THE ROLE OF MINIMAL THE ROLE OF MINIMAL RESIDUAL DISEASE (MRD) RESIDUAL DISEASE (MRD)

EVALUATION IN THE EVALUATION IN THE MANAGEMENT OF LYMPHOMASMANAGEMENT OF LYMPHOMAS

Sara Galimberti - Ematologia Pisa

DOCTORATE IN GENETICS,

ONCOLOGY AND CLINICAL MEDICINE

SUMMARY

NHL: what are they?MRD: what is it?

Which techniques? Which techniques? The FIL MRD NET experienceClinical aspects and role of MRD

I.

NHL: what are they?MRD: what is?

Which techniques? Which techniques? The FIL MRD NET experienceClinical aspects and role of MRD

NHLLymphomas represent 2,9% of all cancers in men

and 3,2% in women, with an incidence of about 20/100000/year. Every year in Italy more than 10000 new cases are diagnosed.

HL & NHL

NHL – CLINICAL FEATURES

Skin involvement

Mediastinum enlargement

Lymph nodes

NHL – CLINICAL FEATURES

Skin involvementMediastinum enlargement

Lymph nodes enlargement

Lymph nodes enlargement

Diagnosis is made by lymph node biopsylymph node biopsy. .

Often the node is superficial and easily accessible.

If it is deep in the abdomen, nodes can be biopsied If it is deep in the abdomen, nodes can be biopsied

under CT guidance or via the laparascope.

WHO CLASSIFICATION

WHO CLASSIFICATION

LIMPH NODE

MOLECULAR PATHWAYS in B -CELL LYMPHOMAGENESIS

GERMINAL CENTER

MARGINAL ZONE

PLASMA CELL

VIRGIN B-CELLS

Mantle cell lymphoma

BCL-1/JH

Primary effusion lymphoma

HHV-8

Lymphoplasmacytoid lymphoma

PAX-5

MALT lymphoma

API2/MLT

MANTLEMEMORY

B-CELS

BCL-2/JH

DLBCLp53

c-MYCp53

BCL-6 mutations Burkitt lymphoma

Follicular lymphoma

NHLMantle cell marginalBurkitt

lymphoplasmocyticfollicularDLBCL

ANN ARBOR STAGING SYSTEM

Stage I - disease in single lymph node or lymph node region.

Stage II - disease in two or more lymph node regions on sameside of diaphragm.

Stage III - disease in lymph node regions on both sides of thediaphragm are affected.

Stage IV - disease is wide spread, including multiple involvement at one or more extranodal (beyond the lymph node) sites, such as the bone marrow

ANN ARBOR STAGING SYSTEM

A: if there is no constitutional symptoms

B: if there is fever, weight loss, night sweats

II.

NHL: what are they?MRD: what is?

Which techniques? Which techniques? The FIL MRD NET experienceClinical aspects and role of MRD

MRD

IMAGINGIMAGING

CITOFLUORIMETRIACITOFLUORIMETRIA

t(14;18), t(11;14), t(8;14)t(14;18), t(11;14), t(8;14)

FLOW CYTOMETRYsIg intensity

dim bright

CD5 +/± CD5 + CD5 +/±

CD23 + / ±

CD23 +

CD23 –

CD10 –

CD10 ++ / ±

CLL

CD49c +

+ – – CD23 + / –

+

v-CLL PLL

MCL PLL

SMZL, SLVL,

IC, CyIg +/–

HCL, CD25 ++ CD11c ++ , CD103 +

FoLy

CD49 + / –

bcl-1, PRAD-1

CD43 – bcl-2 +

MCSS: 3.5–5

MCSS: 2.5–3

MCSS: 2

extra- MCSS

CLL MATUTES' SCORECLL = 3-5

TECHNIQUES

Max SENSITIVITYMax SENSITIVITYNO NEG FALSES

Max SPECIFICITYMax SPECIFICITYNO POS FALSES

TECHNIQUES

FLOW CYTOMETRY:FLOW CYTOMETRY:BM, PB, lymph nodesFew hoursFew hours10-3/10-4

Useful at diagnosis and follow-up

FLOW CYTOMETRY - CLL

CD5+/19+

FMC7-

TECHNIQUES

MOL BIOL:MOL BIOL:BM, PB, lymph nodes2 days2 days10-4/10-5

Useful at diagnosis and follow-up

PB vs BM

Pott, Blood 2006

BM vs PB

Pott, Blood 2010

BM vs PB in MCL at diagnosis: + 19%

TECHNIQUES

MOL BIOL:MOL BIOL:BM, PB, lymph nodes2 days2 days10-4/10-5

Useful at diagnosis and follow-up

BCL2/IGH

IGH REARRANGEMENT

ASO-PCR

Consensus primer 5’ Consensus primer 3’

