Neuron-specific enolase has potential value as a biomarker for [18F]FDG/[68Ga]Ga-PSMA-11 PET mismatch findings in advanced mCRPC patients

ORIGINAL RESEARCH Open Access Neu on-speci ic enolase has po en ial alue as a bioma ke o [ 18 F]FDG/[ 68 Ga]Ga-PSMA- 11 PET misma ch indings in ad anced mCRPC pa ien s Flo ian Rosa 1* , Kalle Ribba 1 , Ma in Ries 1 , Johannes Linxweile 2 , Ma k Ba holomä 1 , S ephan Maus 1 , Ma hias Sch eckenbe ge 3 , Same Ezziddin 1 and Fadi Kh eish 1 Abs ac Backg ound: PSMA- a ge ed adioligand he apy (PSMA-RLT) yielded imp essi e esul s in he me as asized cas a ion- esis an p os a e ca cinoma (mCRPC) se ing. High exp ession o PSMA is essen ial o success ul PSMA- RLT. Howe e , some pa ien s de elop [ 18 F]FDG-a id lesions wi h low o no PSMA exp ession ([ 18 F]FDG/[ 68 Ga]Ga- PSMA-11 misma ch indings on PET/CT) in he cou se o ea men . Those lesions a e no a ec ed by PSMA-RLT and a change in he apy managemen is needed. To enable ea ly misma ch de ec ion, possible blood pa ame e s as indica o s o he occu ence o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch indings on PET/CT we e e alua ed. Me hods: Re ospec i e s udy o N= 66 ad anced mCRPC pa ien s wi h dual [ 68 Ga]Ga-PSMA-11 and [ 18 F]FDG PET/ CT imaging wi hin 4 weeks, who we e e e ed o o ecei ed [ 177 Lu]Lu-PSMA-617 adioligand he apy. P os a e- speci ic an igen (PSA), neu on-speci ic enolase (NSE), gamma-glu amyl ans e ase (GGT), and alkaline phospha ase (ALP) we e es ed as indica o s o he occu ence o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch indings. Addi ional o absolu e alues, ela i e changes (ΔPSA, ΔNSE, ΔGGT, ΔALP) o e a pe iod o 4 ± 1 weeks p io o [ 18 F]FDG PET/CT we e analyzed. Resul s: In o al, 41/66 (62%) pa ien s e ealed a leas one [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch inding on PET/ CT. These misma ch indings we e de ec ed in 13/41 (32%) pa ien s by sc eening o and in 28/41 (68%) pa ien s du ing PSMA-RLT. NSE se um le el (55.4 ± 44.6 μg/l s.18.5 ± 8 μg/l, p< 0.001) and ΔNSE (93.8 ± 124.5% s.2.9 ± 39.5%, p< 0.001) we e signi ican ly highe in he misma ch g oup han in he non-misma ch g oup. No signi ican di e ences we e ound o se um PSA (p= 0.424), ΔPSA (p= 0.417), se um ALP (p= 0.937), ΔALP (p= 0.611), se um GGT (p= 0.773), and ΔGGT (p= 0.971). Fo NSE and ΔNSE, he maximum alue o he Youden index in ROC analysis was a a cu -o le el o 26.8 μg/l (sensi i i y 78%, speci ici y 96%) and a + 13.9% (sensi i i y 84%, speci ici y 75%), espec i ely. An in oduced sco ing sys em o bo h pa ame e s achie ed a sensi i i y o 90% and a speci ici y o 88% o he occu ence o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch. (Con inued on nex page) © The Au ho (s). 2020 Open Access This a icle is licensed unde a C ea i e Commons A ibu ion 4.0 In e na ional License, which pe mi s use, sha ing, adap a ion, dis ibu ion and ep oduc ion in any medium o o ma , as long as you gi e app op ia e c edi o he o iginal au ho (s) and he sou ce, p o ide a link o he C ea i e Commons licence, and indica e i changes we e made. The images o o he hi d pa y ma e ial in his a icle a e included in he a icle's C ea i e Commons licence, unless indica ed o he wise in a c edi line o he ma e ial. I ma e ial is no included in he a icle's C ea i e Commons licence and you in ended use is no pe mi ed by s a u o y egula ion o exceeds he pe mi ed use, you will need o ob ain pe mission di ec ly om he copy igh holde . To iew a copy o his licence, isi h p://c ea i ecommons.o g/licenses/by/4.0/. * Co espondence: [email p o ec ed] 1 Depa men o Nuclea Medicine, Saa land Uni e si y—Medical Cen e , Ki be ge S . 