Ph-like (BCR/ABL1-like): un apporoccio baato sul target molecolare

Transcription

1 Ph-like (BCR/ABL1-like): un apporoccio baato sul target molecolare Sabina Chiaretti, MD PhD LE SFIDE DELLA MEDICINA DI PRECISIONE IN EMATOLOGIA Bologna 28 Giugno, 2018

2 Topics: BCR/ABL1-like and other subgroups Molecular background Incidence Diagnosis Outcome and MRD Treatment

3 Topics: BCR/ABL1-like Molecular background Incidence Diagnosis Outcome Treatment

4 First report in adult ALLs Chiaretti et al, CCR 2005 Haferlach et al, Blood : first identification, by GEP, of a subset of adult B-lineage ALL clustering together with BCR/ABL1+ ALL cases

5 Characterization of BCR-ABL1-like in children Deletions are visualised in red, whereas amplifications are shown in blue. GEP: Identification, within children (n=297), of a subset with a transcriptional profile resembling that of BCR/ABL1+ cases ( 15-20%) Clinical features: Hyperleukocytosis, poor response to VCR, ASP and DNM, poor prognosis (reduced DFS at 5 years and increased resistance to induction) Array-CGH: IKZF1, PAX5, TCF3 and VPREB1 deletions, CRLF2 deregulation Den Boer et al, Lancet 2009; Mullighan et al, NEJM 2009

6 Survival probability Impact of CRLF2 expression on B-ALL survival CRLF2 deregulation 100 CRLF2-low DCt>8 CRLF2-high DCt<8 del(x)(p22.33p22.33) del(y)(p11.32;p11.32) P2RY8/CRLF2 75 CRLF2-low t(x;14)(p22;q32) t(y;14)(p11;q32) d-crlf2 50 Point mutations (F232C) 25 CRLF2-high 5-7% of pediatric BCR/ABL1-negative cases >50% of cases associated with Down's syndrome 5-15% of adult BCR/ABL1-negative cases Associated with mutant JAK and IKZF1 p= Months Associated with the BCR/ABL1-like profile, but not useful as a diagnostic marker Chiaretti et al, Leuk Res 2016

7 In high risk ALL, RNA-seq has identified novel mutations that involve TKs in the majority of cases. They appear to have transforming capability and to respond to TKIs. Roberts KG. Cancer Cell 2012; 22: Roberts KG, et al. N Engl J Med 2014;371:

8 BCR/ABL1-like ALL in adults. Genetics Higher frequency of other kinase involvement. Some cases do no have any lesions. Roberts KG et al, JCO 2016

9 Kinase fusions identified in Ph-like ALL Kinase gene Fusion partners, n Patients, n 5 genes ABL ETV6, NUP214, RCSD1, RANBP2, SNX2, ZMIZ1 ABL2 3 7 PAG1,* RCSD1,* ZC3HAV1* CSF1R 1 4 SSBP2* PDGFRB 4 11 EBF1, SSBP2,* TNIP1,* ZEB2* CRLF IGH, P2RY8 JAK ATF7IP,* BCR, EBF1,* ETV6, PAX5, PPFIBP1,* SSBP2, STRN3, TERF2,* TPR* EPOR 2 9 IGH, IGK* DGKH 1 1 ZFAND3* IL2RB 1 1 MYH9* NTRK3 1 1 ETV6 PTK2B 2 1 KDM6A,* STAG2* TSLP 1 1 IQGAP2* TYK2 1 1 MYB* Roberts KG, et al. N Engl J Med 2014;371:

10 Topics: BCR/ABL1-like Molecular background Incidence Diagnosis Outcome Treatment

11 BCR-ABL1-like. Incidence Incidence is higher in AYA (10% in children; 27% in AYA). NEVER detected in cases positive for known fusion transcripts (BCR/ABL1, KMT2A-r, TCF3/PBX1) However: - It highly depends on the denominator (all B-lineage ALL or B-neg ALL) and - on the assay used for BCR/ABL1-like identification - More adult cases are being evaluated incidence in adults is almost equal to AYA 25% Roberts KG, NEJM : ; Heroldt T, NEJM :2235; Chiaretti S et al, in press

12 Topics: BCR/ABL1-like Molecular background Incidence Diagnosis Outcome Treatment

13 BCR/ABL1-like ALL diagnosis: a difficult issue Difficult to identify these cases by techniques other Well identified subgroup by GEP than GEP Furthermore, non univocal gene signature Poor prognosis documented: therapy?

