From genetics to the clinic of CLLplan.medone.co.kr/70_icksh2019/data/ES02-3_Wei_Xu.pdf ·...

From genetics to the clinic of CLL

Wei XU (徐卫)Department of Hematology

First Affiliated Hospital of Nanjing Medical UniversityJiangsu Province Hospital

2

I have no personal or financial interests to declare:

I have no financial support from an industry source at the current presentation.

대한혈액학회 Korean Society of Hematology

COI disclosureName of author ：Wei Xu

3

• CLL is the most common leukemia in Western countries, but is less frequent in Asian countries.

• It has geographical variability and familial risk (7 to 8 fold).

• Diagnosis of CLL is based on unique immunophenotype of clonal B lymphocytes.

• Progression of CLL is associated with genetic complexity.

4

5

The role of genetic lesions in the

prognosis of CLL

Subclone and CLL clonal evolution

6Fabbri G, et al. Nat Rev Cancer. 2016;16:145

• CLL may originate at the stem cell stage, and subsequent antigenic stimulationmay lead to selection and expansion of mature B cells.

• IGHV-M CLL seem to originate from post-GC CD5+CD27+ B cells and IGHV-UMCLL seem to arise from pre-GC CD5+CD27- B cells.

7Gruber M, et al. Semin Hematol. 2014;51:177.

• The introduction of NGS has led to a increase in the knowledge of the molecular underpinnings of CLL over the last ten years.

8

• CLL-associated genetic lesions with a frequency of ≥1.5%. • The frequency of genetic alterations in unselected cases and in the two IGHV-M and

IGHV-UM CLL subgroups is reported in grey, orange and purple, respectively. • Grouped according to the biological programme or pathway in which they are involved.

Fabbri G, et al. Nat Rev Cancer. 2016;16:145

9Lazarian G, et al. J Clin Oncol. 2017;35:984.

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Pathways Involved genesNOTCH signaling NOTCH1, FBXW7

NF-κB pathwayMYD88, RIPK1, DDX3X, TRAF3, TRAF2, SAMHD1,BIRC3, BRAF, EGR2, IRF4, BCOR, BTK, NFKBIE,

TNFAIP3, CD79B, ITPKB, CARD11, PLCG2Wnt signaling MED12, FUBP1, MGA, MYC

DNA repair and cell cycle regulation POT1, ATM, CHEK2, DYRK1A, BRCC3, TP53

EpigeneticsHIST1H1B, HIST1H1E, DDX3X, ZMYM3, CHD2, EZH2,

KMT2C, KMT2D, BAZ2A, TET2, CREBBP, EP300,ASXL1, IKZF3

RNA and ribosomal processing SF3B1, EWSR1, NXF1, XPO4, RPS15

Others FAT1, FAT3, IGLL5, MTOR, CACNA1E, BCL6, PRDM1, BCL2, PIM1, KIT, KLHL6, CXCR4, PIK3CA, SOCS1

Xu Wei, et al. Data unpublished

• We performed targeted gene sequencing (TGS) of 93 Chinese CLL patients. • This TGS panel consists of 63 genes

11

The most frequently mutated genes: • IGLL5 (n=21)• MYD88 (n=15)• LRP1B (n=11)• KMT2D (n=7)• CREBBP (n=6)• TET2 (n=6)• CHEK2 (n=5)• SF3B1 (n=4)

• 195 driver mutations were found across 50 genes • 80 patients (86%) harbored at least one mutated gene• 57 cases (61.3%) harbored mutations involving at least

two genes

Xu Wei, et al. Data unpublished

12

Gene Our cohort Landau et al Puente et al P value (vs Landau et al)

P value (vs Puente et al)