100, Geb. 50, 66421 Hombu g, Ge many Full lis o au ho in o ma ion is a ailable a he end o he a icle Rosa e al. EJNMMI Resea ch (2020) 10:52 h ps://doi.o g/10.1186/s13550-020-00640-2 (Con inued om p e ious page) Conclusion: We obse ed a signi ican ly highe absolu e se um concen a ion and a highe ela i e inc ease o NSE in ad anced mCRPC pa ien s wi h [ 18 F]FDG-a id and insu icien PSMA exp essing me as ases ([ 18 F]FDG/[ 68 Ga]Ga- PSMA-11 misma ch indings on PET/CT) in ou coho . NSE migh be used as a po en ial labo a o y indica o o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch indings, i his obse a ion is con i med in u u e, ideally p ospec i e, s udies in la ge pa ien coho s. Keywo ds: P os a e cance , mCRPC, PSMA PET/CT, FDG PET/CT, Misma ch In oduc ion Wi h o e 1,000,000 new cases and app oxima ely 300,000 dea hs wo ldwide in 2012, p os a e ca cinoma is one o he mos equen malignan diseases in men [1]. A signi ican ac ion o p os a e ca cinoma pa ien s’p og esses o he le- hal me as asized cas a ion- esis an p os a e ca cinoma (mCRPC) se ing [2,3]. Du ing he las decade, howe e , he de elopmen o new ea men op ions o men wi h mCRPC has led o an imp o ed su i al ime [4]. In addi ion o chemo he apy wi h doce axel o cabazi axel [5, 6] and nex -gene a ion and ogen ecep o signaling inhib- i ion wi h abi a e one o enzalu amide [7,8], adioligand he apy a ge ing he p os a e-speci ic memb ane an igen (PSMA) is a po en ial op ion in pallia i e se ings [9,10]. PSMA- a ge ed adioligand he apy (PSMA-RLT) wi h [ 177 Lu]Lu-PSMA-617 yielded imp essi e esul s in pallia i e se ings while causing only mode a e side e ec s [11,12]. PSMA- a ge ed based posi on emission omog aphy (PET)/compu e omog aphy (CT) wi h adiolabeled PSMA ligands, such as [ 68 Ga]Ga-PSMA-11, is equen ly used o imaging o p os a e cance in clinical ou ine [13]. PSMA- a ge ed PET/CT is no only used o he s aging o p os- a e cance , bu i is also a use ul ool o he apy moni o - ing [14,15] and indispensable o e i ying PSMA exp ession p io o PSMA-RLT [16]. High exp ession o PSMA is essen ial o success ul PSMA-RLT in pa ien s wi h mCRPC. Howe e , some pa ien s de elop lesions wi h low o no PSMA exp ession unde ongoing ea men [17]. Those lesions a e no a ec ed by PSMA-RLT and a change in he apy managemen is needed [9,18]. [ 18 F]FDG PET/ CT using 18 F-labeled luo odeoxyglucose ([ 18 F]FDG) in addi ion oaPSMA- a ge edPET/CTmaybeasui able me hod o de ec ion o hose lesions. Mos ly, p os a e ca - cinoma cells ha e a low glucose me abolism due o ene gy gain by lipids and o he ene ge ic molecules bu in ad- anced la e-s age disease he glucose me abolism is highly inc eased by he Wa bu g e ec and shi ing o ae obic glycolysis a e nume ous mu a ion e en s [19]. The e o e, a combina ion o [ 18 F]FDG and PSMA- a ge ed PET/CT may be able o de ec clinically ele an glucose me abolic iable umo lesions wi h low o no PSMA exp ession (mis- ma ch lesions) in he ad anced mCRPC se ing. A ecen ly p ospec i e phase-II ial o [ 177 Lu]Lu-PSMA-617 RLT excluded 16% o sc eened pa ien s due o missing o low PSMA exp ession in [ 18 F]FDG-a id me as ases [20]. Ea ly de ec ion o hese misma ch indings is hus needed o p o- ide pa ien s wi h al e na i e he apy op ions, addi ionally o o ins ead o PSMA-RLT [18]. Howe e , equen ly pe - o ming [ 18 F]FDG PET/CT in addi ion o PSMA- a ge ed PET/CT is e y cos in ensi e and associa ed wi h add- i ional adia ion exposu e o he pa ien , which should be a oided e en in a pallia i e se ing. In his s udy, selec ed se um pa ame e s we e he e o e es ed as indica o s o he occu ence o [ 18 F]FDG/ [ 68 Ga]Ga-PSMA-11 misma ch indings on PET/CT in his s udy, including he p os a e-speci ic an igen (PSA) as he ou ine con ol and esponse pa ame e [21], alkaline phospha ase (ALP) as known o be ele a ed by bone me- as ases [22], gamma-glu amyl ans e ase (GGT) as a pa - ame e o li e unc ion a ec ed by li e me as ases [23], and neu on-speci ic enolase (NSE) as a possible pa ame e o ansdi e en ia ing o a neu oendoc ine