1. Adult AML Slide Deck Master
Clofarabine for T-cell Lymphomas
Not as yet preliminary results from a phase I/II trial
Steven M Horwitz Assistant Attending Memorial Sloan Kettering Cancer Center New York, New York

2. clofarabine structure
Adult AML Slide Deck Master
clofarabine structure
12/6/2018 5:49 AM
Clofarabine is a Hybrid Purine Nucleoside Analog N H N H N N H H N H 2 2 2 2 2 N N N N N N N N N N N N N N N N C l N F N N C l N H O O H O O H H O O O O H O O H O F
Clofarabine is a next generation nucleoside analog
Clofarabine, FLU (fludarabine) and CLAD (cladribine) are halogenated cogneners of deoxyadenosine.
Addition of Fluorine in the arabinose position on the C2 position of the ribose sugar imparts resistance to phosphorolysis H O H O O H H O O H O
deoxyadenosine
Cladribine
Fludarabine
Clofarabine
Montgomery, J. Med. Chem. 1992, 35:

3. Addition of chlorine provides resistance to deamination
Adult AML Slide Deck Master
12/6/2018 5:49 AM
clofarabine stability
Addition of chlorine provides resistance to deamination N H 2 N N
Addition of fluorine provides resistance to phosphorylytic cleavage N C l N H O O
This slide reviews the structural features of Clofarabine that impart the inherent stability of the compound
Addition of Chlorine makes Clofarabine resistant to deamination (click to advance animation)
Addition of Fluorine in the arabinose position makes Clofarabine resistant to phosphorylytic cleavage and stable at acid pH F H O
Montgomery, J. Med. Chem. 1992, 35:
Xie KC, Plunkett W, Cancer Research. 1996, 56:

4. comparison of metabolism
Adult AML Slide Deck Master
12/6/2018 5:49 AM
comparison of metabolism
Fludarabine Cladribine Clofarabine
Glycosidic bond cleavage
pH < 3 stable labile stable
Major metabolites
in cells TP MP+TP MP+TP
Triphosphate Half-life
hrs in CLL > >24

5. Adult AML Slide Deck Master
stereospecificity for cytotoxicity The arabino configuration is important
Parker, Mol Pharmacol. 55, 515, 1999

6. Mechanism of Action: Clofarabine
Adult AML Slide Deck Master
Mechanism of Action: Clofarabine
12/6/2018 5:49 AM
moa
Inhibits ribonucleotide reductase (RnR), resulting in decreased nucleotide pools
Substrate for DNA polymerases to prevent DNA strand elongation
Clofarabine disrupts the mitochondrial membrane.
dNTP pool
dNTP pool
dNTP pool
dNTP pool
dNTP pool
DNA
incorporation
Inhibition of
DNA synthesis/
repair
RNR
incorporation P P P P P P P P P P P P
Polymerase
Polymerase
Polymerase
Polymerase
Polymerase a a a a a , , , , , ε ε ε ε ε
Clofarabine
Inhibition of
DNA synthesis/
Mitochondria ΔΨ ΔΨ ΔΨ ΔΨ ΔΨ 5 5 5 ’ ’ ’ ’ NT NT NT
dCK
dCK
dCK
dCK m m m m m
Cytochrome C
Cytochrome C
Cytochrome C
Cytochrome C
Clofarabine is a prodrug which enters the cell through both equilibrative and concentrative nucleoside transporters.
Once in the cell, Clofarabine is initially phosphorylated by deoxycytidine kinase or dCK to Clofarabine-Monophosphate and is subsequently phosphorylated by other kinases to its active form, Clofarabine-Triphosphate.
dCK is upregulated in transformed cells
Clofarabine acts through several mechanisms to lead to cell death.
Inhibition of DNA synthesis by clofarabine is due to the inhibition of ribonucleotide reductase activity and inhibition of chain elongation by DNA polymerases
dysruption of mitochondrial membrane integrity leading to the release of pro-apoptotic mitochondrial proteins , cytochrome C and apoptosis-inducing factor.
Clofarabine
Release
Release
Release
Release
Transporter
CELL DEATH
Clofarabine
Parker, Cancer Res 1991, 51:
Xie and Plunkett, Cancer Res 1996;56:
Genini, Blood 2000, 96:

