液體活檢-p在腫瘤精準(zhǔn)治療中研究進(jìn)展

內(nèi)容研究進(jìn)展一.液體活檢1）cfDNA&ctDNA

&

CTC概念和2）液體活檢應(yīng)用和意義二.CTC/CTSC研究方法和進(jìn)展三.

的研究液態(tài)活檢來源于腫瘤組織，存在于血液中：循環(huán)腫瘤細(xì)胞（CTC）循環(huán)腫瘤DNA（ctDNA）腫瘤外泌體exosome提示腫瘤發(fā)展進(jìn)程及抗藥性等信息，指導(dǎo)化精準(zhǔn)治療Diaz

LA

Jr,

Bardelli

A.Liquid

Biopsies:

Genoty579–586Circulating

Tumor

DNA.

J

Clin

Oncol.

2014;

32(6):cfDNAWhat:循環(huán)游離DNA，機(jī)體細(xì)胞入外周血循環(huán)后發(fā)生部分降解的內(nèi)源性DNA。高度片段化，約170bp。Source:正常細(xì)胞、異常細(xì)胞（如腫瘤細(xì)胞）， DNA等State:半衰期短，動態(tài)表達(dá)量，核小體以單個(gè)、雙聯(lián)或三聯(lián)的形式進(jìn)入血液，并逐步分解。cfDNA（cell-free

DNA）ctDNAWhat:循環(huán)腫瘤DNA，是指腫瘤細(xì)胞DNA經(jīng)脫落或細(xì)胞凋亡后進(jìn)入循環(huán)系種特征性的腫瘤生物標(biāo)記物，可以被定性、定量和追蹤。

Source:來源于腫瘤細(xì)胞的cfDNA，屬于cfDNA的一種類型。不同患者的水平差異顯著，受腫瘤負(fù)荷，分期等因素的影響ctDNA（circulating

tumorDNA）cfDNA

&

ctDNAEllen

Heitzer,Peter

Ulz,and

Jochen

B.Geig.Circulating

Tumor

DNA

as

a

Liquid

Biopsy

for

Cancer.Clinical

Chemistry

.

2015,112–1231

：

CAPP-Seq

（

cancerby

deep

sequencing

）alized個(gè)profiling體化深度NimbleGen分析方法:利用定制化的SeqC Z

Choice

Library作為”篩選器“對樣本靶向捕獲后再深度測序，靈敏度高，特異性強(qiáng)且經(jīng)濟(jì)可行。2：利用數(shù)字錯(cuò)誤抑制（

IDES

）的方法和

CAPP-seq（ctDNA

方法），能夠檢測到頻率低至0.004％的突變等位AM

,

Lovejoy

A

F

,

Klass

DM,

et

al.

Integrated

digital

error

suppression

for

improved

detectionof

circulating

tumor

DNA.

Nature

Biotechnology(2016)

:10.1038/nbt.3520ctDNA-高通量cfDN段譜系分析Deep

sequencingofcirculating

cell-freeDNA

yields

a

dense,genome-wide

map

ofnucleosomeoccupancythat

enablesidenti?cationofits

cell-types

oforigin,potentially

enabling

thenoninvasive

monitoring

of

a

broader

set

of

clinical

conditionsMatthew

W.

Snyder,

Martin

Kircher,Andrew

J.

Hillet

al.

Cell-free

DNA

Comprises

an

In

Vivo

NucleosomeFootprint

that

Informs

Its

Tissues-Of-Origin.Cell.