ype o p os- a e ca cinoma [24]. Ma e ials and me hods S udy design Re ospec i e monocen e s udy o mCRPC pa ien s wi h dual [ 68 Ga]Ga-PSMA-11 and [ 18 F]FDG PET/CT imaging wi hin 4 weeks, who we e e e ed o o e- cei ed [ 177 Lu]Lu-PSMA-617 adioligand he apy a he clinic o nuclea medicine a Saa land Uni e si y Med- ical Cen e om Oc obe 2015 ill Augus 2019. Pa ien s wi h seconda y malignancies we e excluded o a oid po- en ial in e e ence o image in e p e a ion. Pa ien s and e hics N= 66 o in o al 167 mCRPC pa ien s e e ed o o ecei ed PSMA-RLT in ou cen e we e included in his e ospec i e s udy. Two o he 167 pa ien s we e ex- cluded because o incomple e blood examina ion, 3/167 because o seconda y malignancies and he emaining, and 96/167 due o missing o un imely [ 18 F]FDG PET/ CT. The pa ien s ecei ed a [ 68 Ga]Ga-PSMA-11 PET/ CT and [ 18 F]FDG PET/CT wi hin a sho ime pe iod p io o in ended commencemen o PSMA-RLT (n= 14/66) o in he cou se o PSMA-RLT (n= 52/66). The mean ime be ween bo h PET/CT scans was 7.3 ± 10.7 Rosa e al. EJNMMI Resea ch (2020) 10:52 Page 2 o 10 days (95% con idence in e al o he mean (CI) [4.6; 9.9]). The mean age o he pa ien s was 69 yea s [ ange 45–89 yea s]. All pa ien s ecei ed se e al p e ea men s. De ailed in o ma ion abou he p e ea men s and he pa ien cha - ac e is ics is p esen ed in Table 1. And ogen dep i a ion he apy (ADT) was con inued unchanged in all pa ien s o a oid a ia ion o PSMA exp ession. [ 68 Ga]Ga-PSMA-11 and [ 18 F]FDG PET/CT we e pe o med on a compassiona e use basis unde he Ge man Pha maceu ical Ac §13 (2b). Pa ien s ga e w i en consen a e being ho oughly in- o med abou he isks and po en ial side e ec s o his in e en ion. Addi ionally, pa ien s consen ed o publica ion o any esul ing da a in acco dance wi h he Decla a ion o Helsinki. Re ospec i e analysis app o al was wai ed by he local ins i u ional e iew boa d. PET acquisi ion and analysis Fo PET imaging, a mean ac i i y o 124.1 ± 14.4 MBq [ 68 Ga]Ga-PSMA-11 (CI [120.6; 127.6]) and 268.6 ± 28.7 MBq [ 18 F]FDG (CI [261.6; 275.7]) was adminis e ed, ollowed by a 500-ml in usion o NaCl 0.9%. Fas ing mean blood glucose alue was 98.1 ± 17.3 mg/dl (CI [93.8; 102.4]) be o e adminis a ion o [ 18 F]FDG. The mean up ake ime was app oxima ely 60 min (61.8 ± 6.6 min, CI [60.1; 63.4]) o [ 68 Ga]Ga-PSMA-11 acco ding o s anda d p ocedu es o p os a e cance imaging [25] and 90 min (91.6 ± 8.7 min, CI [89.4; 93.7]) o [ 18 F]FDG, acco ding o he ou s anda d p ocedu e and Ge man guideline o umo imaging [26]. Be o e da a acquisi ion, all pa ien s we e ad ised o emp y hei bladde . No diu e ics we e applied. All PET/CT scans we e pe o med using a Biog aph 40 mCT PET/CT scanne (Siemens Medical Solu ions, Knox ille, TN, USA) wi h EANM Resea ch L d. acc edi a ion. The PET acquisi ion was pe o med om e ex o mid- emu wi h 3-min acquisi ion ime pe bed posi ion. A bed pos- i ion co e s 21.4-cm ex ended ield-o - iew (T ueV). The PET da ase s we e econs uc ed using an i e a i e 3-dimensional OSEM (o de ed-subse expec a ion maximiza ion) algo i hm (3 i e a ions; 24 subse s) wi h Gaussian il e ing and a slice hickness o 5 mm. Ran- dom co ec ion, decay co ec ion, sca e a enua ion, and a enua ion co ec ion we e applied. The CT was pe o med in low-dose echnique using an X- ay ube ol age o 120 keV and a modula ion o he ube cu en by applying CARE Dose4D wi h a maximal ube cu en - ime p oduc o 30 mAs. All PET/CT da a se s we e isu- ally analyzed using ce i ied analysis so wa e (Sec a PACS—Sec a Medical Sys ems GmbH, Cologne, Ge many). [ 18 F]FDG and [ 68 Ga]Ga-PSMA-11 PET/CT we e ead simul aneously by h ee expe ienced physicians (a leas 5 yea s o expe ience in PET eading) sea ching o misma ch indings. A misma ch inding was de ined as me as asis wi h ema kable [ 18 F]FDG up ake and no o conside ably less conco dan [ 