved in surviving patients. Efficacy In the MPC group, 50% of patients were able to successfully tolerate the wean from LVAD support for 30 minutes at 90 days, compared to 20% in the control group (p=0.24). Based on these results, the posterior probability that MPCs increase the likelihood of successful weaning is 93% (Figure 3). The duration of temporary LVAD wean, for those who tolerated it, was greater in MPC than control patients at each time point (Figure 4). None of the control patients and four (20%) of the MPC treated patients were able to tolerate the LVAD wean at the 30-day time 10 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 point. The mean LVEF at the conclusion of the temporary wean, for those who tolerated LVAD turn down, was 24% (MPC; n=10) and 22.5% (Control; n=2) at 90 days (p=0.56), and the median 6MW was 883 (Q1, Q3 [first and third quartiles] 750, 1042) feet in the treatment arm and 1080 (Q1, Q3 871, 1289) feet in the control arm (p=0.35). At 12 months, there was no difference between groups in the ability to tolerate temporary weaning; 30% of MPC and 40% of control patients (p=0.69) weaned from LVAD support. Eighty-five percent (85%) of MPC patients tolerated one or more temporary LVAD weans over the 12 month follow-up period, compared to 40% of control patients (p=0.03) (Figure 5). Importantly, mportantly, heart failure therapy, including angiotensin receptor and aldosterone nee aantagonists, ntag agon onis on ists is t, ts beta blockers, diuretics, and inotropic therapy, was similar betweenn the two groups at 90 days. At A one on ne year, yeaar, ye ar the the regimens regi gim gi me remained similar across ggroups mens r ups with the eexception ro x ep xc pti tion on of angiotensin antagonists an ntaago g nists (M (MPC: MPC P : n= n=14 144 [[100%]; 100% 100% 0%]; ]; ccontrol: ontrol: n= ont n=4 =4 [57% [57%]). 7%]).. 7% Hospitalizations Ho osp spit ital it aliz izzat atiions ionss There was no o ddifference i fe if fere renc re ncee be nc between etw tweeen gr groups rou o ps w with ithh re it rrespect spec sp ectt to ec o hhospitalizations. ospi os pita pi tali ta l zaati li t on onss. Th Thee me medi median dian di an length of stay of index hospitalization was 29.5 days in the MPC and 35.0 days in the control group (p= 0.91). By 90 days post-randomization, 26 patients had been discharged from the index hospitalization; of those 22% (4/18) of MPC patients and 38% (3/8) of control were readmitted. The rate of re-hospitalization per person-year was 2.15 (MPC) and 2.14 (control). The median time to first readmission was 91 days (Q1, Q3 44, 263) in the MPC group and 51 days (Q1, Q3 10, 150) in the control group. The most frequent reasons for readmission were noncardiovascular in both groups, driven by infection (8.8%) and bleeding (70.6%); 96% of the latter were gastrointestinal in origin (Table 3). 11 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Discussion This is the first randomized trial of allogeneic MPCs in patients undergoing LVAD implantation for the management of advanced heart failure.20-22 Early experience with mesenchymal stem cells, or their subpopulations, suggests that they may be more effective than unfractionated bone marrow mononuclear cells in clinical applications. MPCs are multipotent cells with extensive proliferative potential that secrete numerous antiapoptotic, angiogenic factors, and growth factors.23-24 Since MPCs are immune privileged, they can be transplanted into unrelated recipients without the need for HLA matching or immunosupression, thereby creating the possibility of an allogeneic, off-the-shelf cell product, readily available for administration.13,15,25 The predominant mechanism of MPC therapy in cardiovascular disease is generally genera raall llyy considered co ons nsid ider id ered er ed to o be mediated by the paracrine effects of the cells, since both long-term engraftment and transdi differentiation ifffer eren enti en tiat ti atio at ionn into io in nto cardiomyocytes cardiomyocytes are unlikelyy based bassed on previous previou us studies; stud ud die ies; s neither mechanism 15,26-28 ,2626-2 28 28 can for biological ca an ac aaccount count fo or th tthee bi biol olog ogic og icaal ic al aactivity ctiv ct ivit iv i y ddemonstrated it emoonsstrattedd in n nnumerous um merrou ouss stud sstudies. tud die i s.15, IIndeed, ndee nd e d, ee d MPCs MPC PCss are arre kn know known o n to ssecrete ow eccreete ssignificant ig gni nifi f caant aamounts m un mo ntss ooff po potentially ote t ntiiallly re rele relevant leva le vaant n ggrowth rowt ro w h and wt and an angi angiogenic g og gi ogeenic icc ffactors, acto ac to orss, such uch as stromal stroma maal cell–derived ceell ll–d –d der erived ed factor-1, faccto torr-1, 1 hepatocyte hep epat atoc at ocyt oc y e ggrowth yt ro owtth fa fact factor-1, ctor ct or-1 or -1 1, in insu insulin-like suli su linli n li nlike kee ggrowth rowt ro wthh factor-1, wt vascular endothelial growth factor, and interleukin-6.25,28 Mechanistic effects of the MPCs employed in this trial will be examined in further biospecimen analyses. The majority of patients enrolled