2016

Jan

14;164(1-2):57-68.CTCWhat：循環(huán)腫瘤細(xì)胞，是指從實(shí)體瘤中脫離出來并進(jìn)入外周血液循環(huán)的腫瘤細(xì)胞。Source:腫瘤和轉(zhuǎn)移灶都會CTC?？蓹z測到的CTC是腫瘤細(xì)胞在循環(huán)系統(tǒng)微環(huán)境下生存競爭的結(jié)果。CTC（circulating

tumorcells）液體活檢應(yīng)用價(jià)值（監(jiān)測腫瘤特異性突變發(fā)現(xiàn)腫瘤復(fù)發(fā)）Ellen

Heitzer,Peter

Ulz,and

Jochen

B.Geig.Circulating

Tumor

DNA

as

a

Liquid

Biopsy

for

Cancer.Clinical

Chemistry

.

2015,112–123液體活檢應(yīng)用價(jià)值（分析耐藥株的遺傳變異）Ellen

Heitzer,Peter

Ulz,and

Jochen

B.Geig.Circulating

Tumor

DNA

as

a

Liquid

Biopsy

for

Cancer.Clinical

Chemistry

.

2015,112–123液體活檢優(yōu)勢及前景液態(tài)活檢無侵入性、可頻繁多次檢測及快速反應(yīng)能力均體現(xiàn)出發(fā)展?jié)摿薮髴?yīng)用價(jià)值。將削減無效腫瘤醫(yī)療開支、降低醫(yī)療成本。適應(yīng)癥廣泛，如

、結(jié)直腸癌、肺癌、胃癌、食管癌等常見腫瘤均可用液態(tài)活檢技術(shù)進(jìn)行

與監(jiān)測。2015年

發(fā)布的腫瘤治療計(jì)劃的四大舉措之一：“液態(tài)活檢”血漿開發(fā)新方法來評估治療反應(yīng)以及抵抗可能的耐藥性。根據(jù)中國國家

中心發(fā)布的數(shù)據(jù)，我國5年內(nèi)

為

且仍存活的病例數(shù)約為749萬。也是液態(tài)活檢巨大的市場。CTC研究概況CTC與轉(zhuǎn)移的關(guān)系CTC異質(zhì)性與腫瘤治療CTC應(yīng)用和意義CTC研究方法CTCs

(circulating

tumor

cells,

CTCs)Ashworth早在1896年就提出了循環(huán)腫瘤細(xì)胞的概念。侵入循環(huán)系統(tǒng)的腫瘤細(xì)胞，大部分由于機(jī)體的免疫識別、機(jī)械及自身凋亡在短期內(nèi)，只有極少數(shù)存，在適于自身存活和增殖的或組織形成轉(zhuǎn)移灶。CTC與腫瘤轉(zhuǎn)移Simon

A

Joosse,

Tobias

M

G es,

and

Klaus

Pan .

Biology,

detection,

and

clinical

implications

ofcirculating

tumor

cells.

EMBO

Mol

Med.

2015

;

7(1):

1–11.Brouwer

A,

De

Laere

B,

Peeters

D,

et

al.consequences

of

heterogeneity

in

the

circulating

tumor

cellcompartment.

Onco .

2016Mar

9. :10.18632/onco

.8015.CTC異質(zhì)性CTC應(yīng)用和意義CTCs腫瘤轉(zhuǎn)移腫瘤治療CTCs數(shù)量改變反映了腫瘤細(xì)胞對各種治療方案的敏感性和有效性，為

化治療提供依據(jù)。監(jiān)測早期轉(zhuǎn)移、

癌結(jié)腸癌、肺癌、胃癌、肝癌、胰

多種腫瘤監(jiān)測CTCs

表達(dá)／突變，找到藥物敏感的靶點(diǎn)，指導(dǎo)

化治療CTC優(yōu)化治療方案2014年GoldKorn，接受多西他賽聯(lián)合阿曲生坦治療的轉(zhuǎn)移性去勢抵抗性

癌患者，基線循環(huán)腫瘤細(xì)胞CTC數(shù)量可預(yù)后，3周時(shí)CTC升高提示總生存較差，有望作為重新指導(dǎo)和優(yōu)化治療的較好指標(biāo)。Goldkorn

