Biomarkers to Improve Prognostication in Heart Failure

Laura C. van Vark

Biomarkers

to improve

prognostication

in heart failure

Thesis lay-out: Ilse Modder | www.ilsemodder.nl Printed by: Gildeprint Enschede | www.gildeprint.nl

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Copyright © 2020 L.C. van Vark

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Biomarkers voor het voorspellen van uitkomsten bij hartfalen

Proefschrift

ter verkrijging van de graad van doctor aan de Erasmus Universiteit Rotterdam

op gezag van de UHFWRUPDJQLȴFXV Prof. dr. R.C.M.E. Engels

en volgens besluit van het College voor Promoties. De openbare verdediging zal plaatsvinden op

donderdag 9 juli 2020 om 09:30 uur

door

Laura Charlotte van Vark

geboren te Zwijndrecht

Promotor: Prof. dr. ir. H. Boersma

Overige leden: Prof. dr. D.J.G.M. Duncker

Prof. dr. Y.M. Pinto

Prof. dr. J.L. Hillege

Co-promotor : Dr. K.M. Akkerhuis

Financial support by the Dutch Heart Foundation for the publication of this thesis is gratefully acknowledged.

&KDSWHUɅGeneral introduction and outline of thesis

PART I: BIOMARKERS IN HEART FAILURE

&KDSWHUɅPrognostic value of serial galectin-3 measurements in  ɅSDWLHQWVZLWKDFXWHKHDUWIDLOXUH

 ɅJ Am Heart Assoc. 2017

&KDSWHUɅPrognostic value of serial ST2 measurements in patients  ɅZLWKDFXWHKHDUWIDLOXUH

 ɅJ Am Coll Cardiol. 2017

&KDSWHUɅReply: Value of serial ST2 measurements in acute heart  ɅIDLOXUHPL51$UHJXODWLRQDQGJHQHWLFIDFWRUV

 ɅJ Am Coll Cardiol. 2018

&KDSWHUɅPrognostic value of serial measurements of multiple  ɅELRPDUNHUVGXULQJIROORZXSRISDWLHQWVZLWKDFXWH  ɅKHDUWIDLOXUH

 ɅSubmitted

&KDSWHUɅSerially-measured circulating microRNAs and adverse  ɅFOLQLFDORXWFRPHVLQSDWLHQWVZLWKDFXWHKHDUWIDLOXUH

 ɅEur J Heart Fail. 2018

&KDSWHUɅLipoprotein-associated phospholipase A2 activity and risk  ɅɄRIKHDUWIDLOXUH7KH5RWWHUGDPVWXG\

 ɅɄEur Heart J. 2006

&KDSWHUɅBiomarkers of heart failure with normal ejection  ɅIUDFWLRQDV\VWHPDWLFUHYLHZ

 ɅEur J Heart Fail. 2013

PART II: HEALTH RELATED QUALITY OF LIFE IN HEART FAILURE PATIENTS &KDSWHUɅSymptoms and depression in acute heart failure  ɅSDWLHQWV  ɅSubmitted 11 23 45 67 71 93 125 143 175

ɅɅSubmitted

PART III: RAAS INHIBITORS AND OUTCOME IN HYPERTENSIVE PATIENTS &KDSWHUɅAngiotensin-converting enzyme inhibitors reduce mortality  ɅɅLQK\SHUWHQVLRQDPHWDDQDO\VLVRIUDQGRPL]HGFOLQLFDO  ɅɅWULDOVRIUHQLQDQJLRWHQVLQDOGRVWHURQHV\VWHPLQKLELWRUV  ɅɅLQYROYLQJSDWLHQWV

 ɅɅEur Heart J. 2012

&KDSWHUɅImpact of renin-angiotensin system inhibitors on mortality  ɅɅDQGPDMRUFDUGLRYDVFXODUHQGSRLQWVLQK\SHUWHQVLRQ$  ɅɅQXPEHUQHHGHGWRWUHDWDQDO\VLV

 ɅɅInt J Cardiol. 2015

&KDSWHUɅSummarizing discussion.

 ɅɄDutch Summary (Nederlandse samenvatting)

 ɅɅ AppendicesɅ/LVWRISXEOLFDWLRQV  ɅɅ3K'SRUWIROLR  ɅɅ$ERXWWKHDXWKRU  ɅɅ'DQNZRRUG 221 245 265 278 289 293 295 296

Chapter 1

Biomarkers and heart failure

Heart failure (HF) is a major and growing public health problem.1 _{Prevalence of HF is}

KLJKHVSHFLDOO\LQWKHHOGHUO\ as about 10% of men and 8% of women over the age of \HDUVDUHD΍HFWHGLQLQGXVWULDOL]HGFRXQWULHV_{0RUHRYHULQWKH5RWWHUGDPVWXG\}

DSRSXODWLRQEDVHGFRKRUWVWXG\WKHOLIHWLPHULVNRIGHYHORSLQJ+)ZDVIRUPHQ and 29% for women at the age of 55.3_{'HVSLWHWKHVLJQLȴFDQWDGYDQFHVLQWKHUDSLHVDQG}

SUHYHQWLRQPRUWDOLW\DQGPRUELGLW\LVVWLOOKLJKDQGTXDOLW\RIOLIHSRRUb

HF is a clinical syndrome characterized by typical symptoms that may be accompanied E\VLJQVFDXVHGE\DVWUXFWXUDORUIXQFWLRQDOFDUGLDFDEQRUPDOLW\UHVXOWLQJLQDUHGXFHG cardiac output or elevated intracardiac pressures at rest or during stress.4 _{HF is}

FRPSOH[LQYROYLQJVHYHUDOXQGHUO\LQJSDWKRSK\VLRORJLFDOSURFHVVHV2_{)XUWKHUPRUH+)LV}

a progressive disease. HF patients are in need of permanent medication use and acute GHFRPSHQVDWLRQ LV FRPPRQ LQ ZKLFK FDVH DGYDQFHG WUHDWPHQW DQG KRVSLWDOL]DWLRQ is necessary. It is essential to improve prognostication in heart failure patients to be able to help physicians diagnose patients early and anticipate in time to intensify heart failure treatment and patient monitoring.5

$ ELRPDUNHU LV D PHDVXUDEOH LQGLFDWRU UHȵHFWLQJ D ELRORJLFDO VWDWH IRU H[DPSOH D VXEVWDQFHPHDVXUHGLQWKHEORRGΖQKHDUWIDLOXUHELRPDUNHUVPD\UHȵHFWPHFKDQLVPV VXFK DV LQȵDPPDWLRQ R[LGDWLYH VWUHVV QHXURKRUPRQDO DFWLYDWLRQ P\RFDUGLDO DQG PDWUL[ UHPRGHOOLQJ5 _{There is growing evidence that biomarkers released as}

a consequence of these underlying pathophysiological processes may not only LPSURYH RXU SDWKRSK\VLRORJLFDO NQRZOHGJH EXW DUH DOVR XVHIXO IRU LGHQWLȴFDWLRQ RI KHDOWK\ SHRSOH ZKR DUH DW ULVN IRU GHYHORSLQJ +) IRU WKH GLDJQRVLV RI +) SDWLHQWV and for improved risk prediction in HF patients.2 _{Circulating microRNAs have been}

SURSRVHG DV DQ DWWUDFWLYH QHZ FODVV RI ELRPDUNHUV EHFDXVH RI WKHLU DVVRFLDWLRQ ZLWK GL΍HUHQW KHDUW IDLOXUH DHWLRORJLHV DQG GLVHDVH SURJUHVVLRQ WKHLU VWDELOLW\ LQ WKH FLUFXODWLRQ DQG WKHLU HQVXLQJ UHOLDEOH DVVHVVPHQW LQ HDVLO\ DFFHVVLEOH VDPSOHV6

'XH WR GL΍HUHQW KHDUW IDLOXUH DHWLRORJLHV GL΍HUHQW XQGHUO\LQJ SDWKRSK\VLRORJLFDO PHFKDQLVPVDQGGLVHDVHSURJUHVVLRQRYHUWLPHRQHVLQJOHELRPDUNHUPHDVXUHPHQW PD\QRWDGHTXDWHO\UHȵHFWWKH+)GLVHDVH)LUVWRIDOOFRPELQLQJGL΍HUHQWELRPDUNHUV LVSURSRVHGWREHXVHIXOIRUSURYLGLQJDGGLWLRQDOULVNVWUDWLȴFDWLRQ5_{6HFRQGO\PXOWLSOH}

biomarker measurements over time are assumed to propose additional information.7

7KLUGO\FHUWDLQELRPDUNHUVPD\EHXVHIXOLQVSHFLȴFKHDUWIDLOXUHSRSXODWLRQVVXFKDV HF with a preserved ejection fraction (HFpEF).8

Health related quality of life in heart failure patients

)URPDSDWLHQWȇVSHUVSHFWLYHTXDOLW\RIOLIHLVYHU\LPSRUWDQW6WXGLHVKDYHIRXQGWKDW patients value quality of life at least as important as longevity._{$PRQJ+)SDWLHQWV}

KHDOWKUHODWHGTXDOLW\RIOLIH+54R/LVZRUVHFRPSDUHGWRWKHJHQHUDOSRSXODWLRQDQG is worse than that of patients with other chronic conditions.10-12_{0RUHRYHUDQLPSDLUHG}

HRQoL is also related to adverse outcome.13

ΖW LV NQRZQ WKDW +54R/ LV LQȵXHQFHG E\ WKH RFFXUUHQFH RI +) V\PSWRPV  _HF

V\PSWRPVDUHPRUHSUHYDOHQWLQGHSUHVVHGSDWLHQWVZKLFKLVDFRPPRQFRPRUELGLW\ in HF patients.16_{7KHUHIRUHPRUHLQVLJKWLVQHHGHGLQWKHUHODWLRQEHWZHHQGHSUHVVLRQ}

DQGKHDUWIDLOXUHV\PSWRPV%HVLGHVV\PSWRPVRIGHSUHVVLRQWKHSUHVHQFHRIQRQ cardiac comorbidities may also impact HRQoL. Determinants related to HRQoL may be LQȵXHQFHGWRLPSURYHKHDUWIDLOXUHWUHDWPHQW

7UHDWPHQWHϝHFWRIUDDVLQKLELWRUVLQK\SHUWHQVLYHSDWLHQWV

The renin-angiotensin-aldosteron system (RAAS) plays an important role in the regulation of hemodynamic stability in the human body by controlling electrolytes DQG ȵXLG EDODQFH 7KH 5$$6 IXQFWLRQV WKURXJK GLUHFW DQG LQGLUHFW H΍HFWV RQ VHYHUDO organ systems and interacts with autonomic nervous system and several vasoactive hormones._{([FHVVLYHVWLPXODWLRQRIWKH5$$6FDXVHVSDWKRORJLFFKDQJHVLQDZLGH}

YDULHW\RIRUJDQV\VWHPV2YHUDFWLYH5$$6LVDVVRFLDWHGZLWKK\SHUWHQVLRQUHQDOLQMXU\ atherosclerosis and left ventricular dysfunction. _{Therefore the blockade of RAAS has}

EHFRPH D NH\ WKHUDSHXWLF WDUJHW LQ D ZLGH YDULHW\ RI SDWLHQWV VXFK DV K\SHUWHQVLYH patients and RAAS blockade is also a cornerstone in treatment of heart failure patients. 7KHFOLQLFDOO\PRVWLPSRUWDQWH[DPSOHVRISKDUPDFRORJLFDJHQWVWKDWEORFNWKH5$$6 currently are the angiotensin-converting enzyme (ACE) inhibitors and AT1 receptor blockers (ARBs).