in this trial had non-ischemic cardiomyopathy, which, although atypical for the epidemiology of the broader heart failure population, is representative of the breakdown by etiology of the advanced heart failure LVAD subgroup.4,29 Nearly 70% of trial patients received an LVAD for long term use, with approximately a third of the patients receiving LVAD support for BTT. The BTT population in particular, influenced the dose selection for this trial, since these patients are in general younger and may be more likely to 12 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 experience myocardial recovery, but also are at unique risk of jeopardizing their transplant eligibility if they become immune sensitized. For this reason, we selected a dose of 25M cells, a comparatively low dose relative to other trials of cell based therapies. This early trial demonstrates that MPCs are safe; no primary safety endpoint adverse events occurred within 90 days after randomization or over the course of the 12 month followup. As observed, one of the major safety concerns in the BTT LVAD population is HLA sensitization. Interestingly, more control patients developed donor-specific antibodies (DSA) within the first 90 days post-randomization than those who received MPCs, and by one year all donor-specific sensitization had resolved. All three control patients who developed DSA received eceived transfusions following randomization, perhaps contributing to the sensitization. sensit ittizat izat a io on. n. No No sensitization ensitization that developed during the trial was associated with any clinical findings. These data provide pr rovvid idee su sufficient uff ffic icient ic nt ssafety a ety experience to explore hi af high higher gher doses of MP gh MPCs PCs iin n th this vulnerable patient population. pop pu pulation. Serious Seri Se riou ri o s adv ou aadverse dvers versse ev eevent entt ra en rates ate tess at at 990 0 da day days ys aand nd ove oover ver er oone ne yyear earr we ea were r ssimilar re im mil ilar ar bbetween etwe et ween we nM MPC PC an and nd nd control groups. groups ps. Furthermore, ps Furt Fu rthe rt herm he rm morre, adverse adv dveersse ev even event entt ra en rate rates tess ge te gene generally n raall ne llyy we were re ssimilar, im millar ar,, ex exce except c pt ffor ce or bbleeding, l eding, to le those previously reported for the LVAD population.2,4,30 Major bleeding was adjudicated more conservatively in this trial than is current practice. In this trial bleeding events were defined by transfusion requirements at 24 hour increments, regardless of the presence of a clinical bleeding episode, and an ongoing bleed over time was captured as multiple events based on transfusions per time period. This categorization may have contributed to the higher rate of major bleeding observed in this trial relative to the published literature. Nearly 75% of bleeding events occurred more than 30 days following LVAD surgery and intramyocardial injections. Respiratory failure, although theorized to be associated with trapping of cells in the lungs in the setting of cell-based 13 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 therapies, was experienced at similar rates across the treatment groups in this trial, and was consistent with existing benchmarks in LVAD patients.30-31 Direct comparison of suspected and confirmed device thrombus rates seen in this small population with those in the literature is challenged by differences in definitions and duration of LVAD support. That said, the 90 day and 12 month rates observed in the trial population are not dissimilar to those recently reported,32 and are the same between treatment groups. (See Supplemental Table 1 for 30 day AE rates). The median length of stay for the index hospitalization was similar between the two groups, and although somewhat prolonged, was not dissimilar to length of stay data previously reported in the LVAD population.33 The frequency of readmissions by both 90 days and one year post-randomization for patients discharged after their index hospitalization was aalso lso si lso similar imi mila larr la between the treatment groups, and consistentt with the expected range for the LVAD population. po opu pula lati la tion ti on n.34 Interestingly, Intter ereestingly, es although the rate of re read readmissions dmissions wass ssimilar im millar between the treatment ggroups, rouups p , the median medi me dian di n time tim imee too first fir irsst st readmission rea eaddmis dmisssionn was was earlier earlieer er in in the t e control th co ontro roll group groupp (51 grou (51 days dayys da ys [10, [100, 150]) 1500]) 15 than han iin n th thee tr trea treatment eatm tm ment ent ggroup rooup oup (9 (91 91 da days ys [[44, 44, 26 44, 263] 263]). ]). T The hee m most osst comm ccommon omm mmon onn ccause au usee of of readmission, reead a mi misssio io on, similar sim milar i la to o what has been beeen shown sho hown wn previously pre revi viou vi ou usl slyy in i continuous con onti tinu ti nuou nu ouss flow ou f ow fl o device dev evic icee trials, ic tria tr i lss, was ia was ga gast gastrointestinal stro