IGLL5 21/93 (22.6%) 12/538 (2.2%) 0/452 (0.0%) <0.0001 <0.0001

MYD88 15/93 (16.1%) 16/538 (3.0%) 18/452 (4.0%) <0.0001 <0.0001

LRP1B 11/93 (11.8%) 16/538 (3.0%) 0/452 (0.0%) 0.0007 <0.0001

KMT2D 7/93 (7.5%) 4/538 (0.7%) 5/452 (1.1%) 0.0002 <0.0001

TET2 6 /93 (6.5%) 3/538 (0.6%) 0/452 (0.0%) 0.0005 <0.0001

CREBBP 6 /93 (6.4%) 3/538 (5.8%) 3/452 (0.7%) 0.0005 <0.0001

CHEK2 5/93 (5.4%) 5/538 (0.9%) 0/452 (0.0%) 0.0086 <0.0001

BCL2 3/93 (3.2%) 0/538 (0.0%) 0/452 (0.0%) 0.0193 <0.0001

PLGC2 2/93 (2.1%) 0 (0%) 0/452 (0.0%) <0.0001 <0.0001

SF3B1 4/93 (4.3%) 113/538 (21.0%) 39/452 (8.6%) 0.0309 0.2055

RPS15 0/93 (0%) 23/538 (4.3%) 0/452 (0.0%) 0.0031 <0.0001

Xu Wei, et al. Data unpublished

13

Genetic lesions in CLL

Subclone and CLL clonal evolution

14Rossi D, et al. Br J Cancer. 2016;114:849.

• The genes that are recurrently mutated in each cellular programme and theclinical implications of gene mutations.

15Fabbri G, et al. Nat Rev Cancer. 2016;16:145

• CLL can be classified into two subgroups on the basis of the presence orabsence of mutations in IGHV.

16

65.8%

34.2%

48.9% 51.1%

M-IGHV UM-IGHV

Chinese Italian（N= 631） （N= 792）

P<0.001

• Chinese patients with CLL are more likely to carry mutated IGHV gene than Italian CLL patients.

Marinelli M, Ilari C, Xia Y et al. Oncotarget. 2016 ;7:20520.

17

15.5%

3.3%

46.8%

28.7%

2.4% 1.6% 1.7%

25.1%

3.3%

46.0%

20.6%

3.2% 1.1% 0.8%

VH1 VH2 VH3 VH4 VH5 VH6 VH7

Chinese Italian

P<0.001

• VH3 is the most frequently used subgroup in both Chinese and Italian patients with CLL.

• VH1 usage was significantly higher in Italian patients.• VH4 usage was more common in Chinese patients.

Marinelli M, Ilari C, Xia Y et al. Oncotarget. 2016 ;7:20520.

P<0.001

18

Tobin G, et al. Blood.2003;101:4952. Ghiotto F, et al. J ClinInvest. 2004;113:1008. Stamatopoulos B, et al. Blood.2007;53:1757. Chu CC, et al. Blood. 2008;112:5122. Catera R, et al. Mol Med. 2008;14:665. Rossi D, et al. Clin Cancer Res. 2009;15:4415. Agathangelidis A, et al. Blood. 2012;119:4467. Strefford JC, et al. Leukemia.2013;27:2196. Rossi D, et al. Blood.2013;121:4902. Papakonstantinou N, et al. MolMed. 2013;19:115. Mansouri L, et al. J Exp Med. 2015;212:833.

1/3 of patients with CLL exhibit highly similar ‘stereotype’ heavy-chaincomplementarity-determining region 3 (HCDR3) sequences, suggesting a commonantigenic determinant for the development of CLL.

19

27.3%

21.2%

16.7%

6.1% 6.1%4.5%

3.0%1.5%

4.3%

20.9%

11.3%

7.0%4.3%

1.7%

6.1%

10.4%

#8 #1 #4 #5 #6 #77 #3 #2

Chinese Italian

• The proportion of known stereotyped BCR was significantly lower in Chinese CLL patients (19.7%) than that in Italian CLL patients (25.8%).

• The incidence of subset #8 was remarkably higher in Chinese CLL cases. • The frequency of subset #2 was significantly lower in Chinese CLL cases.

P<0.001

P=0.03

Marinelli M, Ilari C, Xia Y et al. Oncotarget. 2016 ;7:20520.