A，Ely

B，Quinn

DI，et

al．Circulating

Tumo

cell

counts

are

Prognostic

of

overall

survival

in

SWOG

S0421：a

Phase

IIITrial

of

docetaxel

with

or

without

atrasentan

For

Metastatic

castration

resistant

prostate

cancer.J

Clin

Oncol2014

32(11)：1136-1142．Bian

L，Wang

T，Liu

Y，et

al．Evaluation

of

treatment

response

for

breast

cancer：are

we

Entering

the

eraof

”biological

Complete

remission”?．Chin

J

Cancer

Res．2012，24：403-407．2012年Bian,乳癌患者在接受化療（白蛋白結(jié)合型紫杉醇+卡培他濱+曲妥珠單抗）前后CTC變化。治療前：CTC

3，HER-2+；1療程：CTC

0。6療程：病理學(xué)CR。證實(shí)CTC值在

治療早期發(fā)生顯著變化，能夠準(zhǔn)確預(yù)示療效，能夠科學(xué)指導(dǎo)選擇治療方案。CTCs的研究方法Ramdane

Harouaka

,

Zhigang

Kang

,Si-Zheng

,

Liang

Cao.

Circulating

tumor

cells:

Advances

inysis,

and

challenges

for

clinical

applications.

Pharmacology

&

Ther

utics.2014;isolation

and141:209–221.研究方法（1）-Cell

SearchCellSearch系統(tǒng)是唯一經(jīng)FDA批準(zhǔn)進(jìn)入臨床應(yīng)用的CTCs檢測方案。原理：EpCAM抗體標(biāo)識的微磁珠從全血中捕獲細(xì)胞，用免疫熒光識EpCAM+CK+CD45?進(jìn)行磁性分離。CTCEpCAMCKDAPICD45CellSearch數(shù)據(jù)解讀:7.5ML外周血判斷閾值：轉(zhuǎn)移性CTCs≥5

細(xì)胞癌CTCs

≥5細(xì)胞結(jié)腸癌：CTCs≥3細(xì)胞表明預(yù)后不良3.目前FDA只批準(zhǔn)用于以上三種腫瘤CTCs的研究方法（2）-

CTCs的研究方法（3）-Xiaohui

Nia,

Minglei

Zhuo,

Zhe

Sua，et

al.

Reproducible

copy

number

variation

patterns

among

singlecirculating

tumor

cells

of

lung

cancer

patients.

PNA,2013,52(110)

21083-21088.Xiaohui

Nia,

Minglei

Zhuo,

Zhe

Sua，et

al.

Reproducible

copy

number

variation

patterns

among

singlecirculating

tumor

cells

of

lung

cancer

patients.

PNA,2013,52(110)

21083-21088.Ramdane

Harouaka

,

Zhigang

Kang

,

Si-Zheng

,

Liang

Cao.

Circulating

tumor

cells:

Advances

inysis,

and

challenges

for

clinical

applications.

Pharmacology

&

Ther

utics.2014;isolation

and141:209–221.CTCs的分離方法（1）CTCs的分離方法（2）-體外培養(yǎng)Min

Yu

,

Aditya

Bardia

,

Nicola

Aceto,

et

al.

Ex

vivo

culture

of

circulatingbreast

tumor

cellsforindividualized

testingof

drug

susceptibility.

Science.

2014

;

345(6193):

216–220.Papanicolaou

staining

foruncultured

primaryRepresentative

images

ofnonadherent

CTCimmunofluorescentstaining

：CK：

red；Ki67：yellow；CD45

：green；DAPI：

blue.CTSC研究概況CTSC定義CSC,CTC,CTSC三者關(guān)系CTSC研究進(jìn)展CTSC研究難點(diǎn)和未來方向CTSC背景介紹循環(huán)腫瘤干細(xì)胞（circulating

tumor

stem

cells,CTSCs）：CTCs中具有干細(xì)胞特性，即自我更新及分化潛能的一小群細(xì)胞。作為標(biāo)志物CTSC具有更重要的臨床價(jià)值。EMT可使CTCs獲得干細(xì)胞特性，而具有藥物耐受性和高侵襲性。CTSC已在包括肝癌，