roin ro inte in test te stin st inal bleeding.35-41 The 90-day mortality rate was 30% in the control group and zero in the MPC group. In a post-hoc analysis, the posterior probability that mortality at 90 days is reduced in the MPC group exceeded 80%. At one year, the mortality rates were the same for both groups (30%). Factors known to increase mortality in LVAD recipients, such as prior cardiac surgery, history of stroke, diabetes, and dialysis, were balanced across the treatment groups and within the expected range for the advanced heart failure population, and no patient received a right ventricular assist device implantation at the time of surgery.30,38-41 14 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Despite the low dose of cells deployed in this trial, a likely efficacy signal was observed; a greater proportion of MPC patients experienced successful temporary weans at 90 days. Moreover, the total number of temporary weans tolerated by MPC patients was double that of the control group. Similarly, the significantly lower early mortality rate and fewer hospitalizations in MPC patients compared to control are promising. However, the treatment effect was not seen at one-year. This argues for evaluating higher doses in this population, especially since sensitization concerns were addressed at the lower dose. Consideration also should be given to re-dosing after 90 days to determine whether this might improve the durability of a treatment effect. Re-dosing by systemic intravenous infusion is being employed in another trial of the same MPCs in a non-cardiovascular application. This trial has several limitations. It is a small, exploratory trial; the efficacy endpoints such uch h as as weaning, weean a ing, g, LVEF, LVE V F, and 6MW at 90 days are arre based based on a com comparison mpa p riiso sonn of 10 patients in the MPC MP C and 2 patients paatiient ents in in the th he control cont co ntro nt roll group, grou gr ouup, limiting lim mittingg the th he insight insigh ins sigh ht that thaat can th can be be drawn. draw dr awn. aw n. A larger lar a ge gerr followfoll foll l owup trial triial is is being bein be ingg planned. planne pla annedd. d. In In addition, a di ad diti tion ti on,, efficacy on effi effi fica cacy ca c endpoints, cy e dpo en dpoinnts, nts, such suchh ass functional fun unct cttio ionnal nal status s at st a uss and and rereehospitalizations, hospitalizatio ons ns,, as ooften ften ft en n uused sedd in se n oother th her hheart eart ea rt ffailure ail i ur u e tr tria trials, ials ls,, ar ls aree no nott st straightforward tra raig ight ig htfo ht forw fo r ar rw ardd wi w within t in the th context of LVAD support.42-43 We selected an efficacy endpoint that combines tolerance of LVAD weaning (without symptoms of cardiovascular compromise) with additional assessment of functional status. However, the ability to wean also is affected by non-cardiovascular factors, such as debilitation secondary to co-morbidities or inability to optimize anticoagulation, which, in turn, is critical to safely turning down the pump speed. These factors are unrelated to the intervention and in a small trial may impact a potential efficacy signal. LVADs offer a unique “clinical laboratory” for evaluation of adjunctive cardiac regenerative therapies. The cells used in this trial have many potential advantages including, “off 15 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 the shelf” availability and the potential for immunologic privilege. This exploratory trial confirms feasibility and safety, and suggests early efficacy of MPCs, opening up the field for further clinical investigation. Acknowledgments: The investigators acknowledge the scientific leadership of Dr. Sonia Skarlatos whose contributions to this trial and the clinical investigation of cell-based therapies were substantial. Funding Sources: Trial support was through cooperative agreements (U01 HL088942, HL088957, HL088951, HL088955, HL088939, HL088953) funded by: National Heart Lung and ), the National Institute of Neurological g Blood Institute ((NHLBI), Diseases and Stroke of the NIH,, Bethesda, MD, and the Canadian Institutes for Health Research. This trial was conducted condu d ct du cted ed by by the th Cardiothoracic Surgical Trials Network (CTSN) in collaboration with the Cardiovascular Cell Therap apyy Research Network. Preliminary developm p ent of the MPC-cell line was supported by an Therapy development NH HLB LBII gr ran nt (H HL0 L077 7 09 096) 6).. Inve vestig igatiion nal a pro odu duct c (for in ct inve v stig gat atiiona nall use) us was was pprovided rovi v de d d byy NHLBI grant (HL077096). Investigational product investigational M essoblast, so Inc c. Mesoblast, Inc. Conf nfli lictt of of Interest In nte tereest Disclosures: Discl clos o ur ures es: A utho ors rreport epor ep ortt thee fo foll llow owin ing: g D .D. As .D A chei ch e m, m nnone; on ne; E.. Conflict Authors following: D.D. Ascheim, Bagi g ella, no one ne;; T ewey ew ey, no non ne;; E. E E icchh hhor orrn, nnone; o e; on e; D Feld ldma ld man, ma n T hora ho rate ra tecc Inc. te IInc., nc , He Hear