20

• Subset #1: significantly worse than other CLL patients with IGHV unmutated– median OS： 28.0 months vs 85.1 for unmutated IGHV vs 165.1

for mutated IGHV, P<0.0001) – Median TTT： 0.8 months vs 4.8 for unmutated IGHV vs 46.2 for

mutated IGHV, P<0.0001)

• Subset #8: similar to other CLL patients with IGHV unmuated CLL– median OS： 82.2 months – median TTT：1.7 months– had a significantly increased risk of developing Richter’s

syndrome (n=3/16) (P= 0.0095), which was consistent with a previous study from western populations.

Marinelli M, Ilari C, Xia Y et al. Oncotarget. 2016 ;7:20520.

21

NJ_23 #NEW1 IGVH3-23 IGHJ4 IGHD2-21 mutated 20 CAKGYRDNYDGDQSSVFDSWNJ_186 #NEW1 IGVH3-23 IGHJ4 IGHD3-22 mutated 20 CAKGYRDNYDGDQSSVFDSWTJ_38 #NEW10 IGVH1-18 IGHJ4 IGHD6-6 unmutated 15 CARLQYIPMYSLDYWTJ_146 #NEW10 IGVH1-18 IGHJ4 IGHD6-13 unmutated 15 CARLQYIPMYSLDYWTJ_153 #NEW11 IGVH3-23 IGHJ4 IGHD2-2 mutated 10 CVTGGQAGDWTJ_156 #NEW11 IGVH3-23 IGHJ4 IGHD2-2 mutated 10 CVTGGQAGDWTJ_134 #NEW12 IGVH4-34 IGHJ3 IGHD6-19 mutated 17 CARPYEVAVAPGAFDVWTJ_136 #NEW12 IGVH4-34 IGHJ3 IGHD6-19 mutated 17 CARPYEVAVAPGAFDVWTJ_123 #NEW1 IGVH4-4 IGHJ3 IGHD5-12 mutated 20 CARGASGYGLLGADDGFDVWTJ_124 #NEW13 IGVH4-4 IGHJ3 IGHD5-12 mutated 20 CARGASGYGLLGADDGFDVWTJ_91 #NEW14 IGVH3-30 IGHJ6 IGHD5-12 mutated 13 CARRGHDSYMDLWTJ_150 #NEW14 IGVH3-30 IGHJ6 IGHD5-12 mutated 13 CARRGHDSYMDVWTJ_151 #NEW14 IGVH3-30 IGHJ6 IGHD5-12 mutated 13 CARRGHDSYMDVWTJ_56 #NEW15 IGVH3-74 IGHJ4 IGHD1-26 mutated 9 CSWDHFDSWTJ_143 #NEW15 IGVH3-74 IGHJ4 IGHD5-12 mutated 9 CTYDHFDSWNJ_78 #NEW2 IGVH4-59 IGHJ4 IGHD3-22 unmutated 22 CARGAAGYYDSSGYKEYYFDYWNJ_168 #NEW2 IGVH4-59 IGHJ4 IGHD3-22 unmutated 22 CARRGSGYYDSSGYPEYYLDYWNJ_254 #NEW3 IGVH3-21 IGHJ4 IGHD3-22 unmutated 21 CARESYYYDISGYFLYYFDYWNJ_266 #NEW3 IGVH3-21 IGHJ4 IGHD3-22 unmutated 21 CARESYYYDISGYFLYYFDYWNJ_136 #NEW4 IGVH3-74 IGHJ3 IGHD6-19 unmutated 14 CARAVAGIDAFDIWNJ_42 #NEW4 IGVH3-74 IGHJ3 IGHD6-19 unmutated 14 CARAVAGIDAFDIWHK_85 #NEW5 IGVH4-4 IGHJ4 IGHD3-22 unmutated 18 CARGGGYSSGYPYYFDYWNJ_300 #NEW5 IGVH4-4 IGHJ4 IGHD6-19 unmutated 18 CARGGGYSSGLPYYFDYWHK_19 #NEW6 IGVH3-7 IGHJ6 IGHD3-10 unmutated 20 CAAGRGSGRYYYYYYGMDVWNJ_71 #NEW6 IGVH3-7 IGHJ6 IGHD1-26 unmutated 20 CAGGSGSYQYYYYYYGMDVWTJ_03 #NEW7 IGVH3-9 IGHJ6 IGHD6-13 mutated 24 CAKDRVCCIAAAGPSLRYYGMDVWTJ_155 #NEW7 IGVH3-9 IGHJ6 IGHD6-13 mutated 24 CAKDRVCCIAAAGPSLRYYGMDVWTJ_27 #NEW8 IGVH3-23 IGHJ4 IGHD3-16 mutated 29 CAKDVRGGREEFADYIWVSYRSGIAFDYWTJ_45 #NEW8 IGVH3-23 IGHJ4 IGHD3-16 mutated 29 CAKDVRGGREEFADYIWVSYRSGIAFDYWTJ_32 #NEW9 IGVH4-34 IGHJ5 IGHD2-21 mutated 21 CARRRDPVYCGDDCPTGVDSWTJ_34 #NEW9 IGVH4-34 IGHJ5 IGHD2-21 mutated 21 CARRRDPVYCGDDCPTGVDSWNJ_15 #NOVEL 1 IGVH4-59 IGHJ6 IGHD3-22 unmutated 25 CARGNYYDSSGYYYVGYYYYYMDVWNJ_31 #NOVEL 1 IGVH4-59 IGHJ6 IGHD3-22 unmutated 25 CARGDYYDSSGYYYVGYYYYYMDVW