68 Ga]Ga-PSMA-11 up ake based on isual analysis. The decision o a misma ch ind- ing was aken in consensus o all PET eade s. Se um pa ame e s Selec ed se um pa ame e s used in ou clinical p ac ice we e es ed as indica o s o he occu ence o [ 18 F]FDG/[ 68 Ga]Ga- PSMA-11 misma ch indings in PET/CT: p os a e-speci ic an igen (PSA), neu on-speci ic enolase (NSE), gamma- glu amyl ans e ase (GGT), and alkaline phospha ase (ALP). Absolu e alues we e es ed in all 66 pa ien s. Addi ionally, ela i e changes (ΔPSA, ΔNSE, ΔGGT, ΔALP) o e a pe iod o 4 ± 1 weeks p io o [ 18 F]FDG PET/CT we e analyzed in 55/66 pa ien s (83.3%). In 11/66 cases (16.7%) no labo a o y da a om p e ious blood samples we e a ailable. S a is ical analysis and sco ing sys em Fo s a is ical analysis, Mann-Whi ney U es was ap- plied using P ism 8 (G aphPad So wa e, San Diego, Table 1 Pa ien cha ac e is ics Cha ac e is ic Value Age [yea s] 69 (45–89) PSA [ng/dl] 70 (0.3–4742) Time om ini ial diagnosis [yea s] ≤2 18 (27.3%) >2 o≤5 20 (30.3%) > 5 28 (42.4%) P io he apy P os a ec omy 29 (44%) Radia ion 39 (59%) ADT 66 (100%) Enzalu amide 50 (76%) Abi a e one 47 (71%) Doce axel 37 (56%) Cabazi axel 24 (36%) Xo igo 7 (11%) ECOG PS be o e i s cycle 0 18 (27%) 1 41 (62%) 2 5 (8%) 3 2 (3%) Si e o me as asis Bone 60 (91%) Lymph node 49 (74%) Li e 29 (44%) Lung 10 (15%) Da a a e p esen ed as n(%) o median ( ange) Abb e ia ions:PSA p os a e-speci ic an igen, ECOG PS Eas e n Coope a i e Oncology G oup Pe o mance S a us, ADT and ogen dep i a ion he apy Rosa e al. EJNMMI Resea ch (2020) 10:52 Page 3 o 10 USA) o de e mine signi ican di e ences be ween he g oups. A p alue < 0.05 was ega ded as s a is ically sig- ni ican . Fo powe calcula ion o Mann-Whi ney U es , we calcula ed he e ec size; alues highe han 0.5 we e conside ed as a s ong e ec size. Fo each pa ame e ha u ned ou o be s a is ically signi ican , a ecei e ope a - ing cha ac e is ic (ROC) analysis was pe o med and a cu -o poin was de e mined using he maximal alue o Youden index. In addi ion, a sco ing sys em combining all signi ican pa ame e s was in oduced. This sco ing sys em was cons uc ed o ease o clinical use and on he basis o sensi i i y-speci ici y analysis o he s onges pa ame e , assis ed by he o he signi ican pa ame e s, enhancing he powe and accu acy o he sco e. Resul s In o al, 41/66 (62%) pa ien s included in his analysis e- ealed a leas one [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 mis- ma ch inding on PET/CT, and 25/66 (38%) had no misma ch indings. O e all, 390 misma ch lesions we e de ec ed: 211 in bone (in 24/41 pa ien s), 62 in lymph nodes (in 21/41 pa ien s), 100 in he li e (in 20/41 pa- ien s), 4 in he lung (in 1/41 pa ien s), and 13 in o he lo- ca ions (in 9/41 pa ien s). Figu e 1p esen s an example o a pa ien wi h mul iple misma ch indings in he li e . In he misma ch g oup, 13/41 (32%) pa ien s had hei mis- ma ch indings (121/390 misma ch lesions) de ec ed a baseline imaging be o e in ended commencemen o PSMA-RLT, whe eas in he emaining 28/41 (68%) pa- ien s he misma ch (269/390 misma ch lesions) was diag- nosed in he cou se o PRLT a e ha ing ecei ed a mean o 4 ± 2 cycles o PSMA-RLT. When compa ing hese 269 misma ch lesions o he ini ial [ 68 Ga]Ga-PSMA-11 PET/ CT be o e s a ing PSMA-RLT, 29/269 (11%) we e ini- ially in ensely PSMA-posi i e, 52/269 (19%) we e mode - a ely PSMA-posi i e, and he majo i y, 188/269 (70%), we e no iden i iable on ini ial [ 68 Ga]Ga-PSMA-11 PET/ CT. Figu e 2shows an example o wo misma ch lesions, one in ensely PSMA-posi i e and one no iden i iable on he ini ial [ 68 Ga]Ga-PSMA-11 PET/CT. NSE se um le el (55.4 ± 44.6 μg/l s.18.5 ± 8 μg/l, p< 0.001) and ΔNSE (93.8 ± 124.5% s.2.9 ± 39.5%, p< 0.001) we e signi ican ly highe in he misma ch g oup han in he non-misma ch g oup. No signi ican di e - ence was