FIRST PCR NOT TUMOR-SPECIFIC

FR1 FR2 FR4FR3CDR1 CDR2L CDR3

NOT TUMOR-SPECIFIC

FR1 FR2 FR4FR3CDR1 CDR2L CDR3

primer 3’ specific patientprimer 5’ specific patient

SECOND PCRTUMOR-SPECIFIC

TECHNIQUES

FLOW vs PCRFLOW vs PCR

92 samples analyzedby FC and ASO PCR

85% concordant

Sensitivity:

Bottcher, Leukemia 2004

Sensitivity:5 x 10-3 FC5 x 10-3-2 x 10-4 PCR

FIL-MRD NETWORK

ORBAL

CAND

TO

NO

GE

FI

PE

MO

REPR BO

RA

ANPG

TR

UD

VR

VI PD

ROZZANO HUMANITAS

BSMI

MONZA

TN

BZ

PV

PC

IVREA

BR

BA

TA

TRANI

NU

CA

ROMA

LT

NANOCERA INF

PA

CT

MERC

CZ

CS

RIONERO PT

TRICASE

NHL-Task 2 RESULTSMRD measurement of 8 Follow-up

LAB CODE

MRD-FU1 MRD-FU2 MRD-FU3 MRD-FU4 MRD-FU5 MRD-FU6 MRD-FU7 MRD-FU8

TO NEG POS POS NEG POS POS NEG POS

NHL-Task 2 RESULTSMRD measurement of 8 Follow-up

RO NEG POS POS

debole

NEG POS POS NEG POS

PI NEG POS POS NEG POS POS NEG POS

BO NEG POS POS NEG POS POS NEG POS

Task 3 RESULTS

LAB S QR SLOPE CORR.C. FU1 FU2 FU3 FU4 FU5 FU6 FU7 FU8

TO 1E-05 5E-05 3,64 1,00 1E-08 2E-01 6E-05 1E-08 2E-01 4E-04 1E-08 3E-04

borderline samples

QUANTITATIVE PCR

RO 1E-05 1E-04 3,7 0,98 1E-08 2E-01 1E-06 1E-08 4E-01

4E-04

1E-08 2E-04

PI 1E-05 5E-05 3,5 0,99 1E-08 3E-01 1E-06 1E-08 5E-01 4,6E-04 1E-08 1,5E-04

BO 1E-05 1E-04 3,7 1,00 1E-08 1E-01 1E-06 1E-08 9E-02 4E-04 1E-08 3E-04

linea

altre patologie

II.

NHL: what are they?MRD: what is?

Which techniques? Which techniques? The FIL MRD NET experienceClinical aspects and role of MRD

HOT TOPIC

Adapted from Ferrero, Hematol Oncol 2011

MRD & LETTERATURAyearyear markermarker methodmethod ptspts statusstatus txtx PFSPFS

1991 Bcl2 Q-PCR 114 Rel ASCT <0.05

1995 Bcl2, IgH Q-PCR 24 Rel ASCT <0.05

1999 Bcl2 Q-PCR 113 Rel ASCT <0.05

2000 Bcl2 Q-PCR 53 Rel ASCT Ns

2002 Bcl2, IgH Q-PCR 42 Dx ASCT <0.05

2004 bcl2., IgH Q-PCR 35 Dx ASCT <0.05

2006 Bcl2, IgH Q-PCR 42 Rel ASCT <0.05

2008 Bcl2, IgH QT-PCR 60 Dx HD/R-CHOP <0.05

2009 Bcl2 QT-PCR 127 Dx zevalin <0.05

CHOPCHOP--RR

PB

PCR+ predicts the FFS also in multivariate analysis

Rambaldi, Blood 2002

RR--CHOP / RCHOP / R--BB

179 pts receiving R -CHOP vs R-Bendamustine

@ dx: PCR + 63% - QUANTITATIVE PCR

2y-EFS= 82% pts low PCR vs 47% high PCR

Post -tx: PCR - = 86%Post -tx: PCR - = 86%median 2y- PFS= 9 m pts PCR + vs n.r. PCR -

Outcome is significantly conditioned Outcome is significantly conditioned

by the PCR status @ diagnosis and postby the PCR status @ diagnosis and post--therapytherapyZohren, ASH 2009

RR--CHOP CHOP EORTC 20981

238 pts with relpased follicular lymphoma randomized to CHOP vs R-CHOP

Pts in PR/CR randomized to Rituximab maintenance vs observation

Van Oers, JCO 2010

RR--CHOP CHOP EORTC 20981

PFS is conditioned by the PCR status @ diagnosis...

