1. New Targets – New therapies
Hairy Cell Leukemia:
New Targets – New therapies
Robert J. Kreitman, M.D.

2. HAIRY CELL LEUKEMIA B-cell leukemia
2% of all leukemias, ~1000 cases/yr
‘Fried egg’ appearance
Cytoplasmic projections
CD19, CD22, CD103, CD11c, CD25

3. 313 HCL randomized between DCF and IFN
NEJM (1984) 310: CR, 4 PR (n=8) (other trials, CR 5-10%, PR 30-70%, DFS 1.5-3Y)
NEJM (1987) 316: CR, 10 PR (n=27) (other trials: CR 70-75%, PR 3-25%, DFS: 8Y 76%
JCO, 13:974, 1995
313 HCL randomized between DCF and IFN
CR rates: DCF 76% vs IFN 11%, (p<0.001)
DCF CR 66% in patients crossing over after IFN

4. Single 5-7 day course of treatment
NEJM (1990) 322: /12 CR, 1/12 PR
CR 75-87%, PR 11-25%, DFS: 4Y 79-84%, 7Y 60-66%
Single 5-7 day course of treatment
Evidence is lacking that cladribine can eradicate HCL:
Saven et al., Blood 92:1918, 1998, Goodman et al, JCO 21:891, 2003
No plateau seen on DFS curves
Sigal et al., Blood, 115:1893, 2010.
Of 358 HCL, MRD was present in 10 of 19 patients still in CR 16y later.

5. Outcome of HCL after purine analog
Else et al., BJH, 145:733, 2009
Median RFS 16y (1st line), 11y (2nd line), 6.5y (3rd line)
Median time to purine refractory disease ~ 34 years
Golomb et al, Ann Int Med, 89:677, 1978 – Median OS of HCL = 4 years

6. HCLc and HCLv Clinical features: Type HCLc HCLv
% of HCL ~80-90% ~10-20%
Spleen enlargement Mild Severe
Lymph nodes - +
Cytopenias + -
Lymphocytosis - +
CD25 + -
CD /-
Annexin A1 + -
Response to CdA, DCF 70-87% CR 8% CR

7. Classic
HCL
HCLv

8. Classic
HCL
HCLv
VH4-34+ HCL
Arons et al., Blood, 14:4687, 2009

9. Mab-based treatment for HCL
Blood (2003) 102: % CR, 13% PR (n=24)
All patients had prior purine analog and needed treatment
Better results if no prior purine analog, or no cytopenias.
Rituximab with cladribine or pentostatin better, but more toxic

10. Cladribine followed by Rituximab For Early HCL
Rituximab followed Cladribine by 1 month
100% CR in 31 patients, MRD+ in 6 (21%) of 28
In HCLv, 2 of 5 had CRs lasting 12 and 35 mo
Ravandi et al., Blood, 118:3818, 2011

11. Pentostatin Cladribine Benzimidazole Ring Nitrogen mustard
Structure of bendamustine compared with other purine analogs
NH2 N N N N N N Cl N N N
Pentostatin
Cladribine O O
HOCH2
HOCH2 OH Cl OH
Benzimidazole
Ring N N Cl
Nitrogen mustard
COOH N
Bendamustine
CH3

12. C1D24 C7D42 Response in HCL adenopathy to bendamustine
Kreitman et al., Leuk & Lymphoma, 52:1153, 2011

13. Burotto et al., CCR, in Press
Bendamustine and rituximab for multiply relapsed/refractory HCL
Cy1 Cy2 Cy3 Cy Cy Cy6
| | | | | | | | | | | |
D1 D15 D1 D15 D1 D15 D1 D15 D1 D15 D1 D15
Bendamustine 70 mg/m2 d1,2 (n=6)
Bendamustine 90 mg/m2 d1,2 (n=6+28)
Rituximab 375 mg/m2 d1, 15
Bendamustine Dose
(mg/m2 d1, 2)
Patients enrolled 6 6
ORR (CR+PR) 6 (100%) 6 (100%
CR 3 (50%) 4 (67%)
MRD-negative 2 (33%) 4 (67%)
Med Time to CR 223 days 111 days
Burotto et al., CCR, in Press

14. Recombinant immunotoxins for HCL
Blood (1999) 94: % CR, 75% PR (n=4)
(LMB-2)
NEJM (2001) 345: % CR, 13% PR (n=16)
JCO (2005) 23: % CR, 19% PR (n=31)
(BL22)

15. BL22
BL22

16. Moxetumomab pasudotox: High affinity mutant of BL22
>1000
CLL
HCL
BL22 V V V V L L L H
Median ratio II Ib
III
1000
2.4 (n=32) - S
Moxe
100 V V V V L L L H II Ib
III
IC50 (ng/ml) - S 10
SSY  THW at 100a, b, c.
Affinity  14x (lower koff).
Originally HA22, renamed CAT-8015, then moxetumomab pasudotox.
Salvatore et al, CCR, 8:995, 2002 1
Median ratio
8.3 (n=30)
0.1
BL22
Moxe
BL22
Moxe