Reductions in both cardiovascular morbidity and mortality have been well GHPRQVWUDWHG ZLWK 5$$6 LQKLELWRUV DFURVV VSHFLȴF SRSXODWLRQV VXFK DV KHDUW IDLOXUH SDWLHQWV KLJK FDUGLRYDVFXODU ULVN SDWLHQWV DQG VWDEOH FRURQDU\ GLVHDVH SDWLHQWV21-23

ΖQWKHVHWULDOVSDWLHQWVZHUHVHOHFWHGIRUDFULWHULRQRWKHUWKDQK\SHUWHQVLRQSHUVH and so only half of the patients enrolled in these trials had prevalent hypertension. 7KHEHQHȴFLDOH΍HFWVRI5$$6LQKLELWRUVLQK\SHUWHQVLYHSDWLHQWVRQPRUWDOLW\KDYHQRW EHHQFRQYLQFLQJO\GHPRQVWUDWHGDVPRVWWULDOVZHUHXQGHUSRZHUHGIRUWKLVHQGSRLQW

Aim and outline of this thesis

2XUPDLQREMHFWLYHLQWKLVWKHVLVLVWRHYDOXDWHELRPDUNHUVWKDWUHȵHFWWKHXQGHUO\LQJ SDWKRSK\VLRORJLFDO SURFHVVHV LQ +) VXFK DV PHFKDQLFDO RYHUORDG DWKHURVFOHURVLV LQȵDPPDWLRQDQGFDUGLDFȴEURVLV2 _{To take into account the dynamic and progressive}

QDWXUH RI +) RYHU WLPH ZH GHVLJQHG WKH 75DQVODWLRQDO ΖQLWLDWLYH RQ 8QLTXH DQG novel strategies for Management of Patients with Heart failure (TRIUMPH) study.24

TRIUMPH is a multi-centre observational cohort study in acute HF patients. Patients were enrolled when they where hospitalized with decompensation of known chronic +)RUQHZO\GLDJQRVHG+)ΖQ75Ζ803+ELRPDUNHUVZHUHHYDOXDWHGIRUWKHLUSURJQRVWLF properties using a unique design of seven planned repeated measurements during 1-year follow-up. To be able to account for repeated biomarker measurements in our DQDO\VHVMRLQWPRGHOVZHUHXVHG

%DVHG RQ SUHYLRXV FOLQLFDO DQG HSLGHPLRORJLFDO VWXGLHV 67 DQG JDOHFWLQ ZHUH marked as biomarkers with high potential for improving prognostication. _Although

the AHA/ACC Guidelines advise physicians to consider using ST2 and galectin-3 as an DGGLWLRQDOELRPDUNHUWRQDWULXUHWLFSHSWLGHOHYHOVIRUSURJQRVWLFDWLRQLQ+)SDWLHQWV26

the ESC guidelines state that none of the newer biomarkers has reached the stage of being recommended for routine use.4 _{Therefore in chapter two, three and four67}

DQGJDOHFWLQZHUHH[WHQVLYHO\HYDOXDWHG6XEVHTXHQWO\ZHSHUIRUPHGDPXOWLPDUNHU PXOWLWLPH SRLQW DQDO\VHV WR DVVHVV WKH LQGHSHQGHQW SURJQRVWLF YDOXH RI UHSHDWHG PHDVXUHPHQWVRI17SUR%13FDUGLDFWURSRQLQΖ67JDOHFWLQDQGFUHDWLQLQHIRUDOO cause and cardiovascular mortality (FKDSWHUȴYH).

+HQFHZHHYDOXDWHGWKHSRWHQWLDOIRUSURJQRVWLFDWLRQRIVHULDOPHDVXUHGPLFUR51$V )LUVWDQ51$VHTXHQFLQJGLVFRYHU\H[SHULPHQWLQSLJVZDVXVHGWRLGHQWLI\WKHPRVW promising novel microRNA (miRœ1306œS6HFRQGO\PXOWLSOHPL5VNQRZQWREHFDUGLDFœ enriched or previously linked to HF were evaluated (miRœPL5œ22œSPL5œ345œS miRœ378aœSPL5œ423œSPL5œDPL5œ133aœSPL5œEPL5œ499aœSPL5œ622 and miRœ208aœ3p) (chapter six).

(OHYDWLRQRILQȵDPPDWRU\PDUNHUVKDYHDOVREHHQDVVRFLDWHGZLWKDQLQFUHDVHRIWKH development of HF.27 _{Lipoprotein phospholipase A2 (Lp-PLA2) is proposed as a}

pro-LQȵDPPDWRU\PDUNHUDQGLVDQLQGHSHQGHQWSUHGLFWRURIFDUGLRYDVFXODUGLVHDVH28 _We

DVVHVVWKHDVVRFLDWLRQEHWZHHQ/S3/$DSURLQȵDPPDWRU\ELRPDUNHUDQGLQFLGHQW +)LQDUDQGRPVDPSOHRIWKH5RWWHUGDP6WXG\DSRSXODWLRQEDVHGFRKRUWVWXG\DPRQJ persons aged 55 years and over (chapter seven).29 _{In chapter eight ZH UHYLHZHG WKH}

associations of several biomarkers with the occurrence and prognosis of HF with a preserved ejection fraction.

The aim of the second part of this thesis is to evaluate symptom burden and health-related quality of life (HRQoL) determinants in acute HF patients. We evaluated the occurrence DQGEXUGHQRI+)V\PSWRPVLQDFXWH+)SDWLHQWVZLWKDQGZLWKRXWGHSUHVVLRQEDVHG on data from the TRIUMPH cohort (chapter nine6HFRQGO\ZHLQYHVWLJDWHGWKHUHODWLRQ EHWZHHQ SUHVHQFH RI QRQFDUGLDF FRPRUELGLWLHV GLDEHWHV PHOOLWXV FKURQLF NLGQH\ G\VIXQFWLRQ&23'RUSULRU&9$DQG+54R/)XUWKHUPRUHZHHYDOXDWHGGHWHUPLQDQWV RI+54R/LQDFXWH+)SDWLHQWVZLWKDQGZLWKRXWFRPRUELGLWLHVchapter ten).

ΖQWKHȴQDOSDUWRIWKLVWKHVLVZHH[DPLQHGZKHWKHU5$$6LQKLELWRUVUHGXFHDOOFDXVH and cardiovascular mortality in hypertensive patients. We considered RAAS inhibitors DVDFODVVRIGUXJVDVZHOODV$&(LQKLELWRUVDQG$5%VVHSDUDWHO\chapter eleven). In DGGLWLRQZHDVVHVVHGWKHH΍HFWLYHQHVVRI5$$6LQKLELWRUVWRSUHYHQWDOOFDXVHPRUWDOLW\ DQGFDUGLRYDVFXODUGHDWKP\RFDUGLDOLQIDUFWLRQDQGVWURNHLQK\SHUWHQVLYHSDWLHQWV:H performed a ‘number needed to treat’ analyses to give insight to the absolute treatment H΍HFWRI5$$6LQKLELWRUVLQVWHDGRIUHODWLYHULVNUHGXFWLRQDORQHZKLFKLVFRQVLGHUHGDQ important measure to accurately communicate risk (chapter twelve).

)LQDOO\LQFKDSWHUWKLUWHHQDJHQHUDORYHUYLHZDQGGLVFXVVLRQRIDOOUHVXOWVGHVFULEHGLQ this thesis is given.

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24. TRIAL ID: NTR1893. TRanslational Initiative on Unique and novel strategies for Management of Patients with Heart failure (TRIUMPH).

25. 8HODQG7$XNUXVW3%URFK.$DNKXV66NDUGDO50XQWHQGDP3DQG*XOOHVWDG/*DOHFWLQLQKHDUWIDLOXUHKLJK levels are associated with all-cause mortality. Int J Cardiol. 2011;150:361-4.

26. <DQF\&:-HVVXS0%R]NXUW%%XWOHU-&DVH\'(-U&ROYLQ00'UD]QHU0+)LOLSSDWRV*6)RQDURZ*&*LYHUW] 00+ROOHQEHUJ60/LQGHQIHOG-0DVRXGL)$0F%ULGH3(3HWHUVRQ316WHYHQVRQ/:DQG:HVWODNH&$&& AHA/HFSA Focused Update of the 2013 ACCF/AHA Guideline for the Management of Heart Failure: A Report of the American College of Cardiology/American Heart Association Task Force on Clinical Practice Guidelines and the Heart Failure Society of America. J Am Coll Cardiol. 2017;70:776-803.