7. clofarabine as a single agent in relapsed/refractory adult AML
Adult AML Slide Deck Master
12/6/2018 5:49 AM
clofarabine as a single agent in relapsed/refractory adult AML
Study n
Regimen
Response
Therapeutic Area
Safety
Profile
Kantarjian
J.Clinical Oncology 2003 51
(32 AML)
Phase I
CLO mg/m2 /d x 5
ORR (AML/ALL) = 16%
CR=6%
Solid Tumors,
Acute Leukemias, Myeloma,
Lymphoma
DLT= Transient Gd 3/4 elevations in bilirubin and transaminases
Blood 2003 62
(31 AML)
Phase II
CLO 40 mg/m2/d x 5
CR(AML)=42%
ORR(AML)=55%
*ORR 56% in pts with unfavorable cytogenetics
AML
MDS,CML-BP, ALL
Myelosuppression, infections, transient elevations in bilirubin and transaminases, rash, N/V
*Foran
Clo 221
ASCO 2003 Poster 32
(21 eval)
1 CRp, 1 PR
Median age 63 years
N/V, diarrhea, transient transaminitis, stomatitis
*Genzyme-sponsored study

8. Adult AML Slide Deck Master
anecdotes
23 y/o male with relapsed T-lymphoblastic lymphoma (s/p ALL-2)
Biopsy at relapse showed NK/T-cell lymphoma (CD3+, CD56+, TdT-)
Refractory Hyper-CVAD
CR to first cycle of clofarabine, then proceeded to allo transplant
In previous cloforabine studies
2 other patients with T-cell disase, T-ALL, both CR
Dose and schedule for Phase 1/2 combination studies of clofarabine and Ara-C were based on in vitro studies.

9. clofarabine in combination
Adult AML Slide Deck Master
clofarabine in combination
12/6/2018 5:49 AM
Study n
Regimen
Response
Disease
Safety Profile
*Faderl Blood 2005 32
CLO 40 mg/m2/d x5 + Ara-C 1 g/m²/d x5
CR (AML) =28%
ORR (AML) =40%
AML, MDS, ALL, CML-BP
Myelosuppression, infection, LFT, N/V, rash, diarrhea, PPE, mucositis,
Faderl
Blood 2006 60
CLO 40 mg/m2 + Ara-C 1g/m2
CR= 52%
CRp=8%
ORR=60%
AML or High Risk MDS
Myelosuppression, infections, diarrhea, N/V, headache, rash, PPE,, LFT
Foran
ASH 2005
Abstract 4
CLO 30 mg/m2/d x 5 + Ara-C 100 mg/m2/d x 7
CR = 50%
AML
> 60 years
2 infection related deaths
Dose amended to 25% reduction
Burnett British Soc Hem 2006 30
ClO mg/m²/d x 5 + Daunorubicin 50g/m²/d x3
CR=83%
Suitable for intensive chemo
Not available

10. Adult AML Slide Deck Master
rationale
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Anecdotal responses in T-cell and NK/T lymphoma
Long intracellular half-life
Moderate dose dependent toxicities
Good penetration into tissues (extranodal disease)
Potential for combination studies with other active drugs for T-cell lymphoma
Dose and schedule for Phase 1/2 combination studies of clofarabine and Ara-C were based on in vitro studies.

11. objectives-phase I/II study
Adult AML Slide Deck Master
objectives-phase I/II study
12/6/2018 5:49 AM
Determine the MTD and DLT for clofarabine on a q3 week cycle in relapsed/refractory T-cell lymphoma
Preliminary assessment of Efficacy in T-cell lymphoma
Explore basis for future combination studies
Dose and schedule for Phase 1/2 combination studies of clofarabine and Ara-C were based on in vitro studies.

12. Adult AML Slide Deck Master
design
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Phase I/II Study
Relapsed/Refractory NK/T or T-cell lymphoma
Post transplantation allowed
clofarabine
clofarabine 22 23 24 1 2 3 4 5
Dose and schedule for Phase 1/2 combination studies of clofarabine and Ara-C were based on in vitro studies.
Accelerated titration design-start at 4 mg/m2 daily x 3
Grade 2 non-hematologic toxicity-traditional dose escalation 3-6 pt/cohorts
Can modify to 5d regimen of q2week regimen
Treat expanded cohort at MTD
So far-2nd dose level (8mg/m2)