ound o se um PSA (p= 0.424), ΔPSA (p= 0.417), se um ALP (p= 0.937), and ΔALP (p= 0.611) be ween he misma ch and he non-misma ch g oup, e- spec i ely. Twen y-nine o 66 (44.0%) pa ien s had li e me as ases, and 20/29 had a leas one misma ch inding in he li e . In pa ien s wi h li e me as ases, GGT (p= 0.773) and ΔGGT (p= 0.971) also e ealed no signi i- can di e ence be ween he misma ch and he non- misma ch g oup. Fo NSE and ΔNSE, he e ec size was 0.70 and 0.59, which we e conside ed as s ong. Summa- ized s a is ics o he da a a e p esen ed in Table 2. All hese s a is ical compa isons a e illus a ed by box-plo diag am o ma o each es ed pa ame e (Fig. 3). A subg oup analysis compa ing he pa ien s diagnosed wi h a misma ch by sc eening o PSMA-RLT wi h he pa ien s diagnosed wi h a misma ch a e se e al cycles PSMA-RLT e ealed no signi ican di e ence o se um NSE alue (67.5 ± 51.7 μg/l s.49.9 ± 40.8 μg/l, p= 0.12). The p edic i e impac o NSE and ΔNSE ega ding misma ch de ec ion is disce nible om he wa e all plo s wi h highligh ing o misma ch and non-misma ch indi iduals (Fig. 4). ROC analyses o NSE and ΔNSE Fig. 1 Pa ien wi h hepa ic misma ch indings. Maximal in ensi y p ojec ion (MIP), PET/CT, and PET da a o [ 68 Ga]Ga-PSMA-11 PET/CT (a) and o [ 18 F]FDG PET/CT (b) Rosa e al. EJNMMI Resea ch (2020) 10:52 Page 4 o 10 e ealed an a ea unde he cu e (AUC) o 0.92 and 0.84, espec i ely (Fig. 5). Fo NSE, he maximum alue o he Youden index (J= 0.74) was a a se um le el o 26.8 μg/l wi h a sensi i i y o 78% and a speci ici y o 96% o he occu ence o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch lesions, whe eas he maximum alue o he Youden Index o ΔNSE (J= 0.59) was a + 13.9% in- c ease wi h a sensi i i y o 84% and a speci ici y o 75%. Only one pa ien had misma ch indings wi h se um NSE < 15 μg/l. On he o he hand, we obse ed se e al misma ch (13/41) and non-misma ch (16/25) pa ien s wi h NSE in he ange be ween 15 and 30 μg/l (Fig. 4a). To dicho omize hese pa ien s and o imp o e he sensi- i i y wi hou ma ked loss o speci ici y we in oduced a sco ing sys em (Combined NSE Sco e) based on bo h pa ame e s, which is p esen ed in Fig. 6a. The main pa o he sco ing sys em was assigned o he absolu e alue o NSE being he s onges pa ame e in ROC analysis. Ze o poin s we e gi en o NSE in he physiological ange o < 15 μg/l. Fo he ease o clinical use, NSE was classi ied in he c i ical ange (15–30 μg/l) in s eps o 5μg/l, s a ing wi h 1 poin o 15–20 μg/l up o 3 poin s o 25–30 μg/l co e ing he Youden’s index (26.8 ng/ml) o he sensi i i y-speci ici y analysis. G ea e in e al s eps we e chosen o NSE abo e 30 μg/l (4 poin s 30– 50 μg/l, 5 poin s 50–100 μg/l and 6 poin s > 100 μg/l). ΔNSE was addi ionally included o enhancing he powe and accu acy o he sco e. Addi ional poin s o ΔNSE was se o −1 poin when NSE was dec easing (mo e han −20 %), o 0 poin s wi h s able (−20 o + 20 Fig. 2 Example o wo misma ch lesions de ec ed a e 5 cycles o PSMA-RLT: one in ensely PSMA-posi i e (g een a ow) and one no iden i iable (blue a ow) lesion on he ini ial [ 68 Ga]Ga-PSMA-11 PET/CT a baseline p io o PSMA-RLT (a). [ 68 Ga]Ga-PSMA-11 PET/CT (b) and [ 18 F]FDG PET/CT (c) a e 5 cycles o PSMA-RLT showing [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch in bo h lesions Table 2 Desc ip i e s a is ics o se um pa ame e s G oup nMedian (IQR) Mean (± SD) Minimum Maximum p alue NSE [μg/l] Misma ch 41 35 (30.0–66.9) 55.4 (± 44.6) 15 188.6 < 0.001 Non-misma ch 25 16.9 (13.0–20.4) 18.5 (± 8.0) 10 50 PSA [ng/ml] Misma ch 41 168 (83.0–602.5) 367.6 (± 407.9) 0.3 1360 0.424 Non-misma ch 25 190 (116.8–743.0) 666.3 (± 1086.8) 3 4742 GGT [U/l] Misma ch 22 75 (52.5–184.5) 120.5 (± 104.5) 22 378 0.773 Non-misma ch 9 76 (30.0–163.0) 127 (± 149.9) 20 497 ALP [U/l] Misma ch 41 140 (86–242.5) 192.2 (± 163.5) 12 818 0.937 Non-misma ch 25 