27. 9DVDQ566XOOLYDQ/05RXEHQR΍5'LQDUHOOR&$+DUULV7%HQMDPLQ(-6DZ\HU'%/HY\':LOVRQ3:'ȇ$JRVWLQR5% DQG)UDPLQJKDP+HDUW6ΖQȵDPPDWRU\PDUNHUVDQGULVNRIKHDUWIDLOXUHLQHOGHUO\VXEMHFWVZLWKRXWSULRUP\RFDUGLDO

infarction: the Framingham Heart Study. Circulation. 2003;107:1486-91.

28. 2HL++YDQGHU0HHUΖ0+RIPDQ$.RXGVWDDO3-6WLMQHQ7%UHWHOHU00DQG:LWWHPDQ-&/LSRSURWHLQDVVRFLDWHG phospholipase A2 activity is associated with risk of coronary heart disease and ischemic stroke: the Rotterdam Study. Circulation. 2005;111:570-5.

29. +RIPDQ$*UREEHH'(GH-RQJ37DQGYDQGHQ2XZHODQG)$'HWHUPLQDQWVRIGLVHDVHDQGGLVDELOLW\LQWKHHOGHUO\ the Rotterdam Elderly Study. Eur J Epidemiol. 1991;7:403-22.

30. *LJHUHQ]HU*:HJZDUWK2DQG)HXIHO00LVOHDGLQJFRPPXQLFDWLRQRIULVNBMJ. 2010;341:c4830.

31. &RRN 5- DQG 6DFNHWW '/ 7KH QXPEHU QHHGHG WR WUHDW D FOLQLFDOO\ XVHIXO PHDVXUH RI WUHDWPHQW H΍HFW BMJ. 1995;310:452-4.

Part I

Chapter 2

3URJQRVWLFYDOXHRIVHULDOJDOHFWLQ

measurements in patients

with acute heart failure

L.C. van Vark, I. Lesman-Leegte, S.J. Baart, D. Postmus, Y.M. Pinto, R.A. de Boer, F.W. Asselbergs, E.M.C.J. Wajon, J.G. Orsel, E. Boersma, H.L. Hillege, K.M. Akkerhuis; TRIUMPH Investigators

$EVWUDFW

Background

Several clinical studies have evaluated the association between galectin-3 levels DQG RXWFRPH LQ SDWLHQWV ZLWK KHDUW IDLOXUH +) +RZHYHU RQO\ OLWWOH LV NQRZQ DERXW the predictive value of repeated galectin-3 measurements. This study evaluates the prognostic value of repeated time-dependent galectin-3 measurements in acute HF patients.

Methods

ΖQ WKH 75Ζ803+ FOLQLFDO FRKRUW VWXG\  DFXWH +) SDWLHQWV ZHUH HQUROOHG LQ  hospitals LQ 7KH 1HWKHUODQGV EHWZHHQ  DQG . Repeated blood samples (seven) were drawn during 1-year follow-up. Associations between repeated biomarker measurements and the primary endpoint were assessed using a joint model.

Results

Median age was 74 years and 37% were women7KHSULPDU\HQGSRLQWFRPSRVLWHRI DOOFDXVHPRUWDOLW\DQG+)UHKRVSLWDOL]DWLRQZDVUHDFKHGLQSDWLHQWVGXULQJ a median follow-up of 325 days (IQR 85-401). Median baseline galectin-3 level was 24 ng/ml (IQR 18-34). The mean number of galectin-3 measurements available per patient was 4.3.

$IWHU DGMXVWPHQW IRU FOLQLFDO IDFWRUV DQG 17SUR%13 WKHUH ZDV D ZHDN DVVRFLDWLRQ between baseline galectin-3 and risk of the primary endpoint. When repeated PHDVXUHPHQWV ZHUH WDNHQ LQWR DFFRXQW WKH DGMXVWHG KD]DUG UDWLR per 1 standard deviation increase of the galectin-3 level (on the log2 scale) at any time point increased to &ΖȂSYDOXH$IWHUDGGLWLRQDODGMXVWPHQWIRUUHSHDWHG 17SUR%13PHDVXUHPHQWVWKHDVVRFLDWLRQUHPDLQHGVWDWLVWLFDOO\VLJQLȴFDQW

Conclusions

Repeated galectin-3 measurements appeared a strong predictor of outcome in acute +) SDWLHQWV LQGHSHQGHQW RI 17SUR%13 +HQFH JDOHFWLQ PD\ EH KHOSIXO LQ FOLQLFDO practice for prognostication and treatment monitoring.

Introduction

Most studies on serum biomarkers in heart failure (HF) populations conducted so IDU KDYH UHODWHG DGYHUVH RXWFRPH GXULQJ IROORZXS ZLWK D VLQJOH PHDVXUHPHQW DW baseline.1-3 _{Although this approach has demonstrated the prognostic value of a variety}

RIELRPDUNHUVDPRQJZKLFKWKHZHOONQRZQQDWULXUHWLFSHSWLGHV4_{LWGRHVQRWH[SORUHWKH}

ELRORJLFDOYDULDWLRQZLWKLQSDWLHQWVZLWKHYROYLQJGLVHDVHΖQIDFW+)LVDKLJKO\YDULDEOH heterogeneous and progressive condition.5_{7KXVUHSHDWHGELRPDUNHUPHDVXUHPHQWV}

PD\EHUHTXLUHGWRPRUHDFFXUDWHO\UHȵHFWWKLVG\QDPLFDQGSURJUHVVLYHQDWXUHRIWKH XQGHUO\LQJSDWKRSK\VLRORJLFSURFHVVHVVXFKDVPHFKDQLFDORYHUORDGDWKHURVFOHURVLV LQȵDPPDWLRQDQGFDUGLDFȴEURVLV7KHUHIRUHZHH[SHFWWKDWULVNPRGHOVWKDWDFFRXQW IRU UHSHDWHG PHDVXUHPHQWV PD\ PRUH DGHTXDWHO\ UHȵHFW WKH FXUUHQW VWDWXV RI WKH patient compared to models that only use single measurements.

The TRanslational Initiative on Unique and novel strategies for Management of Patients with Heart failure (TRIUMPH) study was designed to identify and validate novel biomarkers to improve prognostication in HF.6 _{TRIUMPH was designed as a translational study}

SURJUDPFRPELQLQJELRORJLFDOGLVFRYHU\RIQRYHOELRPDUNHUVWHFKQRORJLFDGYDQFHVDQG FOLQLFDOYDOLGDWLRQLQSDWLHQWVSUHVHQWLQJZLWKDFXWH+)ΖQWKHFOLQLFDOYDOLGDWLRQVWXG\ both the novel and established HF biomarkers were evaluated for their prognostic properties using a unique design of seven planned repeated measurements during \HDUIROORZXS%DVHGRQSUHYLRXVFOLQLFDODQGHSLGHPLRORJLFDOVWXGLHVJDOHFWLQZDV earmarked as a biomarker with high potential for improving prognostication.

*DOHFWLQ LV D PHPEHU RI D ODUJH IDPLO\ RI ſJDODFWRVLGHELQGLQJ DQLPDO OHFWLQV7

*DOHFWLQ H[SUHVVLRQ KDV EHHQ GHWHFWHG LQ PDFURSKDJHV QHXWURSKLOV HRVLQRSKLOV DQG PDVW FHOOV ΖQ UHVSRQVH WR D YDULHW\ RI PHFKDQLFDO DQG QHXURKRUPRQDO VWLPXOL macrophages secrete galectin-3.8_{ *DOHFWLQ VWLPXODWHV DGGLWLRQDO PDFURSKDJHV}

SHULF\WHV P\RȴEUREODVWV DQG ȴEUREODVWV ZKLFK DUH DOO LQYROYHG LQ WKH LQLWLDWLRQ DQG SURJUHVVLRQ RI WLVVXH VFDUULQJ &RQVHTXHQWO\ JDOHFWLQ DSSHDUV WR EH LQYROYHG LQ FDUGLDF ȴEURVLV ΖQ DGGLWLRQ JDOHFWLQ SOD\V DQ LPSRUWDQW UROH LQ WKH LQȵDPPDWRU\ UHVSRQVHZKLFKLVDQLPSRUWDQWVWHSLQWKHSURFHVVRIFDUGLDFUHPRGHOLQJ9-11 _Galectin-3

LVH[SUHVVHGLQQXPHURXVWLVVXHVVXFKDVKHDUWNLGQH\OXQJXWHUXVDQGFRORQ12 _The

OHYHORIJDOHFWLQH[SUHVVLRQLVUHODWLYHO\ORZLQKHDUWWLVVXHXQGHUQRUPDOFRQGLWLRQV but may increase substantially under pathophysiological circumstances.13

Several clinical studies have evaluated the prognostic value of galectin-3. Higher levels of galectin-3 have been associated with an increased risk of incident HF and all-cause

mortality in the general population._{)XUWKHUPRUHVLQJOHJDOHFWLQOHYHOVKDYHVKRZQ}

WR EH DQ LQGHSHQGHQW ULVN IDFWRU RI PRUWDOLW\ LQ ERWK VWDEOH DQG DFXWH +) SDWLHQWV although it still remains uncertain whether galectin-3 confers independent prognostic information when added to NT-proBNP. _{A few studies have been performed to}

assess the prognostic value of galectin-3 when measured multiple times. The change in galectin-3 level over time was predictive of outcome.19-21_{+RZHYHUJLYHQWKHG\QDPLFDQG}

SURJUHVVLYHQDWXUHRI+)WKHQXPEHURIJDOHFWLQPHDVXUHPHQWVQHHGHGIRUDGHTXDWH HVWLPDWLRQRIWKHWUXHJDOHFWLQOHYHOLVH[SHFWHGWREHKLJK7KHUHIRUHLQWKHSUHVHQW VWXG\ZHDVVHVVHGWKHLQGHSHQGHQWDVVRFLDWLRQEHWZHHQWKHHVWLPDWHGLQVWDQWDQHRXV JDOHFWLQOHYHOXVLQJIUHTXHQWO\PHDVXUHGJDOHFWLQOHYHOVDQGWKHLQFLGHQFHRIDOO cause mortality and HF readmission during 1-year follow-up in the 496 patients with acute HF who compose the TRIUMPH clinical cohort.