Xu Wei, et al. Data unpublished

16 new paired clusters were identified among Chinese CLL cases and these new clusters were characterized by the remarkable recurrence of the VH3-23 gene

22Chai-Adisaksopha C, et al. Blood. 2017; 130: 2278.

• A systematic review of 5 randomized trials indicate that FCR can only improve CR, PFS and OS in patients with IGHV-M.

• With a follow-up from 31 to 71 months, median PFS was not reached in the patients with IGHV-M.

23

The CLL3 trial was designed to study intensive treatment including ASCT as part of first-line therapy in patients with CLL.

• PFS and OS were significantly reduced in the patients presence of IGHV-UM

IGHV-UMIGHV-UM

Dreger P, et al. Blood. 2012;119:4851

24

• A similar PFS and OS in patients with unmutated and mutated IGHV.

O'Brien S, et al. Blood. 2018;131:1910

25Döhner H, et al. N Engl J Med. 2000;343:1910.

• 17p- was associated with very poor prognosis and the patients had only 32 months of survival time.

• The patients with isolated 13q-, demonstrates better prognosis, had 133 months of survival time.

• More than 80% of CLL patients were detected with cytogenetic aberrations by interphase FISH examination.

26

• TP53 aberration is the strongest indicator for inferior prognosis of CLL.• The cumulative incidence of TP53 aberration is 10% at the time of first line treatment, and

20–40% in relapsed CLL. • TP53 mutations are often accompanied by 17p13 deletion (60%).

Rossi D, et al. Leuk Lymphoma. 2017;58:1548.

27

0.0

0.2

0.4

0.6

0.8

1

0 6 12 18 24 30 36 42 48 54 60 66

FCFCR

0.0

0.2

0.4

0.6

0.8

1

0 6 12 18 24 30 36 42 48 54 60 66

del(17p)

del(11q)

+12del(13q)

no aberrationdel(11q)

+12

del(13q)

no aberration

del(17p)

Time (months)Time (months)

Ove

rall

surv

ival

Ove

rall

surv

ival

3-year follow-up

Hallek M, et al. Lancet . 2010; 376:1164

• Patients with del(17p) showed a significantly shorter OS than did those in all the other cytogenetic subgroups, irrespective of the treatment given.

OS in del(17p) patients: FCR is not good

28Fischer K, et al. J Clin Oncol. 2012;30:3209

del(17p)

• Patients with del(17p) had a significantly shorter EFS than did those in all the other cytogenetic subgroups

29

• PFS and OS were significantly reduced in the patients presence of 17p-

del(17p) del(17p)

Dreger P, et al. Blood. 2012;119:4851

30Dreger P, et al. Blood. 2010;116:2438.

• There were no significant differences in terms of EFS and OS among the individual genetic subgroups.