17. Moxetumomab pasudotox in HCL7
CR 13 (46%)
PR 11 (39%) 6
SD 3 (11%)
PD 1 (4%) 5
CR 46%, ORR 86% (n=28)
PATIENTS 4
12 (92%) of 13 CRs > 1yr 3 2 1 10 20 30 40 50
dose (ug/Kg x3)
Kreitman et al., JCO, 30:1823,2012

18. NEJM, 364:2305, 2011 Total exome sequencing of 1 HCL patient
BRAF V600E & in 47 other HCL patients vs 0 of 195 other hematologic tumors
BRAF V600E also in 50% melanomas, 7% colon cancers, other cancers
HCLv not characterized
Multiple studies confirmed 100% expression of BRAF V600E in HCL
BRAF WT in 16/16 HCLv, 5/5 classic HCL expressing VH4-34, & 6 others
Thus, HCL is a heterogeneous disease with some variants defined only molecularly, and BRAF can be WT even in classic HCL
MEK mutations have been found in HCLv and VH4-34+ HCL

19. Rapid development of agents targeting the MAPK pathway
6/30/11 (NEJM) Vemurafenib vs DTIC for V600E+ melanoma showed ^OS, PFS, leading to FDA approval for melanoma.
Chapman et al., NEJM, 364:2507, 2011

20. Vemurafenib 240 mg q12 x56 days
Dietrich S et al., NEJM, 366(21): , 2012

21. 6-Month follow-up of HCL treated with Vemurafenib
Dietrich S et al., JCO, 31:e300, 2013

22. Transient response of HCL to Vemurafenib
(240 mg po q12 x58 days)
Pre Day Day 60
Follows et al., BJH, 161:136, 2013
Thus, Vemurafenib can induce response including CR in HCL, but durability and eradication of MRD has not been reported, and relapse occurs.

23. Other anecdotal responses of HCL to Vemurafenib
27y history of HCL, multiply relapsed (14 times).
Began Vemurafenib 960 mg po q12hr x3 weeks
Stopped due to arthralgias
Arthralgias resolved
HCL decreased by BMBx from 68% to <5% by morphology and 10-20% by IHC.
23y history of HCL, multiply relapsed
Plt 24, Hgb 6.9
Began Vemurafenib 240 mg po q12hr x56d
Day 56: Improved Plt to 253, Hgb 9.6
Day 56: BMBx improved to 5% HCL
Day 90: CR achieved, but MRD not reported
Munoz et al., JCO, 31:e351, Peyrade et al., Haematologica, 98:e20, 2013
Thus, Vemurafenib can induce response including CR in HCL, but durability and eradication of MRD has not been reported, and relapse occurs.

24. Rational for combined BRAF-MEK inhibition (preclinical)
BRAFV600E human melanoma xenograft
500
1000
1500
2000 10 20 30 40 50 60 70 80 90
Untreated control
Trametinib (0.3 mg/kg) (anti-MEK)
Mean tumor volume (mm3) + SEM
Dabrafenib (30 mg/kg)
Dabrafenib (300 mg/kg) (anti-BRAF)
Dabrafenib + trametinib
(30/0.3 mg/kg)
Treatment period (days)
Control
BRAFi (150 mg/kg)
BRAFi + MEKi ( mg/kg)
BRAFi alone results in thickening & hyperkeratosis
Co-dosing with MEKi attenuates lesion formation
Conclusions:
Besides more effective, combined treatment is less toxic than single-agent BRAF or MEK inhibition 24

25. Mutated BRAF no inhibition
BRAF V600E inhibition
MEK inhibition
BRAF+ MEK inhibition
B-Raf
V600E
B-Raf
V600E
B-Raf
V600E
B-Raf
V600E
Malignant phenotype
SCC Kerato-acantoma
Rash
Diarrhea
Lack of toxicity

26. Conclusions:
Purine analogs cladribine and pentostatin are highly effective in 1st-2nd line for HCL, but are usually non-curative and MRD is very common
Patients with HCLv or VH4-34+ classic HCL are primarily resistant
Rituximab has limited activity in HCL as a single agent, but high activity combined with purine analogs
Bendamustine or BR is active in relapsed/refractory HCL
LMB-2 (anti-CD25) is active for CD25+ relapsed/refractory HCL
Moxetumomab pasudotox (HA22), an affinity-matured version of BL22, achieves CRs in ~50% of relapsed/refractory HCL without DLT or MRD
BRAF is an important target in most but not all HCL. Vemurafenib is a useful agent in V600E+ HCL but MRD-negative CR is unreported.
New targets in HCL and HCLv are being sought for therapy.