arttWare, Inc. ar Bagiella, none; T.. D Dewey, none; Eichhorn, D.. Fe Feldman, Thoratec HeartWare, consultant; A.C. Gelijns, none; D. Goldstein, Medical advisory board, Thoratec Inc.; Consultant, Heartware Inc.; N. Jeffries, none; C.T. Klodell, none; S. Lee, none; J. Lynch, none; C. Milano, Thoratec Inc., HeartWare Inc. consultant; M.A. Miller, none; Y. Naka, Thoratec Inc. consultant; P.T. O’Gara, none; J.E. Rame, Thoratec, Inc., Heartware Inc. research funding; J.G. Rogers, Thoratec consultant; E.A. Rose, Board of Directors, Mesoblast, Inc.; R. Simari, none; N. Smedira, none; L.A. Moye, none; M.K. Parides, none; D. Skerrett, Chief Medical Officer, Mesoblast, Inc.; B. Sun, Thoratec Inc., Sunshine Heart consultant; A. Szady, none; W. TaddeiPeters, none; D.A. Taylor, Miromatrix Medical Inc, founder; Y.J. Woo, none 16 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 References: 1. Rose EA, Gelijns AC, Moskowitz AJ, Heitjan DF, Stevenson LW, Dembitsky W, Long JW, Ascheim DD, Tierney AR, Levitan RG, Watson JT, Meier P, Ronan NS, Shapiro PA, Lazar RM, Miller LW, Gupta L, Frazier OH, Desvigne-Nickens P, Oz MC, Poirier VL; Randomized Evaluation of Mechanical Assistance for the Treatment of Congestive Heart Failure (REMATCH) Study Group. Long-term use of a left ventricular assist device for end-stage heart failure. N Engl J Med. 2001;345:1435-1443. 2. 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Dixon JA, Gorman RC, Stroud RE, Bouges S, Hirotsugu H, Gorman JH 3rd, Martens TP, Itescu S, Schuster MD, Plappert T, St John-Sutton MG, Spinale FG. Mesenchymal cell transplantation and myocardial remodeling after myocardial infarction. Circulation. 2009;120:S220-229. 28. Williams AR, Hare JM. Mesenchymal stem cells: biology, pathophysiology, translational findings, and therapeutic implications for cardiac disease. Circ Res. 2011;109:923-940. 29. Slaughter MS, Pagani FD, McGee EC, Birks EJ, Cotts WG, Gregoric I, Howard Frazier O, Icenogle T, Najjar SS, Boyce SW, Acker MA, John R, Hathaway DR, Najarian KB, Aaronson KD; HeartWare Bridge to Transplant ADVANCE Trial Investigators. HeartWare ventricular assist system for bridge to transplant: combined results of the bridge to transplant and continued access protocol trial. J Heart Lung Transplant. 2013;32:675-683. 19 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 30. Kirklin JK, Naftel DC, Kormos RL, Stevenson LW, Pagani FD, Miller MA, Timothy Baldwin J, Young JB. Fifth INTERMACS annual report: risk factor analysis from more than 6,000 mechanical circulatory support patients. J Heart Lung Transplant. 2013;32:141-156. 31. Everaert BR, Bergwerf I, De Vocht N, Ponsaerts P, Van Der Linden A, Timmermans JP, Vrints CJ. Multimodal in vivo imaging reveals limited allograft survival, intrapulmonary cell trapping and minimal evidence for ischemia-directed BMSC homing. BMC Biotechnol. 2012;12:93. 32. Starling RC, Moazami N, Silvestry SC, Ewald G, Rogers JG, Milano CA, Rame JE, Acker MA, Blackstone EH, Ehrlinger J, Thuita L, Mountis MM, Soltesz EG, Lytle BW, Smedira NG. Unexpected abrupt increase in left ventricular assist device thrombosis. N Engl J Med. 2014;370:33-40. 33. Slaughter, MS, Bostic R, Tong K, Russo M, Rogers JG. Temporal changes in hospital costs for left ventricular assist device implantation. J Card Surg. 2011;26:535-541. 34. Smedira NG, Hoercher KJ, Lima B, Mountis MM, Starling RC, Thuita L, Schmuhl Schm hm muh u l DM DM,, Blackstone EH. Unplanned hospital readmissions after HeartMate II implantation implantation. Heart on n. JACC J CC H JA e rt ea Fail. 2013;1:31-9. 35. Forest SJ, SJ, Bello R, Friedmann P, Casazza D,, Nucci C, Shin JJ, D'Alessandro D, Stevens G, Goldstein and days out of Gold Go ldst ld stei st einn DJ. ei DJ. Readmissions Reead admi m ssions after ventricular assist asssis i t device: etiologies, etiolog oggies, s, patterns, patterns, at hospital. 2013;95:1276-1281. ho osppit ital. Ann Ann Thorac Thor Th orac or ac Surg. Sur u g. 20 2013 13 3;9 ;95: 5:12 1276 76-1 -128 281. 1. 36. Boilson BA, Clavell 36 6. Hasin Hasin T,, Marmor Marrmor Y, Kremers Kre reemerss W, W, Topilsky Topilsskyy Y,, Severson To Sev ever erso er sonn CJ, so CJ, Schirger Scchirg gerr JJA, A, B o lsson oi nB A, C lavvel la AL, Rodeheffer RJ, Frantz Edwards Pereira Stulak L,, Da R,, Pa Park SJ, AL L, Ro Rode dehe heff he ffer er R J, F J, ranntz ra ntz RP, RP Ed Edwa ward rd ds BS BS, Pe Pere reeirra NL NL,, Stul S tul ulak a JM, ak M, JJoyce oyce oy ce L Daly ly R ark S J, Kushwaha SS. Readmissions after implantation axial ventricular Kush shw waha ha S S.. R eaadm dmissi sion o s af afte terr implan anta tati tion on ooff ax xia iall fl flow ow lleft eftt ve vent ntri r cu ula l r as aassist sist ddevice. si evicce. ev e J Am m Cardiol. Coll Cardiol l. 2013;61:153-163. 201 0 3; 3;61 6 :1 61 153 3-1 -163 6 . 37. Stulak JM, Lee D, Haft JW, Romano MA, Cowger J, Park SJ, Aaronson KD, Pagani FD. Gastrointestinal bleeding and subsequent risk of thromboembolic events during support with a left ventricular assist device. J Heart Lung Transplant. 2014;33:60-64. 