Methods

Objective and study design

TRIUMPH was designed as a translational bench-to-bedside study program encompassing the entire spectrum of biomarker discovery to clinical validation.6 _{The clinical validation}

study was an observational prospective study enrolling patients admitted with acute +)LQKRVSLWDOVLQ7KH1HWKHUODQGVEHWZHHQ6HSWHPEHUDQG'HFHPEHU This cohort study was designed to validate the clinical value of biomarkers successfully SDVVLQJ WKH ELRLQIRUPDWLFV DQG HDUO\YDOLGDWLRQ VWDJHV RI 75Ζ803+ DQG WR IXUWKHU evaluate more established biomarkers of HF. There was a particular interest in the change in biomarker levels over time. The study was approved by the medical ethics committee at all participating centers.

Patient selection

3DWLHQWV Ȳ \HDUV RI DJH ZHUH HOLJLEOH IRU HQUROOPHQW LI WKH\ ZHUH KRVSLWDOL]HG ZLWK GHFRPSHQVDWLRQRINQRZQFKURQLF+)RUQHZO\GLDJQRVHG+))XUWKHUPRUHWKUHHRWKHU FULWHULD KDG WR EH PHW  QDWULXUHWLF SHSWLGH OHYHOV KDG WR EH HOHYDWHG WR Ȳ WLPHV WKHXSSHUOLPLWRIQRUPDO8/1WKHUHKDGWREHHYLGHQFHRIVXVWDLQHGV\VWROLFRU GLDVWROLFOHIWYHQWULFXODUG\VIXQFWLRQDQGSDWLHQWVKDGWREHWUHDWHGZLWKLQWUDYHQRXV GLXUHWLFV3DWLHQWVZLWK+)SUHFLSLWDWHGE\DQRQFDUGLDFFRQGLWLRQE\VHYHUHYDOYXODU G\VIXQFWLRQZLWKRXWVXVWDLQHGOHIWYHQWULFXODUG\VIXQFWLRQRUE\DQDFXWH67VHJPHQW HOHYDWLRQP\RFDUGLDOLQIDUFWLRQZHUHH[FOXGHG)XUWKHUPRUHSDWLHQWVVFKHGXOHGIRUD FRURQDU\UHYDVFXODUL]DWLRQSURFHGXUHRQDZDLWLQJOLVWIRUDKHDUWWUDQVSODQWDWLRQZLWK VHYHUHUHQDOIDLOXUHIRUZKLFKGLDO\VLVZDVQHHGHGRUZLWKDFRH[LVWHQWFRQGLWLRQZLWK

DOLIHH[SHFWDQF\\HDUFRXOGQRWSDUWLFLSDWH$OOVWXG\SDUWLFLSDQWVSURYLGHGZULWWHQ informed consent.

Patient management

3DWLHQWPDQDJHPHQWZDVDWWKHGLVFUHWLRQRIWKHWUHDWLQJSK\VLFLDQDQGLQDFFRUGDQFH with the guidelines of the European Society of Cardiology.22_{ΖPSRUWDQWO\WKHELRPDUNHU}

GDWDWKDWZHUHJHQHUDWHGLQWKHFRQWH[WRIWKLVREVHUYDWLRQDOVWXG\ZHUHQRWXVHGIRU treatment decisions.

Study procedures

'XULQJKRVSLWDOL]DWLRQEORRGVDPSOHVZHUHREWDLQHGDWDGPLVVLRQGD\RQFHGXULQJ GD\V  WR  DQG VXEVHTXHQWO\ RQ WKH GD\ RI GLVFKDUJH $IWHUZDUGV UHSHDWHG EORRG VDPSOHVZHUHDOVRREWDLQHGDWRXWSDWLHQWIROORZXSYLVLWVZKLFKZHUHSODQQHGDWWR ZHHNVPRQWKVPRQWKVDQGWRPRQWKVDIWHUGLVFKDUJHThe baseline blood VDPSOH ZDV GHȴQHG DV WKH ȴUVW VDPSOH REWDLQHG DIWHU LQFOXVLRQ XS WR D PD[LPXP of 2 days after inclusion. $W HDFK YLVLW +) V\PSWRPV ZHUH DVVHVVHG XVLQJ WKH 1<+$ FODVVLȴFDWLRQ0HGLFDWLRQXVHZDVGHWHUPLQHGDWGLVFKDUJHXVLQJWKUHHFDWHJRULHV use of an angiotensin converting enzyme inhibitor (ACE-I) or an angiotensin II receptor DQWDJRQLVW$5%RUERWKXVHRIDEHWDEORFNHUXVHRIGLXUHWLFV3DWLHQWVXQGHUZHQW SK\VLFDO H[DPLQDWLRQ DQG V\VWHPDWLF PHDVXUHPHQWV RI ZHLJKW EORRG SUHVVXUH DQG heart rate.

Blood collection

Non-fasting blood samples were obtained by venipuncture and transported to the clinical chemistry laboratory of each participating hospital for further processing according to a standardized protocol. The collected material was centrifuged at 1700 G 5HODWLYH&HQWULIXJDO)RUFHZKHUHDIWHUFLWUDWH('7$KHSDULQHDQGWUDV\OROSODVPD ZDVVHSDUDWHGDVZHOODVEORRGVHUXP%X΍\FRDWVZHUHFROOHFWHGIURP('7$WXEHVWR HQDEOHDQDO\VLVRIJHQHWLFIDFWRUV'LPHWK\OVXOIR[LGH'062ZDVDGGHGWRDQDGGLWLRQDO EDTA tube for cryopreservation of blood cells. All blood aliquots were subsequently stored at a temperature of -80°C within 2 hours after venipuncture.

Galectin-3 measurements

Serum and heparin-plasma was transported under controlled conditions to a central laboratory (Future Diagnostics Solutions B.V.) for batch analysis of galectin-3 and NT-SUR%13 OHYHOV *DOHFWLQ FRQFHQWUDWLRQV ZHUH GHWHUPLQHG LQ VHUXP XVLQJ WKH %*0 *DOHFWLQ7HVWDVLQVWUXFWHGE\WKHPDQXIDFWXUHU%*0HGLFLQHΖQF:DOWKDP86$ NT-proBNP concentrations were determined in heparin plasma using the Elecsys

17SUR%13 DVVD\ RQ D &REDV  DQDO\]HU 5RFKH 'LDJQRVWLFV /LPLWHG 5RWNUHX] Switzerland). Analysts were blinded for patient characteristics and endpoints.

Endpoints

Information on vital status and hospital readmissions was obtained until at least 9 PRQWKVZLWKDPD[LPXPRIGD\VDIWHUWKHLQGH[KRVSLWDOL]DWLRQ:HDSSURDFKHGWKH FLYLOUHJLVWU\VFUHHQHGDOOPHGLFDOUHFRUGVDQGDVNHGSDWLHQWVIRULQIRUPDWLRQGXULQJ their follow-up visits.

The primary endpoint is the composite of all-cause mortality and readmission IRU +) 5HDGPLVVLRQ IRU +) ZDV GHȴQHG DV DQ XQSODQQHG UHKRVSLWDOL]DWLRQ GXH WR GHFRPSHQVDWLRQ RI +) ZLWK DW OHDVW WZR RI WKH IROORZLQJ WKUHH FULWHULD EHLQJ SUHVHQW HOHYDWHG QDWULXUHWLF SHSWLGH OHYHOV Ȳ WLPHV WKH 8/1 V\PSWRPV RI FDUGLDF GHFRPSHQVDWLRQ UDOHV HGHPD RU HOHYDWHG FHQWUDO YHQRXV SUHVVXUH DQG WUHDWPHQW with intravenous diuretics. Secondary endpoints included the individual components of the primary endpoint and cardiovascular mortality. $QHYHQWDGMXGLFDWLRQFRPPLWWHH EOLQGHGIRUELRPDUNHULQIRUPDWLRQZDVHVWDEOLVKHGIRUUHYLHZLQJDQGDGMXGLFDWLRQRI endpoints.

Statistical analysis

7KH GLVWULEXWLRQV RI FRQWLQXRXV YDULDEOHV LQFOXGLQJ ELRPDUNHUOHYHOV ZHUH HYDOXDWHG IRU QRUPDOLW\ E\ YLVXDO H[DPLQDWLRQ RI WKH KLVWRJUDP DQG .ROPRJRURY6PLUQRY WHVWV 9DULDEOHVZLWKDQRUPDOGLVWULEXWLRQDUHSUHVHQWHGDVPHDQsVWDQGDUGGHYLDWLRQ6' whereas the median and interquartile range (IQR) are presented in case of non-normality. Categorical variables are presented as counts and percentages. Galectin-3 and NT-proBNP levels had a non-normal distribution and were therefore log-transformed for further analysis.

3DWLHQWV ZHUH FODVVLȴHG DFFRUGLQJ WR WKH TXDUWLOHV RI WKH JDOHFWLQ GLVWULEXWLRQ DQG GL΍HUHQFHV LQ EDVHOLQH FKDUDFWHULVWLFV EHWZHHQ WKHVH TXDUWLOHV ZHUH HYDOXDWHG E\ FKLVTXDUHWHVWVFDWHJRULFDOYDULDEOHVDQDO\VLVRIYDULDQFHRU.UXVNDO:DOOLVWHVWVDV appropriate.