14 Still a difficult issue LDA Q-RT-PCR Quantification of expression of 15 transcripts (Kang BW et al, ASH 2013) Quantification of expression of 10 transcripts (Chiaretti S et al, BJH 2018) LDA, FISH, RT-PCR, NGS (RNA-seq, WGS, WES) (Roberts KG et al, NEJM 2014) Integrated algorithms FISH, RT-PCR, Q-RT-PCR, NGS (Fasan A et al, ASH 2015) Known fusion transcripts, JAK2 mutations, CRLF2-r (Herold T et al, Haematologica 2016) CRLF2 expression (FC), JAK2 mutations, FISH for TKrearrangements and CRLF2-r (Jain N et al, ASH 2017) As far as possible, diagnostic assays should be available in most centers (or in centralized laboratories)

15 BCR/ABL1-like ALL diagnosis: a difficult issue Well identified subgroup by GEP Difficult to identify these cases by techniques other than GEP Furthermore, non univocal gene signature To set up a diagnostic assay for a straightforward identification of BCR/ABL1-like ALL cases Poor prognosis documented: therapy? To analyze the clinico-biologic and molecular features of BCR/ABL1-like adult ALL cases To verify BCR/ABL1-like cases response to TKIs (ponatinib)

16 BCR/ABL1-like ALL cases: methods (I) 342 Probsets CRLF2 Harvey R.C. et al Probsets By meta-analysis of published and in house GEP data of B-NEG ALL, selection of 9 BCR/ABL1-like specific transcripts (FC 1.5, p<0.001) + CRLF2 Confirm the differential expression of the 10 transcripts by a Q-PCR approach Build a BCR/ABL1-like predictor on the basis of Q-PCR results (easy, fast and economic!)

17 BCR/ABL1-like ALL cases: methods (II) Discovery panel 52 adult ALL samples previously evaluated by GEP 26 BCR/ABL1- like ALL cases 26 negative B-ALL Q-RT expression values shrinked into 3 principal components. A logistic regression model was used to examine the association among the 3 principal components and BCR/ABL1-like cases. Generation of a score on principal components, used to classify the remaining samples (screening panel).

18 Development of the BCR/ABL1-like predictor and screening Selection of BCR/ABL1-like specific genes and validation by Q-PCR 1. Selection of 10 predictor genes 2. Validation by Q-PCR in 52 B-NEG ALL Development of BCR/ABL1-like predictor* 1. Identification of principal components (PCs) Factor loadings PC1 PC2 PC3 Gene1 2^(-ΔCt) Gene2 2^(-ΔCt) Gene3 2^(-ΔCt) Gene4 2^(-ΔCt) Gene5 2^(-ΔCt) Gene6 2^(-ΔCt) Gene7 2^(-ΔCt) Gene8 2^(-ΔCt) Gene9 2^(-ΔCt) Gene10 2^(-ΔCt) Definition of a score Screening 1. Q-PCR of predictor genes in 129 B- NEG ALL 2. BCR/ABL1-like predictor 54/194 newly identified BCR/ABL1-like patients (28%) - 9.5% children, 29% AYA, 30% adults - * Chiaretti S et al, BJH in press