136 (83.5–306.5) 211.8 (± 209.6) 35 1042 ΔNSE [%] Misma ch 31 47 (18.6–137.3) 93.8 (± 124.5) −12 533 < 0.001 Non-misma ch 24 3 (−20.5 o 15.6) 2.9 (± 39.5) −62 140 ΔPSA [%] Misma ch 31 27 (−4.3 o 109.1) 254.7 (± 890.3) −60 4956 0.417 Non-misma ch 24 15 (−9.7 o 64.4) 63.8 (± 153.5) −89 631 ΔGGT [%] Misma ch 18 65.5 (30.3–143.8) 196.8 (± 464.9) −79 1986 0.971 Non-misma ch 9 70 (2.0–138.5) 205.3 (± 454.6) −31 1406 ΔALP [%] Misma ch 31 0 (−14.0 o 39.0) 35.3 (± 137.6) −33 753 0.611 Non-misma ch 24 12.5 (−13.8 o 40.0) 28.2 (± 76.4) −41 342 Abb e ia ions:NSE neu on-speci ic enolase, PSA p os a e-speci ic an igen, GGT gamma-glu amyl ans e ase, ALP alkaline phospha ase Rosa e al. EJNMMI Resea ch (2020) 10:52 Page 5 o 10 %) o unknown NSE, and o 1 (20–50%) o o 2 (> 50%) poin s o inc easing NSE. The ROC analysis o he de el- oped sco ing sys em (Fig. 6b) e ealed an AUC o 0.91 and a maximum alue o he Youden index o 0.78 a 3 sco ing poin s wi h a sensi i i y o 90% and a speci ici y o 88% o he occu ence o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch lesions. The gain o he Combined NSE Sco e is seen a he sensi i i y le el o abou 90%, whe e he iso- la ed use o NSE achie es only a speci ici y o 76%. Discussion We obse ed a signi ican ly highe absolu e se um con- cen a ion and a highe ela i e inc ease o NSE in ad- anced mCRPC pa ien s wi h [ 18 F]FDG-a id and insu icien PSMA exp essing me as ases ([ 18 F]FDG/ [ 68 Ga]Ga-PSMA-11 misma ch indings) in ou coho . NSE is a highly speci ic ma ke o neu ons and pe iph- e al neu oendoc ine cells, used as a bioma ke o agg es- si e o ms o neu oendoc ine umo s and small cell ca cinoma [27,28]. Ele a ed NSE se um le els ha e also been obse ed in neu oendoc ine sub ypes o p os a e ca - cinoma [29,30].Whileonlyasmallp opo iono p os a e cance pa ien s ep esen neu oendoc ine ea u es a he beginning o disease [31], mo e pa ien s de elop hese unde he apy [24]. Se um NSE migh be inc eased in pa- ien s wi h misma ch indings as a sign o dedi e en ia ion o ans o ma ion o a neu oendoc ine sub ype. This could be a mechanism o esis ance induced by selec i e ea - men p essu e and was pa icula ly obse ed in pa ien s unde going an i-and ogen he apy [32–34]. Neu oendo- c ine di e en ia ion in p os a e ca cinoma is a pheno ypic change by which p os a e cance cells ans-di e en ia e in o neu oendoc ine-like cells. These neu oendoc ine-like cells a e lacking exp ession o and ogen ecep o and p os a e-speci ic an igen. Neu oendoc ine-like cells a e known o p oduce pep ide ho mones and g ow h ac o s o p omo e umo p og ession and a e apop osis- esis an con ibu ing o ea men ailu e [35–37]. Some case e- po s desc ibed ha PSMA- a ge ed PET/CT may be no able o de ec neu oendoc ine p os a e ca cinoma [38–40]. Suppo ing hese obse a ions, Bakh e al. (2018) demon- s a ed in i o on pa ien -de i ed xenog a (PDX) an in- e se co ela ion be ween PSMA gene (FOLH1) and neu oendoc ine bioma ke gene exp ession [41]. Pa icu- la ly, supp ession o he PSMA gene was obse ed in 65% o cases which o e exp essed he NSE gene (ENO2). They also demons a ed ha he mos p og essions’pa hway o neu oendoc ine ans-di e en ia ion unde ongoing Fig. 3 Compa ison o absolu e se um alues o NSE (a), PSA (b) and ALP (c) be ween all misma ch- and non-misma ch pa ien s. Compa ison o se um GGT (d) o pa ien s ha ing misma ch and non-misma ch li e me as ases. Rela i e change o each pa ame e : ΔNSE (e), ΔPSA ( ), ΔALP (g), ΔGGT (h). Ex eme ou lie s a e no shown Rosa e al. EJNMMI Resea ch (2020) 10:52 Page 6 o 10 Fig. 4 Wa e all plo o NSE (a) and ΔNSE (b) alues in descending o de and colo coding in o misma ch ( ed) and a non-misma ch (blue) Fig. 5 ROC cu es o misma ch p edic ion by se um NSE (a) and ΔNSE (b) wi h maximum alue o he Youden Index (J) Rosa e al. EJNMMI Resea ch (2020) 10:52 Page 7 o 10 ea men was associa