:HDSSOLHG&R[SURSRUWLRQDOKD]DUGVPRGHOVWRHYDOXDWHWKHDVVRFLDWLRQRIEDVHOLQH galectin-3 levels with the study endpoints. Subjects were censored at the time of RFFXUUHQFH RI WKH HQGSRLQW XQGHU LQYHVWLJDWLRQ GHDWK DQG DW WKH VFKHGXOHG HQG of follow-up. No deviations of the proportional hazards assumption were found by inspecting log minus log plots of the survival functions. We performed univariate analyses

WRREWDLQWKHFUXGHHVWLPDWHVRIWKHH΍HFWRIEDVHOLQHJDOHFWLQOHYHOPRGHODQDO\VHV WKDWZHUHDGMXVWHGIRUDJHDQGVH[RQO\PRGHODQGDQDO\VHVWKDWZHUHDGGLWLRQDOO\ DGMXVWHGIRUV\VWROLFEORRGSUHVVXUHGLDEHWHVPHOOLWXVOHIWYHQWULFXODUHMHFWLRQIUDFWLRQ /9()SUHYLRXVKRVSLWDOL]DWLRQIRU+)GXULQJWKHODVWPRQWKVLVFKHPLF+)ERG\PDVV LQGH[HVWLPDWHGJORPHUXODUȴOWUDWLRQUDWHH*)5DQGEDVHOLQH17SUR%13OHYHOPRGHO 3). The results are presented as adjusted hazard ratios (HR) per 1 SD increase of the ELRPDUNHUOHYHORQWKHORJVFDOHZLWKFRQȴGHQFHLQWHUYDOV&ΖWe calculated the H*)5XVLQJWKH0RGLȴFDWLRQRI'LHWLQ5HQDO'LVHDVHHTXDWLRQ23

-RLQW PRGHOV ZHUH ȴWWHG WR DVVHVV WKH DVVRFLDWLRQ EHWZHHQ HVWLPDWHG LQVWDQWDQHRXV ELRPDUNHU OHYHOV FDOFXODWHG XVLQJ WKH UHSHDWHG ELRPDUNHU OHYHOV DQG WKH VSHFLȴHG study endpoints$MRLQWPRGHOFRPELQHVDPL[HGH΍HFWVOLQHDUUHJUHVVLRQPRGHOIRUWKH VHULDOPHDVXUHPHQWVZLWKD&R[SURSRUWLRQDOKD]DUGVPRGHOIRUWKHULVNRIWKHVSHFLȴHG study endpoints.24 _{We used FXELFVSOLQHVZLWKNQRWVVHWDWZHHNDQGPRQWKDIWHU}

LQLWLDOKRVSLWDOL]DWLRQ)RUWKHDQDO\VHVZLWKWKHUHSHDWHGJDOHFWLQPHDVXUHPHQWVZH XVHGVLPLODUXQLYDULDWHDQGPXOWLYDULDWHPRGHOVDVPHQWLRQHGDERYHPRGHOVDQG H[FHSWIRUPRGHOZHDGGHGPHGLFDWLRQXVHDWGLVFKDUJHWRWKHPL[HGH΍HFWVOLQHDU regression model. We also tested whether the instantaneous slope of the galectin-3 WUDMHFWRULHVLWVHOIZKHQDGGHGWRPRGHOZDVDQLQGHSHQGHQWSUHGLFWRU)LQDOO\ZH combined the repeated measurements of galectin-3 and NT-proBNP to assess their respective independent prognostic value. Taking into account the limitations of the R SDFNDJHVIRU-RLQW0RGHOLQJZHZHUHDEOHWRFRPELQHWKHHVWLPDWHGJDOHFWLQWUDMHFWRU\ (using a PL[HGH΍HFWVOLQHDUUHJUHVVLRQPRGHO) and the estimated NT-proBNP trajectory XVLQJDWLPHGHSHQGHQW&R[SURSRUWLRQDOKD]DUGVPRGHOLQRQHMRLQWPRGHOSince the PRGHOGLGQRWFRQYHUJHZKHQZHDGMXVWHGIRUDOOWKHFRYDULDWHVLQPRGHOEDVHOLQH V\VWROLFEORRGSUHVVXUHKDGWREHOHIWRXWLQWKLVȴQDOPRGHOPRGHO'LDJQRVWLFVDQG sensitivity analyses were performed to evaluate the joint models. To account for the correlation structure between serial biomarker measurements collected from the same SDWLHQWZHREWDLQHGWKH6'IURPWKHWRWDOYDULDQFHRIDUDQGRPLQWHUFHSWVOLQHDUPL[HG PRGHOȴWWHGRQWKHSRVWGLVFKDUJHGDWD7KHȴQDOUHVXOWVDUHSUHVHQWHGDVDGMXVWHG+5 per 1 SD increase of the biomarker level (on the log2 scale) at any point in time with 95% CI.

7KH75Ζ803+VDPSOHVL]HZDVFKRVHQWRDFKLHYHDSRZHURIſ WRGHWHFW an odds ratio of at least 2.0 (α VLGHGWHVWIRUDELRPDUNHUYDOXHDERYHWKH percentile of its distribution comparing endpoint cases with non-cases. The incidence RI WKH SULPDU\ HQGSRLQW ZDV LQLWLDOO\ HVWLPDWHG DW  EDVHG RQ REVHUYDWLRQV LQ KLVWRULFDOKHDUWIDLOXUHSRSXODWLRQV7KHQSDWLHQWVDUHUHTXLUHG'XULQJWKHFRXUVH

RIWKHVWXG\EDVHGRQHYROYLQJHYLGHQFHWKHHVWLPDWHGLQFLGHQFHZDVDGMXVWHGWR DQGWKHVDPSOHVL]HZDVHYHQWXDOO\GHWHUPLQHGDWSDWLHQWV75Ζ803+HQUROOHG SDWLHQWVDQGUHDFKHGWKHSULPDU\HQGSRLQW

'DWD RQ FRYDULDWHV ZHUH FRPSOHWH LQ  RI SDWLHQWV H[FHSW IRU /9() ZKLFK ZDV complete in 78%. Single imputation was applied to account for missing values of covariates. 'DWDLVLPSXWHGXVLQJSUHGLFWLYHPHDQPDWFKLQJIRUFRQWLQXRXVYDULDEOHV ORJLVWLF UHJUHVVLRQ IRU ELQDU\ YDULDEOHV DQG SRO\WRPRXV UHJUHVVLRQ IRU XQRUGHUHG categorical data. Baseline covariates used in the full model and survival information were used in the imputation. The software used was R package MICE (https://cran.r-project.org/web/packages/mice/mice.pdf). A sensitivity analyses was performed on the full model for the primary end point on the complete cases.

7KH6WDWLVWLFDO3DFNDJHIRU6RFLDO6FLHQFHVYHUVLRQ6366Ζ%0FRUS$UPRQN1< 86$ZDVXVHGIRUGHVFULSWLYHGDWDDQDO\VLV5VWDWLVWLFDOVRIWZDUHYHUVLRQDYDLODEOH at: www.r-project.org) was used for advanced statistical analyses of the longitudinal biomarker data and study endpoints (packages JMBayes and JM). All statistical tests ZHUHWZRWDLOHGDQGSYDOXHVZHUHFRQVLGHUHGVWDWLVWLFDOO\VLJQLȴFDQW

Results

Patients

A total of 496 patients were enrolled in the TRIUMPH clinical cohort. Three patients withdrew their informed consent. Eighteen patients were withdrawn from statistical analyses due to inclusion violation. These patients had no evidence of sustained systolic RUGLDVWROLFOHIWYHQWULFXODUG\VIXQFWLRQRQHFKRFDUGLRJUDSK\$FFRUGLQJO\SDWLHQWV compose the analysis set. Their median age was 74 years (IQR 65-80) and 37% were women (Table 1). Median systolic blood pressure was 125 mmHg (IQR 110-147) and PHGLDQ/9()ZDVΖ45$WGLVFKDUJHXVHGDQ$&(ΖRUDQ$5%RUERWK 78% used a beta-blocker and 93% used diuretics. Median baseline galectin-3 level was 24 ng/ml (IQR 18-34) and NT-proBNP 4152 pg/ml (IQR 2089-9387). Table 2 shows the EDVHOLQHFKDUDFWHULVWLFVRISDWLHQWVLQGL΍HUHQWTXDUWLOHVRIJDOHFWLQOHYHO3DWLHQWVLQ quartiles with a higher galectin-3 level were older and had a worse kidney function. In the higher galectin-3 quartiles more patients had a history of myocardial infarction and GLDEHWHVPHOOLWXVKDGLVFKHPLF+)DQGKDGEHHQDGPLWWHGWRWKHKRVSLWDOIRUKHDUW failure during the last 6 months. In the lower galectin-3 quartiles more patients had newly-diagnosed HF during the initial hospitalization.

Table 1. %DVHOLQHSDUDPHWHUVDFFRUGLQJWRRYHUDOOVDPSOHLQVWXG\SRSXODWLRQ1 

Variables Overall sample

Demographic characteristics, median (IQR) or %

$JH\HDUV 74 (65-80)

Female 37

Caucasian 95

Measurements at baseline, median (IQR) or %

%RG\PDVVLQGH[NJP 28 (25-31) 6\VWROLFEORRGSUHVVXUHPP+J 125 (110-147) 'LDVWROLFEORRGSUHVVXUHPP+J 74 (65-85) +HDUWUDWHESP 85 (72-100) H*)5ml/min/1.73m2 _{46 (34-62)} /HIWYHQWULFXODUHMHFWLRQIUDFWLRQ 30 (21-41) 1<+$FODVVLȴFDWLRQ II III IV 17 55 27 Medical history, %

Newly diagnosed heart failure 36

Heart failure with reduced ejection fraction 83

Previous heart failure admission within 6 months

Ischemic heart failure 2049

Myocardial infarction 40

Hypertension 51

$WULDOȴEULOODWLRQ 42

Diabetes Mellitus 36

Stroke 17

Medication use at discharge, %

ACE-I and/or ARB 78

Beta-blocker 78

Diuretics 93

Biomarkers, median (IQR)

Galectin-3 (ng/ml) 24 (18-34)

NT-proBNP (pg/ml) 4152 (2089-9387)

IQR ΖQWHUTXDUWLOHUDQJHH*)5 HVWLPDWHGJORPHUXODUȴOWUDWLRQUDWH$&(Ζ $QJLRWHQVLQFRQYHUWLQJHQ]\PHLQKLELWRU $5% $QJLRWHQVLQΖΖUHFHSWRUDQWDJRQLVW

Baseline Galectin-3 levels and the incidence of study endpoints

During the median follow-up of 325 GD\V Ζ45   SDWLHQWV  UHDFKHG WKH SULPDU\ FRPSRVLWH HQGSRLQW RI DOOFDXVH GHDWK Q  RU UHDGPLVVLRQ IRU +) Q 7KLVFRUUHVSRQGVZLWKDQLQFLGHQFHUDWHΖ5RISHUSDWLHQW\HDUVIRU WKHSULPDU\HQGSRLQWΖQWKHKLJKHVWTXDUWLOHRIEDVHOLQHJDOHFWLQSDWLHQWV reached the primary endpoint compared to 27 patients (24%) in the lowest quartile. The number of events in the highest quartile compared to the lowest quartile of galectin-3 ZDVDOVRKLJKHUIRUDOOFDXVHPRUWDOLW\Q DQGQ UHVSHFWLYHO\DQG UHDGPLVVLRQIRU+)Q DQGQ UHVSHFWLYHO\

Table 2. Baseline characteristics according to quartiles of galectin-3 level.