，胃癌，結(jié)直腸癌等多種腫瘤中被證實(shí)。CSC

,CTC,CTSC（腫瘤轉(zhuǎn)移等級結(jié)構(gòu)圖）MH,

Imrali

A,

Heeschen

C.Circulating

cancer

stem

cells:

the

importance

to

select.Chin

J

Cancer

Res.2015

Oct;27(5):437-49.MarkersforCTSCMH,

Imrali

A,

Heeschen

C.Circulating

cancer

stem

cells:

the

importance

to

select.Chin

J

Cancer

Res.2015

Oct;27(5):437-49.CTSC研究進(jìn)展學(xué)者們采用不同的表面標(biāo)志物組合來定義CTSC：Epcam/CK/CD44／CD24；

Epcam/CK/ALDH1／CD24Epcam/CK/Vimentin/Fibernectin；Epcam/CK/TWISTI/Akt2PDKaEpcam/Muc1/HER2Baccelli

I，Schneeweiss

A，Riethdorf

S，et

a1．Identification

of

a

population

of

blood

circulating

tumor

cells

from

breast

cancerpatients

that

initiates

metastasis

in

axenografiassay[J]．Nat

Biotechnol，2013，31(6)：539．544．Kasimir—Bauer

S，Hoffmann0，Wallwiener

D，et

a1．Expression

Of

stem

cell

and

epithelial—mesenchymal

transition

markersinPrimary

breast

cancer

Patients

With

circulating

tumor

cells

[J]．Breast

Cancer

Res，2012，14(1)：R15．CTSC研究進(jìn)展Yu

M．Bardia

A，Wittner

BS．et

a1．Circulating

breast

tumor

cells

Exhibit

dynamic

changes

InEpithelial

and

mesenehymal

eompostion[J]．Science，2013，339(6119)：580-584．EPCAM

low

MET

highCD47

high

CD44

high2013年復(fù)旦大學(xué)附屬肝癌

及研究組通過研究123例肝癌患者CTCs相關(guān)腫瘤干細(xì)胞免表標(biāo)記物，包括CD133、ABCG2、CD90、β-catenin、E-cadherin以及

Vimentin。2013年Liu等鑒于ICAM-1可能是肝癌干細(xì)胞標(biāo)志物，也采用流式細(xì)胞術(shù)檢測了肝癌血樣，發(fā)現(xiàn)部分病例可檢出CD45-ICAM-1+細(xì)胞，這樣的細(xì)胞具有較強(qiáng)的體外成球和體內(nèi)致瘤能力。Y.F.,

et

al.,

Circulating

stem

cell-like

epithelial

cell

adhesion

molecule-positive

tumor

cells

indicate

poor

prognosis

of

hepatocellularcarcinoma

after

curative

resection.

Hepatology,

2013.

57(4):

p.

1458-68。Liu

S,

Li

N,

Yu

X,

et

al.

Expression

of

intercellular

adhesionmolecule

1

by

hepatocellular

carcinoma

stem

cells

and

circulating

tumorcells[J].

Gastroenterology,

2013,

144(5):1031-1041肝癌CTSCs研究進(jìn)展的研究CSC

andmiRNACTC

andCTSC項(xiàng)目研究基礎(chǔ)—前期基金支持1.2014年福建省衛(wèi)教聯(lián)合項(xiàng)目：結(jié)直腸癌循環(huán)腫瘤細(xì)胞和腫瘤干細(xì)胞的分子機(jī)制研究（WKJ-FJ-14）個(gè)2012年福建省科技廳重點(diǎn)項(xiàng)目:《循環(huán)腫瘤細(xì)胞與腫瘤干細(xì)胞在體化治療中的應(yīng)用研究》（2012Y0017）；2012年福建省衛(wèi)生廳青年課題:《結(jié)直腸癌循環(huán)腫瘤細(xì)胞及其上皮間質(zhì)變標(biāo)志物的定量檢測》（2012-2-13）；2012年福建省自然科學(xué)基金項(xiàng)目:《miR-148