Van Oers, JCO 2010

RR--CHOP CHOP EORTC 20981

...but not conditioned by the PCR status at the end of therapy...

Van Oers, JCO 2010

CR/CRuoppure

ZEVALIN® (n=208)

Rituximab 250 mg/m2 I.V.

Random

izzazione

Inizio dello studio

Terapia di prima linea con CVP,

CHOP/CHOP-like,

ConsolidamentoInduzione

RADIORADIO--IMMUNO TXIMMUNO TX

STUDIO FIT (409 pts) )

oppure PR

NRPD

Esclusione

Rituximab 250 mg/m2 I.V.nei giorni –7 e 0

+ ZEVALIN® (max 1184 MBq) nel giorno 0

Nessun ulteriore trattamento

(n=20)6)

Random

izzazione

CHOP/CHOP-like, combinazioni

con fludarabina, clorambucile o rituximab

Controllo

ZEVALIN®, n/N* (%)Controllo, n/N* (%)

PCR qualitativa

RADIORADIO--IMMUNO TXIMMUNO TX

61/68 (90%)21/59 (36%)PB

90% converted from PCR+ to PCR –after ZEVALIN ®

RADIORADIO--IMMUNO TX RQIMMUNO TX RQ--PCRPCR

Goff, JCO 2009

@randomization 54% PCR + zevalin 46% PCR+ obs2% HIGH10% INTERMEDIATE88% LOW

+ 3m: red >1 log76% zevalin vs 25%+6m: PCR –84% vs 30%

ZEVALINZEVALIN

Only in PCR+ pts zevalin prolongs PFS

Goff, JCO 2009

AutoSCT & MRD postAutoSCT & MRD post

70 pts affected by indolent NHL

receiving autologours transplantation

Post-autoTMO PCR - : 70% foll

25% SLL

Corradini, JCO 2004

25% SLL

25% MCL

EFS is significantly conditioned

by the PCR status after transplantation

AutoSCT & AFERESIAutoSCT & AFERESI

The contamination molecularly assessed

of aphereses does condition EFS

Corradini, JCO 2004

APHERESES PCR - : 54% foll

25% SLL

12% MCL

OS PFS

aph high

AutoSCT & AFERESIAutoSCT & AFERESI

aph high

aph low

aph intermediate

deathhigh

MCL & ASCTMCL & ASCT160 pts

Pott, Blood 2010

MCL & ASCTMCL & ASCT

PCR status after transplantation does impact

on durability of response

MRD-

Pott, Blood 2010

MRD+

MCL & FCRMCL & FCR

99 pts

Pott, Blood 2010

MCL & FCRMCL & FCR

MRD-

PCR status during rituximab maintenance

does impact on durability of response

Pott, Blood 2010

MRD+

CLL & FCRCLL & FCR

493 pts randomized to FC vs FCR

MRD assessed by 4 -color FC

MRD - = 0.01%

FCR > FCFCR > FC

Bottcher, Blood 2012

CLL & FCRCLL & FCR

2. MRD does impact on OS

Bottcher, Blood 2012

CLL & FCRCLL & FCR

...and PFS for both arms...

Bottcher, Blood 2012

CLL & ASCTCLL & ASCT

40 pts receiving ASCT

FC (100% +) ASO-PCR (97% +)

MRD after 3 and 6 m impacts on PFS and OS

Moreno, Blood 2006

MRD+

MRD+

MRD- MRD-

CLL & ALLOSCTCLL & ALLOSCTThe relapse rate after allogeneic transplantation is conditioned by MRD status

Bottcher, Blood Revies 2011

CLL & TMOCLL & TMO

CLLX3 trial – 90 pts

an MRD-driven approach reduces relapse rate

Dreger, Blood 2010

CONCLUSIONSCONCLUSIONS

• In 2013 the MRD still represents a real “hot topic”

• Perhaps more conclusive data will be produced by the new trials adopting produced by the new trials adopting quantitative PCR

• THE NEW FIL TRIALS ARE MOLECULARLY ORIENTED!!!

THE NEW FOLL12

THANK YOU THANK YOU VERY MUCH TO VERY MUCH TO

MY LABMY LABMY LABMY LABMY COLLEAGUESMY COLLEAGUES

...AND ALL OF YOU......AND ALL OF YOU...