Variables Quartile

1 Quartile 2 Quartile 3 Quartile 4 p-value*

Demographic characteristics, median or %

$JH\HDUV 70 73 76 75 0.010

Female 45 31 33 38 0.13

Caucasian 92 93 97 96 0.27

Measurements at baseline, median (IQR) or %

%RG\PDVVLQGH[NJP2 _{27 27 29 29 0.035} 6\VWROLFEORRGSUHVVXUHPP+J 130 125 125 122 0.29 'LDVWROLFEORRGSUHVVXUHPP+J 80 73 74 70  +HDUWUDWHESP 94 85 84 80 0.002 H*)5ml/min/1.73m2 _{63 55 42 32 } /HIWYHQWULFXODUHMHFWLRQIUDFWLRQ 30 30 34 31 0.020 1<+$FODVVLȴFDWLRQΖΖ III IV 23 50 27 18 51 28 11 63 25 14 60 26 0.12 Medical history, %

Newly diagnosed heart failure 57 40 26 21 

Heart failure with reduced ejection fraction 88 88 76 81 0.080

Previous heart failure admission within 6 months 8 17 24 29 

Ischemic heart failure 40 45 55 56 0.036

Myocardial infarction 28 32 54 48  Hypertension 40 50 56 60 0.016 $WULDOȴEULOODWLRQ 32 44 45 46 0.089 Diabetes Mellitus 20 32 41 50  Stroke 14 14 17 22 0.29 Biomarkers, median 17SUR%13SJPO *DOHFWLQQJPO 318016 397021 437228 754440  *P-value foUGL΍HUHQFHVEHWZHHQJURXSV H*)5 HVWLPDWHGJORPHUXODUȴOWUDWLRQUDWH

Baseline galectin-3 levels were higher in patients who reached a study endpoint when compared with those who remained event-free (Figure 1). The baseline galectin-3 level ZDV DVVRFLDWHG ZLWK DQ LQFUHDVHG ULVN RI UHDFKLQJ WKH SULPDU\ HQGSRLQW DV ZHOO DV ZLWKDOOFDXVHPRUWDOLW\FDUGLRYDVFXODUPRUWDOLW\DQG+)UHDGPLVVLRQ7DEOH$IWHU adjustment for all selected potential confounders including baseline NT-proBNP level PRGHO  WKH DVVRFLDWLRQ EHWZHHQ EDVHOLQH JDOHFWLQ DQG WKH GL΍HUHQW HQGSRLQWV EHFDPHZHDNHUEXWUHPDLQHGSUHVHQW

Table 3. +D]DUGUDWLRVIRUGL΍HUHQWHQGSRLQWVSHU6'LQFUHDVHRIWKHbaseline galectin-3 level (on the log2

scale).

Mean value * Baseline level †

M - SD M M + SD HR (95% CI) P-value 15.9 24.7 38.2 Primary endpoint Model 1 1.50 (1.30 - 1.75)  Model 2 1.49 (1.28 – 1.73)  Model 3 1.12 (0.93 – 1.36) 0.241

Number of events / patients 188/475

All-cause mortality

Model 1 1.54 (1.29 – 1.85) 

Model 2 1.52 (1.26 – 1.83) 

Model 3 1.26 ( 1.01 – 1.59) 0.044

Number of events / patients 113/475

HF hospitalization

Model 1 1.47 (1.22 – 1.76) 

Model 2 1.47 (1.23 – 1.76) 

Model 3 1.05 (0.82 – 1.33) 0.720

Number of events / patients 123/475

Cardiovascular mortality

Model 1 1.60 (1.28 – 1.99) 

Model 2 1.57 (1.26 – 1.97) 

Model 3 1.24 (0.93 – 1.67) 0.147

Number of events / patients 77/475

* Mean ±RQHVWDQGDUGGHYLDWLRQRIWKHSDWLHQWVSHFLȴFJHRPHWULFPean galectin-3 value at baseline (presented on the linear scale).

† Hazard ratios are related to a 1 SD increase of galectin-3 (on the log scale) at baseline.

0RGHOXQDGMXVWHGPRGHODGMXVWHGIRUDJHDQGVH[PRGHODGMXVWHGIRUDJHVH[V\VWROLFEORRGSUHVVXUHGLDEHWHV PHOOLWXV/9()SUHYLRXVKRVSLWDOL]DWLRQIRUKHDUWIDLOXUHGXULQJWKHODVWPRQWKVLVFKHPLFKHDUWIDLOXUHERG\PDVV LQGH[H*)5DQGEDVHOLQH17SUR%13

&ΖFRQȴGHQFHLQWHUYDO+)+HDUWIDLOXUH+5KD]DUGUDWLR0PHDQ6'VWDQGDUGGHYLDWLRQ

Repeatedly measured galectin-3 levels and the incidence of study endpoints

2QDYHUDJHJDOHFWLQZDVDYDLODEOHWLPHVGXULQJIROORZXS7KHPHDQJDOHFWLQ OHYHOGXULQJIROORZXSZDVQJPODQLQFUHDVHRI6'JDOHFWLQOHYHORQWKHORJ scale from the mean was 13 ng/ml. A decrease of 1 SD galectin-3 level on the log2 scale ZDVQJPO$IWHUDGMXVWPHQWIRUDJHDQGVH[PRGHOWKH+5SHU6'LQFUHDVHRIWKH galectin-3 level (on the log2 scale) at any point in time was 2.09 (95% CI 1.71 – 2.56) for the primary endpoint. After adjustment for the broader range of potential confounders LQFOXGLQJ PHGLFDWLRQ XVH DW GLVFKDUJH DQG EDVHOLQH 17SUR%13 OHYHO PRGHO  WKH DVVRFLDWLRQUHPDLQHGKLJKO\VWDWLVWLFDOO\VLJQLȴFDQWZLWKD+5RI&ΖȂ (Table 4). Results were similar for the secondary endpoints. The instantaneous slope of the galectin-3 level trajectories itself was not an independent predictor of the primary endpoint.

Figure 1. Distributions of baseline galectin-3 levels within the subpopulations of patients that had an event and

WKRVHZKRGLGQRWH[SHULHQFHDQHYHQWIRUDWKHSULPDU\HQGSRLQWEWKHVLQJOHHQGSRLQWRIDOOFDXVHPRUWDOLW\ c) the single endpoint of readmission for heart failure; d) the single endpoint of cardiovascular mortality.

Table 4. +D]DUGUDWLRVIRUGL΍HUHQWHQGSRLQWVSHU6'LQFUHDVHRIWKHJDOHFWLQOHYHORQWKHORJVFDOHDWDQ\

SRLQWLQWLPHXVLQJDMRLQWPRGHO

Mean value * Instantaneous level †

M - SD M M + SD HR (95% CI) P-value Primary endpoint 15.4 23.8 36.6 Model 1 2.07 (1.71 – 2.53)  Model 2 2.09 (1.71 – 2.56)  Model 3 1.67 (1.24 – 2.23)  All-cause mortality 15.4 23.8 36.9 Model 1 2.41 (1.83 – 3.15)  Model 2 2.36 (1.78 – 3.08)  Model 3 2.14 (1.47 – 3.16)  HF hospitalization 15.4 23.8 36.6 Model 1 1.87 (1.47 – 2.39)  Model 2 1.92 (1.48 – 2.46)  Model 3 1.41 (1.02 – 1.93) 0.035 Cardiovascular mortality 15.4 23.8 36.9 Model 1 2.46 (1.79 – 3.34)  Model 2 2.43 (1.76 – 3.35)  Model 3 2.22 (1.48 – 3.36) 

* Mean ± onHVWDQGDUGGHYLDWLRQRIWKHSDWLHQWVSHFLȴFJHRPHWULFPean galectin-3 value during follow-up (presented

on the linear scale).

† Hazard ratios are related to a 1 SD increase of galectin-3 (on the log scale) at any point in time.

0RGHOXQDGMXVWHGPRGHODGMXVWHGIRUDJHDQGVH[PRGHODGMXVWHGIRUDJHVH[V\VWROLFEORRGSUHVVXUHGLDEHWHV PHOOLWXV/9()SUHYLRXVKRVSLWDOL]DWLRQIRUKHDUWIDLOXUHGXULQJWKHODVWPRQWKVLVFKHPLFKHDUWIDLOXUHERG\PDVVLQGH[ H*)5PHGLFDWLRQXVHDWKRVSLWDOGLVFKDUJH$&(ΖDQGRU$5%EHWDEORFNHUGLXUHWLFVDQGEDVHOLQH17SUR%13OHYHO

Table 5. +D]DUGUDWLRVIRUGL΍HUHQWHQGSRLQWVSHU6'LQFUHDVHRIJDOHFWLQOHYHORU17SUR%13OHYHORQWKHORJ

scale) at any point in time using repeated galectin-3 and NT-proBNP measurements in a joint model.

Mean value * Instantaneous level †

M – SD M M + SD HR (95% CI) P-value Primary endpoint Galectin-3 15.4 23.8 36.6 1.54 (1.16 – 2.05) 0.003 NT-proBNP 742 2445 8062 2.10 (1.63 – 2.74)  All-cause mortality Galectin-3 15.4 23.8 36.9 1.77 (1.22 – 2.52)  NT-proBNP 739 2480 8321 2.68 (1.90 – 3.86)  HF hospitalization Galectin-3 15.4 23.8 36.6 1.29 (0.92 – 1.81) 0.160 NT-proBNP 742 2445 8062 1.71 (1.27 – 2.25)  Cardiovascular mortality Galectin-3 15.4 23.8 36.9 1.89 (1.25 – 2.85) 0.002 NT-proBNP 739 2480 8321 2.62 (1.70 – 4.27)  * Mean ± RQHVWDQGDUGGHYLDWLRQRIWKHSDWLHQWVSHFLȴFJHRPHWULFPHDQELRPDUNHUOHYHOGXULQJIROORZXSSUHVHQWHGRQ the linear scale).