-靶蛋白網(wǎng)絡(luò)在肝癌干細(xì)胞中調(diào)控機(jī)制研究》（2012J05138）；2011年福建省醫(yī)學(xué)創(chuàng)新課題:《

循環(huán)腫瘤細(xì)胞（CTC）及其單細(xì)胞多的臨床意義》（2011-CX-16）；6.

2011年福建省自然科學(xué)基金項(xiàng)目:《循環(huán)腫瘤細(xì)胞及其耐藥表達(dá)水平在晚期化療敏感性中的應(yīng)用研究》（2011J01141）－基金支持2016年福建省自然科學(xué)基金項(xiàng)目：《miR-194調(diào)控肝癌干細(xì)胞的靶 和

LncRNA

鑒定》（2016J01436）2009年福建省自然科學(xué)基金項(xiàng)目:《靶向

干細(xì)胞的樹突狀瘤苗抗腫瘤的研究》(2009J01118)2008年福建省自然科學(xué)基金項(xiàng)目:《miRNA靶向肝癌SP、NSP細(xì)胞

差異表達(dá)的研究》（2008J0077）MoFlo速流式細(xì)胞分選系統(tǒng)稀有細(xì)胞檢測的靈敏度高分選速度快研究基礎(chǔ)設(shè)備CSC-

Side

population(SP)

assayIt

is

generally

accepted

that

SP

cellspossess

stem

cellcharacteristics.Critical:

Optimize

Hoechst

33342

stainingprocedure

to

achieve

a

consistent

andtumorigenic

SP

subtype

as

SP

approach

isbased

on

the

functional

property

of

CSCs

toexclude

Hoechst-33342-dye

via

ABCtransporters.SPNSP-MoFloysis

and

sorting-Sphere

formation

assayFloating

sphereswereformed

bySP

cells,

while

none

byNSP

cellsHep-3BPLC/PRF/5Huh-7實(shí)驗(yàn)數(shù)據(jù)-Tumors

formed

in

NOD/SCID

micePLC/PRF/5Huh-7Hep-3BWeek

12;1×103

cells;SP:

blue

arrow;NSP:

redarrow.miRNA

profiling

result

by

RT-qPCRmiRNA

IDSP/NSP

Fold

changesP<0.05miR-9-3miR-9*-2.41miR-23b-2.12miR-27b-2.35miR-96-2.8miR-151-3p-2.65miR-151-5p-2.57miR-183*-2.23miR-183-3.16miR-194-2.03miR-146a-3.44miR-148b-3.14miR-340-3p-2.85miR-421-2.32miRNA

IDSP/NSP

Fold

changesP<0.05miR-135b*-3.55miR-489-3.28miR-15a-2.79miR-21-2.67miR-22-2.5miR-365-2.47miR-450a-5p-2.46miR-7a-2.44miR-26b-2.28miR-125a-5p-2.27miR-25-2.22miR-