• In particular, the 13 patients with 17p- had similar survivals with the other patients with other cytogenetic aberrations.

del(17p)del(17p)

31

chlorambucil vs F vs FC

• PFS and OS were significantly shorter in TP53mut patients than in patients with a WT• Poorer outcome for those with isolated TP53mut compared with outcome for those with

neither abnormality did not reach statistical significance for PFS and OS

Gonzalez D, et al. J Clin Oncol. 2011; 29:2223.

32

• TP53mut was associated with significantly decreased PFS and OS in both treatment arms • Both 17p- and TP53mut are independent prognostic factors for PFS and OS

Stilgenbauer S, et al. Blood. 2014; 123:3247.

33

• The mutational status of the TP53 genes had no significant effect on OS and EFS. • Allo-SCT can provide long-term disease control in patients with TP53mut.

Schnaiter A, et al. Blood. 2013; 122:1266.Dreger et P, al. Blood. 2013;121:3284.

34Rossi D, et al. Leuk Lymphoma. 2017;58:1548

Cross-trial comparisons of the response rate and PFS at 12 months between CIT and novel agents in relapsed CLL harboring TP53 lesions.

• The activity of novel agents appears significantly better than that observed in CIT trial.

35Fabbri G, et al. Nat Rev Cancer. 2016;16:145

• PEST domain is involved in the ubiquitination of NOTCH1 and may affect the stability of NOCTH1 protein.

• Mutations within the PEST domain of NOTCH1 have been found in 8% to 12% of patients at diagnosis, in 21% of patients with refractory disease and in 30% of cases with Richter syndrome.

• NOTCH1 mutations have been associated with a shorter TTT and shorter OS independent of other prognostic factors.

36

• The presence of a NOTCH1 mutation was associated with a significant reduction in OS and PFS.

NOTCH1 54.8m vs 74.6m NOTCH1

22.0m vs 26.4月

Oscier DG, et al. Blood. 2013; 121:468.

chlorambucil vs F vs FC

37Stilgenbauer S, et al. Blood.2014;123:3247

• NOTCH1 mutations was also associated with significantly decreased PFS and OS in both treatment arms.

38Zou Y, et al. Cancer Med. 2018;7:1689.

• We detected NOTCH1 mutation in 317 Chinese patients with CLL, mutation rate was 9.1%.

• Subjects with NOTCH1 mutations had briefer TTT and OS than subjects without this abnormality.

39Fabbri G, et al. Nat Rev Cancer. 2016;16:145

• SF3B1 is a splicing factor, which is involved in normal hematopoiesis• Mutations were found in 5% to 17% of CLL patients and were associated with

advanced stage, 11q-, and with short TTT and OS independent of other prognostic factors

40

SF3B1 vs WT：P=0.08P<0.001

Oscier DG, et al. Blood. 2013; 121:468.

• SF3B1 mutations were significantly associated with a reduced OS.• However, wild-type and mutant alleles had a similar PFS.

chlorambucil vs F vs FC

41

• SF3B1 mutation was associated with significantly decreased PFS in both treatment arms and with slightly (not significantly) inferior OS

Stilgenbauer S, et al. Blood. 2014; 123:3247.

42

• The mutational status of the SF3B1 and NOTCH1 genes had no significant effect on OS and EFS.

• Allo-SCT can provide long-term disease control in patients with SF3B1 or NOTCH1 mutations.

Dreger et P, al. Blood. 2013;121:3284.

43

Mutation N=23 (7.8%)

L265P 15 (63.2%)

M232T 3

S219C 3

S243N 1

V271F 1

Chinese Western

At diagnosis 9.5% (15/158)

2-5%Progressive 8.3% (7/84)

Relapsed 4% (1/25) Not reported

Refractory 0%（0/28）

L265P

MYD88 is a critical adaptor molecule of the TLR complexMYD88 mutation rate: 23/295 (7.8%)

• Higher incidence of MYD88 mutations was seen in Chinese cohort compared with westerners.

• The most common mutation site was L265P.