† Hazard ratios are related to a 1 SD increase of biomarker level (on the log scale) at any point in time.

0RGHODGMXVWHGIRUDJHVH[GLDEHWHVPHOOLWXV/9()SUHYLRXVKRVSLWDOL]DWLRQIRUKHDUWIDLOXUHGXULQJWKHODVWPRQWKVLVFKHPLF KHDUWIDLOXUHERG\PDVVLQGH[H*)5PHGLFDWLRQXVHDWKRVSLWDOGLVFKDUJH$&(ΖDQGRU$5%EHWDEORFNHUGLXUHWLFVDQGEDVHOLQH NT-proBNP level

&ΖFRQȴGHQFHLQWHUYDO+)+HDUWIDLOXUH+5KD]DUGUDWLR0PHDQ6'VWDQGDUGGHYLDWLRQ

$IWHU DGMXVWPHQW IRU UHSHDWHG 17SUR%13 PHDVXUHPHQWV PRGHO  WKH DVVRFLDWLRQ between repeated galectin-3 levels and adverse outcome remained statistically VLJQLȴFDQWZLWKD+5RI&ΖȂIRUWKHSULPDU\HQGSRLQWFRUUHVSRQGLQJ with 1 SD increase of galectin-3 level (on the log2 scale) at any point in time (Table 5). The HR corresponding with a 1 SD increase of NT-proBNP level (on the log2 scale) at any point in time was 2.10 (95% CI 1.63 – 2.74) after adjustment for repeated galectin-3 levels.

Figure 2A shows the average estimated galectin-3 level in patients with and without the primary endpoint according to model 3 and the individual galectin-3 measurements. During hospitalization the average galectin-3 level remains steady for patients who remained free of the primary endpoint. For patients who reached the primary endpoint during follow-up the average estimated galectin-3 level decreased slightly after the initial KRVSLWDOL]DWLRQ $SSDUHQWO\ WKURXJKRXW IROORZXS SDWLHQWV ZKR UHDFKHG WKH SULPDU\ HQGSRLQWKDGRQDYHUDJHKLJKHUOHYHOVWKDQWKHLUFRXQWHUSDUWVZKRUHPDLQHGIUHHRI WKHSULPDU\HQGSRLQW)XUWKHUPRUHWKHDYHUDJHHVWLPDWHGJDOHFWLQOHYHOVDSSHDUHGWR HOHYDWHVHYHUDOZHHNVSULRUWRWKHWLPHRIWKHSULPDU\HQGSRLQWȴJXUH%

A

B

Figure2 A. Average estimated galectin-3 pattern during initial hospitalization for decompensated heart failure

IRUSDWLHQWVZLWKDQGZLWKRXWWKHSULPDU\HQGSRLQW7KHȴJXUHLQFOXGHVWKHLQGLYLGXDOJDOHFWLQPHDVXUHPHQWV for patients with and without the primary endpoint.

2 B. Average estimated galectin-3 pattern prior to the primary endpoint or end of follow-up for patients with and

ZLWKRXWWKHSULPDU\HQGSRLQW7KHȴJXUHLQFOXGHVWKHLQGLYLGXDOJDOHFWLQPHDVXUHPHQWVIRUSDWLHQWVZLWKDQG without the primary endpoint.

7KH DYHUDJH HVWLPDWHG JDOHFWLQ OHYHOV DUH DGMXVWHG IRU DJH VH[ V\VWROLF EORRG SUHVVXUH GLDEHWHV PHOOLWXV /9()SUHYLRXVKRVSLWDOL]DWLRQIRUKHDUWIDLOXUHGXULQJWKHODVWPRQWKVLVFKHPLFKHDUWIDLOXUHERG\PDVVLQGH[ H*)5PHGLFDWLRQXVHDWKRVSLWDOGLVFKDUJH$&(ΖDQGRU$5%EHWDEORFNHUGLXUHWLFV and baseline NT-proBNP (model 3).

Discussion

7KLV VWXG\ FOHDUO\ GHPRQVWUDWHV WKDW LQ SDWLHQWV DGPLWWHG ZLWK DFXWH +) UHSHDWHG galectin-3 measurements are a strong and independent predictor of the composite endpoint of all-cause mortality or readmission for HF during 1-year follow-up. Our results LOOXVWUDWHWKDWUHSHDWHGPHDVXUHPHQWVRIJDOHFWLQR΍HULQFUHPHQWDOSURJQRVWLFYDOXH WRUHSHDWHGO\PHDVXUHG17SUR%13ZKLFKLVFRQVLGHUHGWKHJROGVWDQGDUGELRPDUNHU in HF patients.

2XUREVHUYDWLRQWKDWEDVHOLQHJDOHFWLQOHYHOZDVDVVRFLDWHGZLWKPRUWDOLW\FRQȴUPV HDUOLHUȴQGLQJVERWKLQDFXWHDQGVWDEOH+)SDWLHQWV_{6LPLODUWRSUHYLRXVVWXGLHV}

the association between baseline galectin-3 level and mortality attenuated after DGMXVWPHQWIRUHVWDEOLVKHGULVNIDFWRUVLQFOXGLQJNLGQH\IXQFWLRQDQG17SUR%13OHYHO  _{The association between baseline galectin-3 level and readmission for HF was less}

DSSDUHQW +RZHYHU WKH GHFLVLRQ to hospitalize a patient for decompensation of HF PD\EHLQȵXHQFHGE\VHYHUDOVXEMHFWLYHSDWLHQWDQGSK\VLFLDQUHODWHGIDFWRUVWKDWDUH XQOLNHO\WRKDYHDQDVVRFLDWLRQZLWKWKHJDOHFWLQOHYHO)XUWKHUPRUHVHYHUDOULVNIDFWRUV VXFKDVNLGQH\IXQFWLRQGLDEHWHVPHOOLWXVDQG17SUR%13OHYHOLQȵXHQFHWKLVGHFLVLRQ DQG DUH UHODWHG WR JDOHFWLQ 7KHUHIRUH WKH DVVRFLDWLRQ EHWZHHQ EDVHOLQH JDOHFWLQ level and heart failure readmission attenuated after adjustment for these risk factors. Since the primary endpoint is a composite of all-cause mortality and readmission for +)WKHUHODWLRQVKLSEHWZHHQWKHJDOHFWLQOHYHODQGWKHPRUWDOLW\HQGSRLQWVSHUVHDUH stronger compared to the primary endpoint.

Repeated galectin-3 measurements were strongly and independently related to the SULPDU\HQGSRLQWDVZHOODVLWVVHSDUDWHFRPSRQHQWV5HSHDWHGPHDVXUHPHQWVWDNH into account the G\QDPLFDQGFRQWLQXRXVFKDQJHLQJDOHFWLQOHYHORYHUWLPHZKLFK EHWWHUUHȵHFWVWKHWUXHQDWXUHRIWKHXQGHUO\LQJSDWKRSK\VLRORJ\LQ+)ΖQWKLVVWXG\WKH number of galectin-3 measurements per patient was high and therefore the repeated galectin-3 measurements could be used to estimate instantaneous galectin-3 levels (i.e. the estimated galectin-3 level at any point in time during the follow-up period). :KHQFRPSDUHGWREDVHOLQHJDOHFWLQOHYHOVWKHHVWLPDWHGLQVWDQWDQHRXVJDOHFWLQ OHYHOVLGHQWLȴHGSDWLHQWVDWDQHYHQKLJKHUULVNIRUUHDFKLQJDQHQGSRLQW7KHHVWLPDWHG LQVWDQWDQHRXVJDOHFWLQOHYHOPRUHDFFXUDWHO\DSSUR[LPDWHVWKHWUXHJDOHFWLQOHYHO DQGWKHUHIRUHUHȵHFWVWKHDFWXDOFRQGLWLRQRIWKHSDWLHQWDWWKDWSRLQWLQWLPHGXULQJ IROORZXS7KLVLVH[SHFWHGWREHLPSRUWDQWVLQFH+)LVDG\QDPLFDQGRIWHQSURJUHVVLYH GLVHDVHLQZKLFKLQȵDPPDWLRQFDUGLDFȴEURVLVDQGUHPRGHOOLQJDUHRQJRLQJSURFHVVHV that cannot be captured in a single biomarker assessment at one point in time.5

)XUWKHUPRUHEDVHOLQHJDOHFWLQPHDVXUHPHQWVZHUHDOOWDNHQGXULQJKRVSLWDOL]DWLRQ IRUGHFRPSHQVDWHGFKURQLF+)RUQHZRQVHW+)ΖWLVNQRZQWKDWJDOHFWLQLQFRQWUDVW WRQDWULXUHWLFSHSWLGHVGRHVQRWUHVSRQGWRYROXPHRYHUORDGDQGXQORDGLQJGLUHFWO\ which occurs during hospitalization.28 _{As galectin-3 is involved in the process of}

P\RFDUGLDOȴEURVLVLWLVPRUHOLNHO\WKDWJDOHFWLQLVRIPRUHSURJQRVWLFYDOXHZKHQ patients enter a more chronic phase of HF.11

ΖQWHUHVWLQJO\WKHVORSHRIWKHJDOHFWLQWUDMHFWRU\GLGQRWDGGSURJQRVWLFLQIRUPDWLRQWR WKHHVWLPDWHGLQVWDQWDQHRXVJDOHFWLQOHYHO$QH[SODQDWLRQFRXOGEHWKDWJDOHFWLQLV helpful in identifying high-risk patients when their galectin-3 level rises above a certain threshold. The change in galectin-3 level prior to reaching this threshold is not essential IRU ULVN VWUDWLȴFDWLRQ +RZHYHU WR EH DEOH WR HVWLPDWH ZKHWKHU D SDWLHQWȇV JDOHFWLQ OHYHOULVHVDERYHWKHWKUHVKROGUHSHDWHGPHDVXUHPHQWVDUHUHTXLUHG$IHZVWXGLHV have been conducted on the prognostic value of multiple galectin-3 measurements in acute and stable HF patients. _{These studies showed that change in galectin-3 level is}

