Non Transplant Options

for Aplastic Anemia

Jong Wook Lee, MD, PhD.

Department of Hematology, Seoul St. Mary’s Hospital

The Catholic University of Korea, Seoul, Korea

CytotoxicT cell

CytotoxicT cell

CytotoxicT cell

CytotoxicT cell

Haemopoieticcell

Interleukin-2

TNF-β

INF-γ

Fas ligand

INF-γ

TNF-β

INF-γreceptor

TNF-βreceptor

Fasreceptor

Expansion of T-cell clones

IRF-1

↑ ICE apoptosis

Jak 1

Stat 1

Stat 3

i NOS

NOToxicity for other cells

dsRNA-PKR

RNaseL

elF-2↓protein synthesis

↓Cell cycling

Immune-mediated aplastic anemia

Anemia

Leukopenia (neutropenia)

Thrombocytopenia

CD34+ cell↓

CFU-GM, BFU-E ↓

LTC-IC ↓

Young N, Maciejewski J. N Engl J Med. 1997;336:1365-72. Young N, et al. Blood 2006;108:2509-19.

Genetic aplastic anemia

Telomere shortening1–3

● Abnormal telomere shorting in AA: relapse after IST, clonal evolution, survival in SAA

1. Ball SE, et al. Blood. 1998;91:3582-92. 2. Brummendorf TH, et al. Blood 2001;97:895-900. 3. Scheinberg P, et al. J Am Med Assoc. 2010;304:1358-64.

4. Yamaguchi H, et al. N Engl J Med. 2005;352:1413-24.5. Fogarty PF, et al. Lancet. 2003;362:1628-30. 6. Yamaguchi H, et al. Blood. 2003;102:916-8.

Mutation in both TERT and TERC genes4–6

● Mutation of TERC: 3/210 of AA, PNH, and DKC ● Mutation of TERC in 2 families ● Mutation of TERT: 7/200 AA vs 0/528 control

– heterozygous mutations in the TERT gene impair telomerase activity and may be risk factors for bone marrow failure

Embryonicstem cells

Adultstem cells

Telomeres

Chromosome

Telomere is a repeating

DNAsequence

Telomeraseinactive

or absent

Telomereshort

Telomerelong

Telomeraseactive

DKC = dominant dyskeratosis congenita; IST = immunosuppressive therapy; PNH = paroxysmal nocturnal haemoglobinuria; SAA = severe AA.

Auto-antibodies (Ab) against distinct proteinsKinectin - a 1300 amino acid protein expressed on human hematopoietic cell, liver, ovary, testis & brain - IgG Anti-kinectin Ab found in 7/18 (39%) of US AA vs. in 3/30 (10%) of Japanese AA

(Hirano, Blood, 2003)

Anti-PMS1 (postmeiotic segregation increased 1) Ab - Japanese AA, 3/10 (30%) vs. US AA, 0% (0/18) (Hirano, BJH, 2004)

Diazepam-binding related protein-1

- anti-DRS-1 in 38% (27/71) of PNH+AA, 6.3% (2/32) of PNH-AA,

38.5% (5/13) of PNH+MDS.

- response to IST in AA patients with DRS-1 Ab (100% vs. 55%) (Feng, Blood, 2004)

Moesin:

- a membrane cytoskeleton linker protein

- antimoesin Abs in 25/67 (37%) of AA - marker for response to IST (Takamatsu, Blood, 2007)

Treatment options of aplastic anemia

1. Allogeneic BMT– age < 50 years– HLA-matched donor (+)– severe or very severe AA

2. Immunosuppressive therapy (IST)– hematological recovery in 60~70% of patients1,2

3. Supportive care– blood transfusions, management of infections, iron

chelation therapy

1. Rosenfeld SJ, et al. Blood. 1995;85:3058-65.2. Bacigalupo A, et al. Blood. 1995;85:1348-53.BMT = bone marrow transplantation.