38. Lazar JF, Swartz MF, Schiralli MP, Schneider M, Pisula B, Hallinan W, Hicks GL Jr, Massey HT. Survival after left ventricular assist device with and without temporary right ventricular support. Ann Thorac Surg. 2013;96:2155-2159. 39. Atluri P, Goldstone AB, Fairman AS, MacArthur JW, Shudo Y, Cohen JE, Acker AL, Hiesinger W, Howard JL, Acker MA, Woo YJ. Predicting right ventricular failure in the modern, continuous flow left ventricular assist device era. Ann Thorac Surg. 2013;96:857-863. 40. Cowger J, Sundareswaran K, Rogers JG, Park SJ, Pagani FD, Bhat G, Jaski B, Farrar DJ, Slaughter MS. Predicting survival in patients receiving continuous flow left ventricular assist devices: the HeartMate II risk score. J Am Coll Cardiol. 2013;61:313-321. 20 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 14. Wever-Pinzon O, Drakos SG, Kfoury AG, Nativi JN, Gilbert EM, Everitt M, Alharethi R, Brunisholz K, Bader FM, Li DY, Selzman CH, Stehlik J. Morbidity and mortality in heart transplant candidates supported with mechanical circulatory support: is reappraisal of the current United network for organ sharing thoracic organ allocation policy justified? Circulation. 2013;127:452-462. 42. Drakos SG, Wever-Pinzon O, Selzman CH, Gilbert EM, Alharethi R, Reid BB, Saidi A, Diakos NA, Stoker S, Davis ES, Movsesian M, Li DY, Stehlik J, Kfoury AG; UCAR (Utah Cardiac Recovery Program) Investigators. Magnitude and time course of changes induced by continuous-flow left ventricular assist device unloading in chronic heart failure: insights into cardiac recovery. J Am Coll Cardiol. 2013;61:1985-1994. 43. George RS, Yacoub MH, Tasca G, Webb C, Bowles CT, Tansley P, Hardy JP, Dreyfus G, Khaghani A, Birks EJ. Hemodynamic and echocardiographic responses to acute interruption of left ventricular assist device support: relevance to assessment of myocardial recovery. J Heart Lung Transplant. 2007;26:967-973. 21 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Table 1. Baseline Characteristics of the Patients* Characteristic Male Age (yr) Race White Black or African American Cardiomyopathy Ischemic Non Ischemic Biventricular Pacemaker Pre-op IABP Creatinine (mg/dL) Cardiac Index (I/min/m2) PVR (Wood Units) LVEF (%) NYHA Class I & II† Class C lass la ss III III Class C lass IV la V Medications M eddications Beta B et Blocker eta Blocke keer A Aldosterone ldos ld oste os tero te ronne ro ne R Receptor eccep epttor tor An A Antagonist taagoni nist ist ACEi/ARB A CEi/ CE i/AR i/ ARB AR B Intropic Therapy Th her erap apyy ap 1 Agent >1 Agent Vasoactive Therapy Indication for LVAD Bridge to Transplantation Destination Therapy MPC (N=20) 17 (85) 55.1 ± 15.4 Control (N=10) 8 (80) 62.2 ± 7.8 14 (70) 6 (30) 8 (80) 2 (20) 7 (35) 13 (65) 11 (55) 3 (15) 1.3 ± 0.4 1.8 ± 0.6 2.7 ± 2.0 17.5 ±3.9 4 (40) 6 (60) 7 (70) 0 (0) 1.2 ± 0.3 1.8 ± 0.5 3.2 ± 1.8 3.2 1.8 19.3 19 .3 ± 55.1 .11 0 (0) 3 (15)) 17 (85 (85) 855) 1 (10) 2 (20) 7 (7 (70) 0) 1144 (70) (700) 1 (55) 11 (555) 7 (3 (35) 5) 5 (5 (50)) 5 (5 (50) 0) 7 (7 (70) 0) 10 (50) 4 (20) 4 (20) 7 (70) 2 (20) 0 (0) 7 (35) 13 (65) 3 (30) 7 (70) *Plus-minus values are means ± SD, categorical values are n (%); †One patient classified as NYHA II heart failure 22 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Table 2. Serious Adverse Events 90 Days MPC (N=20) Control (N=10) (Person Month=59.2) (Person Month=25) No. of Events (Rate/Pt Month) Bleeding Intra-operative Bleeding Major Bleeding Requiring Surgery Major Bleeding Requiring Re-Hosp Cardiac Arrhythmias Cardiac arrest Sustained ined ventricular arrhythmia* Sustained ined supraventricular arrhythmia† Pericardial rdial Effusion‡ Devicee Malfunction Pump p Failure Non-Pump Pump Failure Pump p Thrombus suspected Pump p Thrombus Thr hrom ombu bus us confirmed c nf co nfir irme m d Hemolysis ly ysi siss Hepaticc Dysfunction D sf Dy s un unct c ion Major In Infect Infection n ct c io i n Localized lized i d Non-Device Inf Infection nfec ecti tion on Internal nal a P Pump u p Component Infl um Inflow f ow fl w or Outflow ow Tr T Tract a t Infectio ac Infection on Sepsiss Unspecified eciffie ied d Neurological log gic ical al Dys Dysfunction y fu func nction TIA Ischemic mic Stroke Hemorrhagic h i St Stroke k Toxic Metabolic Encephalopathy Other Renal Dysfunction Acute Chronic Respiratory Failure Right Heart Failure Venous Thromboembolism Other Total P-value 12 Months MPC (N=20) Control (N=10) (Person Year=18.3) (Person Year=7.6) No. of Events (Rate/Pt Year) 2 ( 0.03) 7 ( 0.12) 0 ( 0.00) 1 ( 0.04) 1 ( 0.04) 4 ( 0.16) 2 ( 0.11) 7 ( 0.38) 10 ( 0.55) 1 ( 0.13) 1 ( 0.13) 7 ( 0.93) 0 ( 0.00) 2 ( 0.03) 1 ( 0.02) 3 ( 0.05) 0 ( 0.00) 2 ( 0.08) 2 ( 0.08) 0 ( 0.00) 1 ( 0.05) 2 ( 0.11) 2 ( 0.11) 3 ( 0.16) 0 ( 0.00) 2 ( 0. 0.26 0.26) 26)) 2 ( 0. 00.26) .26 26)) 26 0 ( 0. 00.00) 00)) 00 0 ( 0.00) 0 ( 0.00) 0 ( 0.00) 2 ( 0.03) 1 ( 0.02) 0 ( 0.00) 0.00 0 ) 0 ( 0.00) 0 ( 0.00) 1 ( 0.04) 1 ( 0.04) 0.0 ) 0.04 1 ( 0.04) 0.04 .0 ) 0 ( 0.00) 0.00 .0 ) 1 ( 0.05) 3 ( 0.16) 1 ( 0.05) 4 ( 0. 00.22) 22) 1 ( 0. 00.05) 05) 1 ( 0. 00.05) 05) 0 ( 0.00) 0 ( 0.00) 1 ( 0.13) 1 ( 0.13) 2 ( 0.26) 0 ( 0. 0.00) .00 0 ) 3 ( 0.05) 0 05 0. 05) 2 ( 0.08) 0.08 .0 ) 4 ( 0. 00.22) 22)) 2 ( 0. 0.26) .26 2 ) 0 ( 0.00) 0.00 00)) 0 ( 0.00) 0.00 .00)) 1 ( 0. 