19 BCR/ABL1-like ALL cases: clinical features BCR/ABL1-like (n=54) non-bcr/abl1-like (n=140) P-value Gender (M/F) 36/18 78/62 p=ns Age 32 (6-72) 28 (0-78) p=p=ns Wbc x 10 9 /l 22.6 ( ) 12.4 ( ) p=0.023 Plts x 10 9 /l 47 ( ) 47 (1-308) p=ns Hb g/dl 9.7 ( ) 8.9 ( ) p=ns CR rate 77.8% 89.2% p=0.06 No significant differences in age and gender: significantly higher WBC and lower remission rate

20 Molecular features of the cases identified by the BCR/ABL1-like predictor BCR/ABL1-like (N=28) non-bcr/abl1-like (N=26) 96% of cases had a lesion typical of the BCR/ABL1-like subset JAK/STAT mutations often concurrent with CRLF2 overexpression CRLF2 overexpression detected in 69% of BCR/ABL1-like cases, though not exclusively ABL-class rearrangements detected in 18% of BCR/ABL1-like ALL TK-r cases often do not express high levels of CRLF2 Chiaretti S et al, BJH 2018

21 Topics: BCR/ABL1-like Molecular background Incidence Diagnosis Outcome and MRD Treatment

22 Outcome in children Associated with male gender, hyperleucocytosis and increased MRD levels, also when patients are considered as standard-risk Intensive and MRD-driven treatment seems to abolish the negative prognostic impact of the BCR/ABL1-like signature in childhood ALL Roberts KG et al, JCO 2014

23 Response to induction therapy in adults Contrasting results on the CR rate - Lower than in other B-neg ALL cases (den Boer J et al, Haematologica 2015; Chiaretti S et al, BJH in press) -No differences with other ALL cases (Herold T et al, Haematologica 2016; Jain N et al, Blood 2017) -MRD persistence more frequent in BCR/ABL1-like cases (Roberts KG et al, JCO 2016; Herold T et al Haematologica 2017; Jain N et al Blood 2017; personal data). Not yet clear if in adults MRD negativity overcomes BCR/ABL1-like signature

24 Survival in adults Significantly inferior survival (EFS, DFS, OS) in all reported studies Roberts KG et al, JCO 2016

25 Topics: BCR/ABL1-like Molecular background Incidence Diagnosis Outcome and MRD Treatment: two options

26 Treating the target Requires a deep knowledge of the genomic background of each case. Time and cost consuming. Feasible only in a few centers. 9 R/R pts have been treated. Median age 24 yrs (range 18-62). 8 pts treated on the ruxolitinib arm (7 pts CRLF2-high, 1 with a JAK2 fusion (HMBOX1-JAK2). 1 pt on the dasatinib arm (NUP214/ABL1). No DLT, but no reponse on ruxo or dasa. Jain N et al, ASH 2017 Pui CH et al, Clin Lymp Myel Leuk, 2017

27 Wide-spectrum appraoch. Ponatinib p= In vitro use of ponatinib on primary cells: effect on proliferation and apoptotic response similar in BCR/ABL1+ and BCR/ABL1- like cases (2 EBF1/PDGFRBpositive, 1 JAK2-mutated and P2RY8/CRLF2-positive, 1 RCSD1/ABL1, 3 WT for JAK/STAT and RAS mutations) p=0.023 Chiaretti S et al, BJH 2018

28 Conclusions BCR-ABL1-like ALL represents a novel ALL subtype. - Detected only in B-neg ALL cases and more frequently from adolescence onwards. - The genomic background is highly heterogeneous and variable from case to case. It can be summarized in lesions involving ABL class genes, JAK/STAT genes and other kinases. Some cases are devoid of lesions. - Diagnosis is still not standardized and represents an unmeet need. - Therapy should include a TKI, possibly in combination with steroids and chemotherapy. Upfront? At MRD persistence (different strategies in children and adults). In combination with immunotherapy?

29 Acknowledgments Monica Messina Alessia Lauretti Alfonso Piciocchi Nadia Peragine Giorgio Inghirami Oliver Elemento Antonella Vitale Anna Guarini Robin Foà