126-3p-2.15miR-182-2.14A

total

of

370

miRNA

species

were yzed

in

PLC/PRF/5 SP

and

NSP

cells,

27

miRNAswere

identified

as

differentially

expressed,

all

of

which

were

down-regulated

in

SP

.實(shí)驗(yàn)數(shù)據(jù)-miR-148b在SP中低表達(dá)實(shí)驗(yàn)數(shù)據(jù)--miR-148b改變SP比例實(shí)驗(yàn)數(shù)據(jù)--miR-148b改變增殖與耐藥實(shí)驗(yàn)數(shù)據(jù)-miR-148b改變遷移侵襲與血管生成實(shí)驗(yàn)數(shù)據(jù)-miR-148b過表達(dá)抑制腫瘤生成實(shí)驗(yàn)數(shù)據(jù)-miR-148b沉默促進(jìn)腫瘤生成研究方案——CTC/CTSC研究方案——分選流程已完成200例鼻咽癌患者CTC檢測，100例患者CTC檢測，100例肺癌患者CTC檢測，150例直腸癌患者CTC/CTSC檢測，具備完善的CTC檢測技術(shù)流程。在研項(xiàng)目：150例初診未治腸癌患者，

其全病程臨床資料并抽取各階段血樣，分析患者各個(gè)時(shí)期

CTCs/CTSCs

數(shù)

值

，

以

PSF

為

觀

察

點(diǎn)

，

評

估

CTCs/CTSCs數(shù)值與臨床分期、預(yù)后、治療方案及療效等情況的相關(guān)性，

尤其是手術(shù)配合細(xì)胞治療的療效觀察，開發(fā)CTC/CTSC在療效評估中的應(yīng)用。研究數(shù)據(jù)入組初診未治腸癌患者150例患者全程臨床資料并抽取各階段血樣（TIL組：根治術(shù)前、術(shù)后第一次細(xì)胞治療后、細(xì)胞治療2個(gè)周期后、細(xì)胞治療3個(gè)周期后……對照組：根治術(shù)前、術(shù)后7天、術(shù)后1月，術(shù)后2月……每月一次）利用Moflo對CTCs/CTSCs計(jì)數(shù)患者各個(gè)時(shí)期CTCs/CTSCs數(shù)值與臨床分期、預(yù)后、治療方案及療效等情況相關(guān)性以PSF為觀察點(diǎn)，評估手術(shù)配合細(xì)胞治療的療效觀察難點(diǎn)和CTC/CTSC研究難點(diǎn)稀有性：僅占PBMC的1/106活性：難以保持細(xì)胞活性進(jìn)行后續(xù)研究單細(xì)胞性：細(xì)胞稀少難以進(jìn)行后續(xù)研究分選精度可達(dá)到1/106可保持分選后細(xì)胞的活性進(jìn)行單細(xì)胞水平RNA

提取和預(yù)擴(kuò)增，保證下游基因表達(dá)譜分析的質(zhì)量MoFlo速流式細(xì)胞分選系統(tǒng)REPLI-gWTASingleCell

Kit本項(xiàng)目關(guān)鍵技術(shù)流式分析CTC敏感性專一性檢測研究數(shù)據(jù)研究數(shù)據(jù)CTCs的畫門模式研究數(shù)據(jù)CTSCs的畫門模式CTCs的檢測研究數(shù)據(jù)研究數(shù)據(jù)CTSCs的檢測首診初治結(jié)直腸癌患者CTC/CTSC檢測分析結(jié)果<3≧3Age(years)0.157≦6025(50.0)1015>6025(50.0)1312Sex0.205female20(40.0)1010man30(60.0)1317Tumor

size(cm)0.189<526(52.0)1313≧524(48.0)1014TNM

stage0.072I5(10.0)41II11(22.0)92III29(58.0)218IV5(10.0)14T

stage0.118T12(4.0)11T25(10.0)50T34(8.0)31T439(78.0)2613RLNM0.032N017(34.0)143N121(42.0)1011N212(24.0)102M

(metastasis)0.024M045(90.0)3411M15(10.0)14PatientCharacteristicsNumber(%)CTC、CTSC(CD44+)p-value研究數(shù)據(jù)結(jié)直腸癌患者轉(zhuǎn)移后化療期間CTC/CTSC變化研究數(shù)據(jù)CTC分選分選白細(xì)胞對照：DAPI+/CD45-/Epcam-分選CTC細(xì)胞：DAPI+/CD45-/Epcam+實(shí)驗(yàn)方案研究數(shù)據(jù)-表達(dá)譜聚類圖腫瘤組織

正常組織

CTC細(xì)胞

白細(xì)胞對照實(shí)驗(yàn)方法區(qū)域中

與表達(dá)的相關(guān)分析圖表達(dá)譜比較組雜交近年文章1. Qinying

Liu, mei

Xu,

Shenghong,

et

al.miRNA-148b

suppresses

hepatic

cancer

stem

cell

bying

neuropilin1.