00.05) 05)) 05 0 ( 0.00 0.00) .0 ) 2 ( 0.03) 0 03 0. 0 ) 0 ( 0.00) 0 00 0. 00) 0 ( 0.00) 0.00 00) 0 ( 0.00) 0.00 00)) 00 3 ( 0. 0.16 0.16) 16)) 1 ( 00.05) .05 0 ) 05 1 ( 0.13 0.13) .13)) 0 ( 00.00) .00 00)) 00 0 ( 0.00) 0.00 0. 00)) 0 ( 0.00) 0.00 0 ) 00 0 ( 0.00) 0 00) 2 ( 0.03) 0 ( 0.00) 0 ( 0.00) 0 00 0. 0) 0 ( 0.00) 0.00 00) 0) 1 ( 0.04) 0 04) 0 ( 0.00) 0 ( 0.00) 1 ( 0. 0.05 0.05) 5) 1 ( 00.05) .05 0 ) 0 ( 00.00) 00) 2 ( 0.11) 1 ( 0.05) 0 ( 0.00) 0 ( 0.00) 1 ( 00.13) 13) 0 ( 0.00) 0 ( 0.00) 4 ( 0.07) 1 ( 0.02) 5 ( 0.08) 3 ( 0.05) 1 ( 0.02) 38 ( 0.64) 77 ( 1.30) 1 ( 0.04) 0 ( 0.00) 2 ( 0.08) 1 ( 0.04) 1 ( 0.04) 8 ( 0.32) 29 ( 1.16) 4 ( 0.22) 1 ( 0.05) 5 ( 0.27) 3 ( 0.16) 1 ( 0.05) 61 ( 3.34) 127 ( 6.95) 1 ( 0.13) 0 ( 0.00) 2 ( 0.26) 3 ( 0.40) 1 ( 0.13) 24 ( 3.18) 52 ( 6.89) 0.60 * P-value 0.96 Sustained ventricular arrhythmia requiring defibrillation or cardioversion; †Sustained supraventricular arrhythmia requiring drug treatment or cardioversion; ‡Fluid or clot in the pericardial space that requires surgical intervention or percutaneous catheter drainage 23 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 DOI: 10.1161/CIRCULATIONAHA.113.007412 Table 3. Hospitalization Experience Index Hospitalization Median Length of stay (Days) Readmissions at One Year No. of patients with 1 readmissions Median time to first readmission (days) Total No. of readmissions (rate per patient year) Reasons for readmission (%) LVAD related Cardiovascular Non-LVAD related Non-CV MPC (N=20) Control (N=10) P Value 29.5 (20.5, 43) 35 (17, 45) 0.91 12 (67) 91 (44, 263) 34 (2.1) 6 (75) 51 (10, 150) 14 (2.1) 8 (24) 3 (9) 23 (68) 1 (7) 2 (14) 11 (79) Figure Legends: Figure CONSORT Figu Fi gure gu re 1. CO CONS SOR ORT T Diagram. D ag Di agram. m Figure Survival. Crosses depict patients. Figu Fi gu ure 22.. Su Surv rviv ival iv al.. Cr al Cros osse os s s de se depi pict pi ct ccensored enso en sore so redd pa re pati tien ti ents en ts. ts Figure 3. Posterior Distribution Analysis Curve. Difference = Difference in response-rates (MPC-Placebo); 93% of the area under the curve is between 0-1, corresponding to the probability that MPCs provide better response than placebo. Figure 4. Duration of LVAD Wean. Figure 5. Number of Successful Weans. 24 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 Assessed for eligibility (n=81) Excluded (n=51) Not meeting inclusion criteria (n=34) Declined to participate (n=10) Other reasons (n=7) Randomized (n=30) (n 30) Allocated Allo Al loca cate ca ted te d to M MPC PC C ((Revascor™) R va Re v scor™) (n=20) Received Rece Re c ived ed d allocated allocat atted intervention intervention (n=19) Did receive Did d not rece ceive allocated ce allo al loca cate ted d intervention inte in terv rven nti tion on ((randomized rand ra ndom omiz ized ed panel antibody before pan nel rreactive eacctiv ea ve an ntitibo body bo dy vvalues al es con alue cconfirmed onfirm med d by core lab; treatment tre eattment nt withheld withh held fo for sa safety afety fety concerns concerns) s) (n=1) (n=1 1) Figure 1 Allocated to contr control trol ol ((cryoprotective c yoprotective medium cr alone) (n=10) (n=10 10) 10 (n=10) Received R ceiv Re ivved ed allocated alllloc ocat ated e intervention ed intter erve ve ent ntio ion io n (n (n=1 =10) 0) Did receive Did no nott re rece ceiv ive e allocated a lo al ocate te ed intervention inte in terv r enti rv entiion ((n=0) n=0) n= 0) Lost to follow follow-up w-u up (n=0) (n=0 (n =0)) Discontinued intervention (n=0) Lost Lo ost to to follow-up follow ow-u w up (n=0) (n=0 (n =0)) Discontinued intervention (n=0) Analyzed (n=20) Excluded from analysis (n=0) Analyzed (n=10) Excluded from analysis (n=0) Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 S Figure 2 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 F Figure 3 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 Figure 4 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 Figure 5 Downloaded from http://circ.ahajournals.org/ at MT SINAI SCHOOL MEDICINE on April 4, 2014 SUPPLEMENTAL MATERIAL (e-Publish) SUPPLEMENTAL TABLE: 30 Day Serious Adverse Event Rates SUPPLEMENTAL METHODS: MPC Supplemental Information SUPPLEMENTAL METHODS: Complete Eligibility Criteria SUPPLEMENTAL METHODS: CTSN/CCTRN LVAD MPC Trial Investigators LVAD MPC Trial SUPPLEMENTAL TABLE: 30 Day Serious Adverse Event Rates Serious Adverse Events at Day 30 MPC (N=20) (Person Month=19.7) Control (N=10) (Person Month=9.1) P-value Bleeding Intra-operative Bleeding 2 ( 0.10) 1 ( 0.11) Major Bleeding Requiring Surgery 4 ( 0.20) 1 ( 0.11) Major Bleeding Requiring Re-Hosp 0 ( 0.00) 1 ( 0.11) 2 ( 0.10) 2 ( 0.22) Sustained supraventricular arrhythmia 1 ( 0.05) 2 ( 0.22) Pericardial Effusion 3 ( 0.15) 0 ( 0.00) Device Malfunction Pump Thrombus confirmed 0 ( 0.00) 1 ( 0.11) Hemolysis 1 ( 0.05) 0 ( 0.00) Localized Non-Device Infection 2 ( 0.10) 2 ( 0.22) Sepsis 1 ( 0.05) 0 ( 0.00) Neurological Dysfunction Toxic Metabolic Encephalopathy 1 ( 0.05) 0 ( 0.00) Renal Dysfunction - Acute 3 ( 0.15) 1 ( 0.11) Respiratory Failure 5 ( 0.25) 2 ( 0.22) Right Heart Failure 2 ( 0.10) 1 ( 0.11) Venous Thromboembolism 1 ( 