ed subsequen ly wi h loss o he PSMA exp ession. Thus, al e na i e molecula imaging me hods a e equi ed o neu oendoc ine pheno ype. The glucose anspo e GLUT1 has been ound o be o e exp essed on cells o he ad anced la e-s age p os- a e cance and i s exp ession was ela ed o umo ag- g essi eness [42]. Meziou e al. (2020) assumed ha GLUT1 exp ession may be inc eased in neu oendoc ine p os a e ca cinoma sugges ing ha [ 18 F]FDG PET/CT migh be an impo an imaging ool o examine neu o- endoc ine p os a e ca cinoma [43]. Few s udies demon- s a ed he clinical u ili y o [ 18 F]FDG PET/CT in neu oendoc ine p os a e ca cinoma [44–46]. Sp a e al. demons a ed in 23 pa ien s wi h neu oendoc ine p os- a e cance ha [ 18 F]FDG PET/CT has clinical bene i and epo ed a high de ec ion a e o me as a ic disease, especially o lymph node and isce al me as ases [46]. This hypo hesis o ans o ma ion o p os a e cance wi h neu oendoc ine ea u es in ou misma ch coho was con i med his opa hologically in one pa ien only. Mo e e idence is equi ed o con i m his hypo hesis. Thus, consis en his opa hological examina ion o mis- ma ch me as ases should be pe o med in u u e s udies. Pa ien s wi h mCRPC, p esen ing disco dan [ 18 F]FDG- a id lesions wi h low o no PSMA exp ession, ha e a poo p ognosis wi h e y sho su i al ime [47]. This misma ch phenomenon indica es a mo e agg essi e ype o mCRPC in clinical ou ine and p ecludes pa ien s om PSMA- di ec ed adioligand he apy [9]. Thus, ea ly diagnosis o misma ch indings is e y impo an be o e and du ing PSMA-RLT o p o ide hese pa ien s wi h al e na i e he - apy op ions in addi ion o o ins ead o PSMA-RLT, includ- ing chemo he apy, immuno he apy, bone-seeking adiopha maceu icals, PARP inhibi ion, and o he no el a ge ed ea men s [18,47]. Ou esul s ob ained om 66 mCRPC pa ien s sugges ha se um NSE is a po en ial bioma ke o he exis - ence o he desc ibed [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 mis- ma ch. S a is ical analysis o ou da a p oposed an NSE cu -o alue o a misma ch occu ence o 26.8 μg/l (sensi i i y 78%, speci ici y 96%) and o + 13.9 % inc ease o ΔNSE (sensi i i y 84%, speci ici y 75%). A combin- a ion o bo h pa ame e s may imp o e he p edic i e powe . We achie ed a sensi i i y and speci ici y o al- mos 90% wi h ou in oduced Combined NSE Sco e (cu -o : o al poin s ≥3, Fig. 6). The simplici y o he sco e allows easy clinical applica ion as a guide o u - he diagnos ic p ocedu es in he mCRPC se ing. In con as o NSE, o he se um pa ame e s analyzed in his s udy as PSA, ALP, and GGT we e no able o in- dica e misma ch indings in ou coho . These bio- ma ke s may only e lec o al umo bu den gi ing no speci ic hin owa ds [ 18 F]FDG-a id lesions wi h low o missing PSMA exp ession. We sugges including NSE assessmen s in o ou ine blood es s o mCRPC pa ien s be o e and du ing PSMA- RLT. Howe e , hese esul s should be no iced wi h cau- ion conside ing a po en ial bias due o he e ospec i e Fig. 6 Combined NSE Sco e (a) and ROC cu e o misma ch p edic ion by he sco e (b) wi h maximum alue o he Youden index (J) Rosa e al. EJNMMI Resea ch (2020) 10:52 Page 8 o 10 s udy design and no - ep esen a i e pa ien coho . Qui e o en, [ 18 F]FDG PET/CT was pe o med when iable PSMA-nega i e me as ases we e suspec ed be o e o du - ing PSMA-RLT due o a wo sening cou se o disease o by hin in o he imaging me hods (CT, MRI). Conse- quen ly, mo e misma ch pa ien s (62% in ou coho ) han non-misma ch pa ien s we e included in ou s udy, esul - ing in a p eselec ed coho o mCRPC pa ien s no ep e- sen ing he no mal incidence o misma ch in candida es e e ed o PSMA-RLT. In a p ospec i e phase-II ial o PSMA-RLT, Ho man e al. (2019) epo ed an incidence o 16.3% (n=7/43) o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 mis- ma ch a ime o sc eening o PSMA-RLT [20]. This was he i s p