DVVRFLDWHGZLWKPRUWDOLW\$SRVVLEOHH[SODQDWLRQDVWRZK\LQWKHSUHVHQWVWXG\VORSHRI WKHJDOHFWLQWUDMHFWRU\GLGQRWDGGIXUWKHUSURJQRVWLFLQIRUPDWLRQPLJKWEHWKDWWKH number of galectin-3 measurements during follow-up was substantially higher in our VWXG\ZKLFKDOORZHGXVWRHVWLPDWHDQLQVWDQWDQHRXVVORSHRIWKHJDOHFWLQWUDMHFWRU\ UDWKHUWKDQWKHVORSHRIWKHGL΍HUHQFHȆGHOWDȇEHWZHHQWKHOHYHODWEDVHOLQHDQGWKDWDW Dȴ[HGSRLQWLQWLPH

The statistical method (Joint Model) used to estimate the trajectory of the galectin-3 level takes into account the continuous changes in biomarker levels and adequately analyses WKHUHODWLRQEHWZHHQWKHVHELRPDUNHUWUDMHFWRULHVDQGGL΍HUHQWHQGSRLQWVFRQVLGHULQJ the changing population due to censoring at the time of occurrence of an endpoint. Previous studies presented changes in biomarker level as a ‘delta’ between just two measurements that are separated in time. If more than two samples are taken into DFFRXQWSDWLHQWVKDYHRIWHQEHHQFDWHJRUL]HGDFFRUGLQJWRWKHQXPEHURIKLJKRUORZ ELRPDUNHUOHYHOV2EYLRXVO\ERWKDSSURDFKHVGRQRWIXOO\FDSWXUHWKHWUXHELRPDUNHU SDWWHUQRIWKHG\QDPLFGLVHDVH$GGLWLRQDOO\WKHSRZHUWRSUHGLFWDGYHUVHRXWFRPHLV reduced.

$QLPSRUWDQWȴQGLQJRIWKHSUHVHQWVWXG\LVWKDWUHSHDWHGJDOHFWLQPHDVXUHPHQWV FRQIHUUHG DGGLWLRQDO DQG LQGHSHQGHQW SURJQRVWLF LQIRUPDWLRQ WR WKDW R΍HUHG E\ baseline as well as repeated NT-proBNP measurements. The fact that NT-proBNP and JDOHFWLQUHȵHFWGL΍HUHQWXQGHUO\LQJSDWKRSK\VLRORJLFDOSURFHVVHVLQ+)PD\EHWKH PRVWLPSRUWDQWUHDVRQIRUWKLVREVHUYDWLRQ*DOHFWLQLVDPDUNHURIFDUGLDFȴEURVLV

LQȵDPPDWLRQDQGUHPRGHOOLQJZKHUHDV17SUR%13LVDPDUNHURIYROXPHRYHUORDG

$VVXFKJDOHFWLQPLJKWEHDPDUNHUWKDWPRUHGLUHFWO\UHȵHFWVWKHSDWKRSK\VLRORJLFDO processes that lead to adverse cardiac remodelling and deterioration of cardiac IXQFWLRQ ZKHUHDV 17SUR%13 UHȵHFWV WKH YROXPH RYHUORDG UHVXOWLQJ IURP WKH DFWXDO OHIW YHQWULFXODU G\VIXQFWLRQ ΖQ WKLV ZD\ WKH JDOHFWLQ DQG 17SUR%13 OHYHO SURYLGH FRPSOLPHQWDU\LQIRUPDWLRQRQWKHSDWKRSK\VLRORJLFDOVWDWHDVZHOODVZLWKUHVSHFWWR WKHDVVHVVPHQWRISURJQRVLV:LWKUHVSHFWWRSURJQRVWLFDWLRQLQ+)WKHUHVXOWVRIWKH SUHVHQWVWXG\WKHUHIRUHQRWRQO\SURYLGHHYLGHQFHIRUWKHXVHRIUHSHDWHGJDOHFWLQ PHDVXUHPHQWVEXWDOVRIRUWKHFRPELQHGXVHZLWKUHSHDWHGO\PHDVXUHG17SUR%13 $OWKRXJKWKLVVWXG\LVDODUJHPXOWLFHQWUHSURVSHFWLYHREVHUYDWLRQDOVWXG\LWVHHPVWKDW the studied population is not completely representable for the average HF population. The mean age in our study population is 74 and the women are underrepresented. Moreover only 18% of the included HF patients have a preserved ejection fraction. De Boer et al.30_{VKRZHGWKDWJDOHFWLQOHYHOVGLGQRWGL΍HUEHWZHHQ+)SDWLHQWVZLWK}

a reduced and preserved ejection fraction and the predictive value of galectin-3 was stronger in patients with a preserved ejection fraction. By underrepresenting the HF patients with a preserved ejection fraction in our study we possibly underestimated the prognostic value of galectin-3.

Future studies should evaluate the value of repeated galectin-3 measurements when used to guide treatment decisions. It may be hypothesized that treatment is to be LQWHQVLȴHGLQSDWLHQWVZLWKKLJKJDOHFWLQOHYHOVRUXQIDYRXUDEOHJDOHFWLQSDWWHUQV2Q WKHRWKHUKDQGUHSHDWHGJDOHFWLQPHDVXUHPHQWVPLJKWEHKHOSIXOWRLGHQWLI\SDWLHQWV who are more likely to respond to certain treatments.31_{)XUWKHUPRUHLWUHPDLQVWREH}

DGGUHVVHG ZKHWKHU JDOHFWLQ PD\ EH WDUJHWHG E\ VSHFLȴF DQWLJDOHFWLQ WKHUDSLHV Additional studies should also determine the number of galectin-3 measurements needed for optimal prognostication and therapy monitoring. The frequency by which JDOHFWLQOHYHOVVKRXOGEHPHDVXUHGPD\QRWEHLGHQWLFDOIRUHDFKSDWLHQWEXWGHSHQGV RQWKHFOLQLFDOFRQGLWLRQRIWKHSDWLHQWWKHWUHDWPHQWJLYHQWKHJDOHFWLQOHYHODQGWKH progression of galectin-3 levels during follow-up.

Conclusions

The TRIUMPH study clearly demonstrates that repeated measurements of galectin-3 are a strong and independent predictor of adverse outcome in patients following DGPLVVLRQIRUDFXWH+)7KHHVWLPDWHGLQVWDQWDQHRXVJDOHFWLQOHYHOLGHQWLȴHGSDWLHQWV

at a higher risk of reaching adverse events than baseline galectin-3 levels alone. In DGGLWLRQUHSHDWHGJDOHFWLQPHDVXUHPHQWVR΍HULQFUHPHQWDOSURJQRVWLFYDOXHWRWKDW FRQIHUUHGE\RWKHUNQRZQULVNIDFWRUVDQGLPSRUWDQWO\UHSHDWHGPHDVXUHPHQWVRI17 proBNP. These results suggest that repeated galectin-3 measurements in addition to NT-proBNP measurements may be helpful in clinical practice to identify HF patients who are at increased risk of adverse outcome.

Sources of funding

This work received support within the framework of CTMM (Center for Translational 0ROHFXODU 0HGLFLQH SURMHFW 75Ζ803+ JUDQW & 6DUD - %DDUW LV VXSSRUWHG by grant 2003T083 of the Netherlands Heart Foundation. Folkert W. Asselbergs is VXSSRUWHGE\D1HWKHUODQGV+HDUW)RXQGDWLRQ'HNNHUVFKRODUVKLS-XQLRU6WD΍0HPEHU 2014T001 and UCL Hospitals NIHR Biomedical Research Centre.

Contributors

/LVWRI6LWHΖQYHVWLJDWRUV+DQV/+LOOHJH0'3K'80&**URQLQJHQWKH1HWKHUODQGV ΖYRQQH/HVPDQ/HHJWH3K'80&**URQLQJHQWKH1HWKHUODQGV0LFKLHO-1DJHOVPLW 0' 3K' 7UHDQW =RUJJURHS 6FKHSHU]LHNHQKXLV (PPHQ WKH 1HWKHUODQGV (OO\ 0&- :DMRQ0'0HGLVFK6SHFWUXP7ZHQWH(QVFKHGHWKH1HWKHUODQGV+DQV3HWHU%UXQQHU /D5RFFD0'3K'0DDVWULFKW80&WKH1HWKHUODQGV*HUDUG&0/LQVVHQ0'3K' 7ZHQWHERUJ]LHNHQKXLV$OPHORWKH1HWKHUODQGV:LOOHP)7HUSVWUD0'3K'6OLQJHODQG =LHNHQKXLV 'RHWLQFKHP WKH 1HWKHUODQGV . 0DUWLMQ $NNHUKXLV 0' 3K' (UDVPXV 0&5RWWHUGDPWKH1HWKHUODQGV/DXUD&YDQ9DUN0'(UDVPXV0&5RWWHUGDPWKH 1HWKHUODQGV(ULF%RHUVPD3K'(UDVPXV0&5RWWHUGDPWKH1HWKHUODQGV$QKR+ /LHP 0' 3K' $GPLUDDO GH 5X\WHU =LHNHQKXLV *RHV WKH 1HWKHUODQGV $OH[DQGHU - :DUGHK0'3K'+DDJODQGHQ0&'HQ+DDJWKH1HWKHUODQGV$GG\-0YDQ0LOWHQEXUJ 0' 3K' )UDQVLVFXV *DVWKXLV  9OLHWODQG 5RWWHUGDP WKH 1HWKHUODQGV (ZRXW - YDQ GHQ%RV0'3K'$OEHUW6FKZHLW]HU=LHNHQKXLV'RUGUHFKWWKH1HWKHUODQGV6WLMQ3- GH5LGGHU6W$QQD=LHNHQKXLV*HOGURSWKH1HWKHUODQGV)RONHUW:$VVHOEHUJV0' 3K' 80& 8WUHFKW WKH 1HWKHUODQGV <LJDO 0 3LQWR 0' 3K' $0& $PVWHUGDP WKH Netherlands).

Disclosures

References

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measurements in patients

with acute heart failure

L.C. van Vark, I. Lesman-Leegte, S.J. Baart, D. Postmus, Y.M. Pinto, J.G. Orsel, B.D. Westenbrink, H.P. Brunner-la Rocca, A.J.M. van Miltenburg, E. Boersma, H.L. Hillege, K.M. Akkerhuis; TRIUMPH Investigators