0.05) 1 ( 0.11) Other 20 ( 1.01) 3 ( 0.33) Total 48 ( 2.43) 18 ( 1.97) Cardiac Arrhythmias Sustained ventricular arrhythmia Major Infection 2 0.43 LVAD MPC Trial SUPPLEMENTAL METHODS: MPC Supplemental Information Allogeneic, immunoselected, ex vivo expanded MPCs are derived from bone marrow mononuclear cells, which are obtained from the posterior iliac crest of healthy human donors. These mononuclear cells are immunoselected, expanded, and cryopreserved to produce a cell bank. Cell banks that meet the quality control (QC) in-process release testing are used for production of the allogeneic MPC product Mesoblast has generated a novel monoclonal antibody (STRO-3), which identifies a unique epitope expressed on the extracellular domain of tissue nonspecific alkaline phosphatase. While STRO-1 is able to isolate essentially all multi-, bi- and unipotential clonogenic stromal elements, STRO 3 is able to selectively isolate a subset of human bone marrow STRO-1 bright cells that contains the multipotential MPCs. These lack the phenotypic characteristics of leukocytes and mature stromal elements, are noncycling, and constitutively express telomerase activity in vivo. This STRO-3 positive MPC population demonstrates extensive proliferation and retains the capacity for differentiation into a range of cell types in vitro. Manufacture of MPCs Bone marrow aspirates are acquired in the United States of America, through contractual agreements with a licensed physician and in accordance with 21 CFR Part 1271 (Human Cells, Tissues, and Cellular and Tissue Based Products). Prospective donors complete a Clinical Bone Marrow Donor Program application, read and sign an informed consent document, sign an authorization for human immunodeficiency virus (HIV) testing, and respond to a health questionnaire. Donors then submit a blood sample for infectious disease testing, complete blood count (CBC), a comprehensive metabolic profile, and blood type system (A, AB, B, or O), Rhesus factor, and human leukocyte antigen (HLA) typing. Donors also complete a physical assessment performed by a program physician. The donor documents and laboratory test results are 3 LVAD MPC Trial reviewed and donor eligibility is determined by the Medical Director. Serum samples are retained to ensure that retrospective testing may be performed as new tests are adopted for donor screening. Screening tests for bone marrow donors are listed in the table below. 4 LVAD MPC Trial Bone Marrow Donor Infectious Disease Testing Agent / Disease Tests Specifications HIV-1, HIV-2 Antibody HIV-1, HIV-2 Nonreactive HIV-1 HIV-1 nucleic acid test (NAT) Negative HBV Hepatitis B surface Antigen (HBsAg) Nonreactive HBV Antibody Hepatitis B core (total) Nonreactive HBV HBV NAT Negative HCV Antibody HCV Nonreactive HCV HCV NAT Negative WNV (West Nile Virus) WNV NAT Negative Syphilis Treponemal specific antibody assay Nonreactive Trypanosoma cruzi Enzyme immunoassay Nonreactive HTLV Types I & II Antibody HTLV I/II Nonreactive CMV Antibody CMV Total Nonreactive EBV (Epstein-Barr Virus) Antibody EBV VCA IgM Nonreactive Health questionnaire No exposure Health questionnaire No exposure Human transmissible spongiform Encephalopathy (TSE), including CreutzfeldtJakob disease (CJD) Communicable disease risks associated with xenotransplantation Production of the MPC product is carried out at a contract manufacturing facility under cGMP conditions. A single-tiered Master Cell Bank/Stock (MCB/MCS) is used; there is no Working Cell Bank. Cell banks 5 LVAD MPC Trial that meet the QC in-process release testing are used for production of product doses. The STRO-3 positive cells are isolated by immunoselection, and then expanded before cryopreservation at 30 x 106 cells/cryovial in ProFreeze® NAO Freezing Medium/Alpha MEM and 7.5% DMSO using a Cryomed Controlled Rate Freezer. Cryopreserved vials are stored in the vapor phase of liquid nitrogen (LN2). Production of the final product begins with the thawing of one cryovial of the MCB/MCS. Cells are seeded and culture expanded in sterile, polystyrene cell factories. At the final harvest of cells, two cell counts are performed and samples are removed for release testing (sterility, endotoxin, mycoplasma, cell count and viability, cell markers, and karyoptype). The remaining cells are pelleted and cells resuspended in 50% Alpha-MEM/42.5% ProFreeze® NAO Freezing Medium/7.5% DMSO at 2°C to 4°C. The final product is then cryopreseved in an appropriate container and stored in the vapor phase of LN2. Storage and Handling Allogeneic MPCs are stored in the vapor phase of LN2 at -196ºC to -140ºC until ready for use. For longterm storage, the product should be maintained in a continuously monitored freezer with adequate security, 24-h temperature monitoring and an audible alarm. Thawed product is to be administered within ninety (90) minutes of its thaw. Further instructions for the handling of MPCs are provided in the individual study operations manuals 6 LVAD MPC Trial SUPPLEMENTAL METHODS: Complete Eligibility Criteria Patients with end-stage heart failure, of either ischemic or non-ischemic etiology, who are being evaluated