ospec i e s udy which excluded pa ien s wi h misma ch lesions. A p ospec i e se ing including la ge non-biased pa ien coho s is necessa y o con i m ou indings. Fu he limi a ions a e he missing blood samples in 11/66 (16.7%) pa ien s p io o sc eening. Those pa- ien s we e included in he sco ing sys em as “unknown” ΔNSE, which e lec s a equen clinical si ua ion in mCRPC pa ien s e e ed o e alua ion o PSMA-RLT. As ano he poin o c i icism, no quan i a i e SUV h esh- old de ining low PSMA exp ession on [ 68 Ga]Ga-PSMA-11 PET/CT was se , which is also a global p oblem in li e a- u e su e ing om de ined cu -o s o adequa e PSMA exp ession. Fu he mo e, addi ional se um pa ame e s, which we e ecen ly epo ed as p ognos ic ma ke s in he mCRPC se ing o PSMA-RLT, such as he neu oendo- c ine ma ke ch omog anin A o he lac a e dehyd ogen- ase (LDH) [48–50], we e no a ailable o es ing in his s udy. Se um ch omog anin A alues migh also be highe in pa ien s wi h misma ch indings simila o NSE. LDH, which is a ma ke wi h s ong p ognos ic alue o esponse p edic ion o PSMA-RLT [48], migh also be a po en ial ma ke o misma ch. Bo h pa ame e s should hus also be es ed in u u e s udies. I hose s udies would indica e a misma ch p edic i e alue, hese pa ame e s could be in- cluded o he sco ing sys em o inc ease i s sensi i i y and speci ici y. Las ly, his opa hological con i ma ion o a ans- o ma ion p ocess in misma ch lesions owa ds a neu oen- doc ine pheno ype was pe o med only in one case. His opa hological examina ion o misma ch me as ases should be included in u u e p ospec i e s udies. Conclusions We obse ed a signi ican ly highe absolu e se um concen a ion and a highe ela i e inc ease o NSE in ad- anced mCRPC pa ien s wi h [ 18 F]FDG-a id and insu i- cien PSMA exp essing me as ases ([ 18 F]FDG/[ 68 Ga]Ga- PSMA-11 misma ch indings on PET/CT) in ou coho . NSE migh be used as a po en ial labo a o y indica o o [ 18 F]FDG/[ 68 Ga]Ga-PSMA-11 misma ch indings, i his obse a ion is con i med in u u e, ideally p ospec i e, s udies in la ge pa ien coho s. Abb e ia ions ADT: And ogen dep i a ion he apy; ALP: Alkaline phospha ase; CI: 95% con idence in e al o he mean; CT: Compu e omog aphy; FDG: Fluo odeoxyglucose; GGT: Gamma-glu amyl ans e ase; LDH: Lac a e dehyd ogenase; mCRPC: Me as asized cas a ion- esis an p os a e ca cinoma; NSE: Neu on-speci ic enolase; PET: Posi on emission omog aphy; PSA: P os a e-speci ic an igen; PSMA: P os a e-speci ic memb ane an igen; PSMA-RLT: PSMA- a ge ed adioligand he apy; ROC: Recei e ope a ing cha ac e is ic Acknowledgemen s No applicable Au ho s’con ibu ions Flo ian Rosa , Kalle Ribba , Ma hias Sch eckenbe ge , Same Ezziddin, and Fadi Kh eish con ibu ed o he design o he s udy. Ma in Ries and Fadi Kh eish pe o med da a acquisi ion wi h suppo om Ma k Ba holomä and S ephan Maus. Flo ian Rosa , Kalle Ribba , and Johannes Linxweile analyzed he da a. Flo ian Rosa and Kalle Ribba d a ed his pape , which was e ised by Ma k Ba holomä, Same Ezziddin, and Fadi Kh eish. All au ho s app o ed he inal manusc ip . Funding No hing o disclose A ailabili y o da a and ma e ials The da ase s used and analyzed du ing he cu en s udy a e a ailable om he co esponding au ho on easonable eques . Compe ing in e es The au ho s decla e ha hey ha e no con lic o in e es . E hics app o al and consen o pa icipa e All p ocedu es pe o med in he pa ien s desc ibed he ein we e in acco dance wi h he e hical s anda ds o he Ins i u ional and/o Na ional Resea ch E hics Commi ees and wi h he 1964 Helsinki Decla a ion and i s la e amendmen s, o wi h compa able e hical s anda ds. This epo does no include any animal s udies. W i en in o med consen was ob ained om all s udy pa icipan s Consen o publica ion All pa ien s ha e gi en w i en consen o publica ion. 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