Qin SC, et al. Blood Cancer J. 2017;7:651

44

• No significantly different between mutations and wide type on TTT and OS• In IGHV mutated cases, MYD88 mutated patients with had significantly shorter

TTT, similar to the patients with IGHV unmutated.

Qin SC, et al. Blood Cancer J. 2017;7:651

45Rossi D, et al. Blood. 2013;121:1403.

100

80

60

40

20

0

Cum

ulat

ive

Prob

abili

ty o

f OS

(%)

0 5 10 15

P < 0.0001

Yrs From Diagnosis

Del(13q14)Normal/+12NOTCH1 M/SF3B1 M/del(11q22-q23)TP53 DIS/BIRC3 DIS

Events, n27534157

Total, n15522899101

Median OS, yrs

NR13.48.55.0

95% CI--

12.1-14.75.6-11.53.4-6.5

Del(13q14) vs normal/+12

Normal/+12 vs NOTCH1 M/SF3B1 M/del(11q22-q23)

NOTCH1 M/SF3B1 M/del(11q22-q23)vs TP53 DIS/BIRC3 DIS

P = 0.0406

P = 0.0082

P = 0.0196

29%

37%

57%

69%

46

Genetic lesions in CLL

The role of genetic lesions in the

prognosis of CLL

47

• Mutations were considered subclonal when VAF was <12% and clonal when VAF ≥12%.

Nadeu F, et al. Blood. 2016;127:2122.

48

diagnosis diagnosis first treatment

• OS of cases harboring solely subclonal TP53 mutations was significantly shorter than that of cases with an unmutated TP53 gene and was similar to that of cases harboring clonal TP53 mutations.

Rossi D, et al. Blood. 2014; 123:2139.

49

• Both clonal and subclonal NOTCH1 mutations predicted for a shorter TTT compared with patients having a WT sequence.

• Patients harboring clonal, but not subclonal, NOTCH1 mutations had a significantly shorter OS compared with patients having a WT sequence.

Nadeu F, et al. Blood. 2016;127:2122.

50Lazarian G, et al. J Clin Oncol. 2017;35:984.

• First, acquisition of somatic alterations in CLL leads to its genomic diversification. • Exposure to chemoimmunotherapy as first-line treatment commonly selects TP53-

disrupted subclones. • Each therapy proposed at relapse may differentially shape the CLL architecture and

would select preferential alterations.

51Puente XS, et al. Nat Genet. 2013;45:229.

• Transformation of normal B cells into CLL results from the accumulation of early driver somatic mutations, such as 13q-, +12 and mutations in MYD88 or NOTCH1.

• During CLL progression, subclones harboring mutations such as TP53, ATM, SF3B1 or NRAS, expand in response to intrinsic or extrinsic pressures.

• Most late driver mutations can be detected early during CLL evolution.

52Sutton LA, et al. Semin Cancer Biol. 2015;34:22.

53Woyach JA, et al. N Engl J Med.2014;370:2286.

• Six CLL patients with acquired resistance to ibrutinib. • Whole-exome sequencing was performed. • C481S mutation in BTK at the binding site of ibrutinib were identified in five

patients and three distinct mutations in PLCγ2 were identified in two patients.

54

• For 20 patients with a detectable mutation in BTK or PLCγ2 at time of relapse, an initial clone could be detected at a median of 9.3 months before relapse.

• Mutations in BTK appear early and have the potential to be used as a biomarker for future relapse, suggesting an opportunity for intervention.

Woyach JA, et al. J Clin Oncol. 2017;35:1437.

55

• CLL is a molecularly heterogeneous disease as revealed by recent genomic studies.

• Among genetic lesions that are recurrent in CLL, few clinically validated prognostic markers, such as TP53 lesions, are available for the use in clinical practice to guide treatment decisions.

• Recently, several novel mutations have been identified in CLL. These mutations have shown to be promising to refine the prognostic stratification of patients.

56

• Small subclones harbouring drug resistant mutations anticipate the development of a chemorefractoryphenotype.

• Clonal evolution in longitudinal samples was observed in CLL patients.

• The characterization of the subclonal mutation and its dynamics in the evolution of the disease may be relevant for the management of CLL patients.

57

Thanks for your attention！