Bioscience

Reports,

2015;35(4).2. Xu

YM,

Xie

YQ,

Wang

XR,

et

al.

Identification

of

cancer

stem

cells

from

hepetocellular

carcinoma

cell

linesand

their

related

microRNAs.Oncol

Rep.

2013.

30:

2056-62.3.，

，

,

等.

患者循環(huán)腫瘤細(xì)胞檢測的臨床意義.中華腫瘤防治雜志,

2015,22(13):1009-1013.4. Identification

of

perioperative

circulating

tumor

cells

and

potential

biomarkers

of

circulating

tumor

stem

cellsin

colorectal

cancer

patients

by

flow

cytometry.待5.，等.

流式細(xì)胞術(shù)檢測鼻咽癌患者外周血循環(huán)腫瘤細(xì)胞及臨床相關(guān)性分析第八屆中國腫瘤學(xué)術(shù)大會暨第十三屆海峽 腫瘤學(xué)術(shù)會議

，2014.6.

Zheng

QH,

et

al.

Floating

cells

with

stem

cell

properties

in

gastric

cell

line

SGC-7901.

Tumori.

2011.

97:393-399.Zheng

QH,XuYM,

WeiZQ,

et

al.Isolation

andpurification

of

cancer

stem-like

cells

fromcell

linePLC/PRF-5.Chinese

Tumor

Biological

Therapy

Journal,

2011,

18(2):126-132.Lai

L,

Cui

C,

Jin

J,

et

al.

Mouse

embryonic

stem

cell-derived

thymic

epithelial

cell

progenitors

enhance

T-cellreconstitution

after

allogeneic

bone

marrow

transplantation.

Blood.

2011.

118(12):3410-3418.近年

文章Chen

G,

Wang

L,

Lu

J,

et

al.

Optical

Diagnosis

for

Lung

Cancer

Using

Multiphoton

Imaging.

Scanning.

2013.:10.1002/sca.21076.Yan

J,Tan

C,Gu

F,et

al.Sorafenib

delays

recurrence

and

metastasisafter

livertransplantation

in

a

ratmodel

ofhepatocellular

carcinoma

with

high

expression

of

pERK.

Liver

Transpl.

2013.

:10.1002/lt.23619.Chen

L,

Zhang

WW,

Zheng

QH,

et

al.

Aculeatusquinones

A-D,

novel

metabolites

from

the

marine-derivedfungus

Aspergillusaculeatus.

Heterocycles.

2013.

Published

online.Zheng

QF,

Wang

JJ,YingMG,et

al.Omentoplasty

inpreventing

anastomotic

leakage

ofoesophagogastrostomyfollowing

radical

oesophagectomy

with

three-field

lymphadenectomy.

Eur

J

Cardiothorac

Surg.

2013.

43(2):274-8.Chen

Y,

Zheng

X,

Chen

G,

et

al.

Immunoassay

for

LMP1

in

nasopharyngeal

tissue

based

on

surface-enhancedRaman

scattering.

Int

J

Nanomedicine.

2012.

7:

73-82.Yan

J,

Zhuo

S,

Chen

G,

et

al.

Preclinical

study

of

using

multiphoton

microscopy

to

diagnose

liver

cancer

anddifferentiate

benign

and

malignant

liver

lesions.

J

Biomed

Opt.

2012.

17(2):

026004.

Zhuo

S,

Yan

J,

Chen