for LVAD implantation as a BTT or DT, were candidates for this study. Candidates who met all inclusion criteria and no exclusion criteria were eligible for the trial regardless of gender, race, or ethnicity. Inclusion Criteria 1. Signed informed consent, inclusive of release of medical information, and Health Insurance Portability and Accountability Act (HIPAA) documentation; 2. Age 18 years or older; 3. If the subject or partner is of childbearing potential, he or she must be willing to use adequate contraception (hormonal or barrier method or abstinence) from the time of screening and for a period of at least 16 weeks after procedure; 4. Female subjects of childbearing potential must have a negative serum pregnancy test at screening; 5. Admitted to the clinical center at the time of randomization; 6. Clinical indication and accepted candidate for implantation of an FDA approved implantable, nonpulsatile LVAD as a bridge to transplantation or for destination therapy. Exclusion Criteria 1. Planned percutaneous LVAD implantation; 2. Anticipated requirement for biventricular mechanical support; 3. Cardiothoracic surgery within 30 days prior to randomization; 4. Myocardial infarction within 30 days prior to randomization; 7 LVAD MPC Trial 5. Prior cardiac transplantation, LV reduction surgery, or cardiomyoplasty; 6. Acute reversible cause of heart failure (e.g. myocarditis, profound hypothyroidism); 7. Stroke within 30 days prior to randomization; 8. Platelet count < 100,000/ul within 24 hours prior to randomization; 9. Active systemic infection within 48 hours prior to randomization; 10. Presence of >10% anti-HLA antibody titers 1 with known specificity to the MPC donor HLA antigens 2; 11. A known hypersensitivity to dimethyl sulfoxide (DMSO), murine, and/or bovine products; 12. History of cancer prior to screening (excluding basal cell carcinoma); 13. Acute or chronic infectious disease, including but not limited to human immunodeficiency virus (HIV); 14. Received investigational intervention within 30 days prior to randomization; 15. Treatment and/or an incompleted follow-up treatment of any investigational cell based therapy within 6 months prior to randomization; 16. Active participation in other research therapy for cardiovascular repair/regeneration; 17. Prior recipient of stem precursor cell therapy for cardiac repair; 18. Pregnant or breastfeeding at time of randomization. 1 2 Documented by clinical site laboratory Documented by Core Lab 8 LVAD MPC Trial SUPPLEMENTAL METHODS: CTSN/CCTRN LVAD MPC Trial Investigators Clinical Site Investigators: Baylor Health Care System: Todd Dewey (PI), Robert Smith, Mitchell Magee, Eric Eichorn, J. Edward Rosenthal, Tina Worely, Cecile Mahoney, Darinka Savor; Cleveland Clinic Foundation: Nicholas Smedira (PI), Nader Moazami, Edward Soltesz, Randall Starling, Eiran Gorodeski, Eileen Hsich, Sangjin Lee, Mazen Hanna, Maria Mountis, Randall Starling, Wilson Tang, David Taylor, Carrie L. Geither, Kathy Sankovic, Tiffany Buda, Pam Lackner, Patricia Bouscher, Barbara Gus, Susan Moore, Cindy Oblak; Columbia University Medical Center: Yoshifumi Naka (PI), Hiroo Takayama, Allan Stewart, Ulrich Jorde, Lyn Goldsmith, Sowmyashree Sreekanth, Amanda Alonso, Rosie Te-Frey, Daniello Van Patten; Duke University Medical Center: Carmelo Milano (PI), Joseph G. Rogers, Chetan Patel, Joseph Rogers, Stacey Welsh, Victoria Sutto, Laura Blue, Amanda Hynes, Christopher Koller; Minneapolis Heart Institute Foundation: Benjamin Sun (PI), David Feldman, Tim Henry, Jay Traverse, Barry Cabuay, Kaisa Hryniewicz, Karen Meyer, Lisa Lundquist, Charlene Boisjolie, Carrie Weaver, Jessica Boughton, Elizabeth Carter; Montefiore-Einstein Heart Center: Daniel Goldstein (PI), Ricardo Bello, David D’Alessandro, Joseph DeRose, William Jakobleff, Robert Michler, Julia Shin, Daniel Spevack, Cecilia Nucci, Nadia Sookraj, Roger Swayze; Ohio State University Medical Center: Sai Sudhakar (PI), Robert Higgins, William T. Abraham, Ayesha Hasan, Sherri Wissman, Kelly MacBrair, Anne Knapke, Laura Yamokoski, Asia McDavid; Texas Heart Institute: James Willerson (PI), O.H.(Bud) Frazier, Hari R. Mallidi, William Cohn, Emerson C. Perin, Guilherme Silva, Andrew Civitello, Reynolds Delgado, Deirdre Smith, Jennifer Chambers, Sylvia Carranza, Dia Tisdel-Pickens, Casey Kappenman; University of Florida: Charles T. Klodell (PI), Thomas Beaver, Edward Staples, Anita Szady, Carl Pepine, James Hill, Richard Schofield, Eileen Handberg, Juan M. Aranda Jr., Daniel Pauly, Sarah Long, 9 LVAD MPC Trial Jessica Bell, Jana Reid, Debbie Robertson, Dana D. Leach, Elfrida Prestwood; University of Maryland: Bartley P. Griffith (PI), James Gammie, Gautam Ramani, Erika Feller, Sunjay Kaushal, Dana Beach, Lynn Dees; University of Pennsylvania: Joseph Woo (PI), Michael A. Acker, Atluri Pavan, Kenneth Margulies, J. Eduardo Rame, Joyce Wald, Mary Lou Mayer, Christyna Zalewski, Stephen Cresse, Bhavana Venkataram, Judith Marble, Mary Lou O’Hara; Echo Core Lab, Massachusetts General Hospital: Judy Hung, Xin Zeng; Neurocognitive Core Lab, Duke University: Joseph P. Mathew, Yanne Toulgoat-Dubois, Jeffrey Browndyke. 10