Supportive care

1. Transfusion

Red cell (PRC) : Hb 7.0 g/dl ↓

PLT : 10,000/mm3 ↓

* Irradiation (2,500 rad)

* Leukocyte-depleted filter

* Iron chelation therapy

2. Infection control

Algorithm for treatment

Maciejewski JP, Risitano AM. Hematology Am Soc Hematol Educ Program. 2005:110-7.

BMT

uBMT;androgens, 2nd IST

Adequate blood count

Observation

Blood counts

Acquired AA

Severe Non-severe

Older adults

No family donor

ATG+CsA

R

Follow for relapse,late clonal ds

Allo-BMT;androgens, 2nd IST

Clinical status

ATG+CsA

R

CsA; androgen, 2nd IST

ATG+CsA

Clinical statusTx-

dependentChildren,

young adults

Age

Siblingmatch

HLA typing

Blood count improvement

at 3–6 mo

Non-R

Non-R Non-R

ATG = antithrombocyte globulin; CsA = ciclosporin; ds = disease; R = response; uBMT = unrelated bone marrow transplantations.

ATG

CyclophosphamideCyclosporin

Inhibition of T cell function

Proposed mechanisms through which ATG can interfere with the immune response

1) T-cell depletion in blood and lymphoid tissues through complement-dependent lysis and T-cell apoptosis

2) induction of B-cell apoptosis

3) modulation of cell-surface molecules (adhesion and chemokine receptors) that mediate leukocyte/endothelium interactions

4) Interference with DC functional properties (maturation and migration)

5) Induction of Treg and NK-T cells.

Morthy M. Leukemia 21:1387–1394, 2007

Comparative study of IST in AA

Treatment

(mg/kg/day)

No. of patients (SAA)

RR (CR+PR) Survival

(F/U duration)

German study

ATG+CsA (12)

ATG43 (31)

41 (29)

70%

46% p<0.05

64% (41 mon)

58% p=0.16

NIH ATG+CsA (12)

ATG51 (51)

70 (70)

67%

29% p=0.001

62% (3 yrs)

_

EBMT ATG+CsA (5)

CsA

54 (0)

61 (0)

74%

46% p=0.02

90% (6 mon)*

67% p=0.001

Korean

study

ATG+CsA (5)

ATG

53 (39)

40 (27)

41%

18% p=0.01

98% (6 mon)

90% p=0.16

Factors for predicting response of IST

● Young age● Presence of PNH clone● Higher reticulocyte count (>25x109/L)● Higher lymphocyte counts (>1x109/L)● Early treatment within 21 days after diagnosis● CsA therapeutic level/late tapering● HLA-DR15

Predictors for response to IST (n=324)

Kim SY et al, GENES, CHROMOSOMES & CANCER 49:844–850, 2010

Factors to predict relapse after IST

● Shorter telomeres● Rapid tapering of CsA : General recommendation is to taper CsA by 10% every month,

starting at least 1 year from IST (Sarraco et al, Br J Haematol 2008,

Marsh et al, Br J Haematol 2009)

● Suboptimal therapeutic range of CsA

Improving on ATG+CSA

● Addition of G-CSF

- improved neutrophil counts but similar response or survival

● Addition of other immunosuppressive drugs

+ High dose of methylprednisolone: toxicities ↑

+ MMF: no benefit (Scheinberg, 2006)

+ Rapamycin/sirolimus: not superior (Scheinberg, 2009)

Alternative immunosuppressive regimens

● Cyclophosphamide (CY)1) John’s Hopkins (Brodsky et al, 1996 & 2010)

- CY 50 mg/kg/d for 4d in 33 untreated SAA

- CR at 50 months; 65%, OS 86%

2) NIH trial (Tisdale et al, 2000 & 2002)

- CY (50 mg/kg/d for 4d) + CsA vs. ATG + CSA

- Response rate: 6/13 (46%) vs. 9/12 (75%)

- Higher fungal infection & death in CY arm

● Alemtuzumab 1) EBMT study : OR 58% (Risitano et al, Br J Haematol 2010)

2) NIH study : 10mg/d, IV for 10 days (Scheinberg et al, Blood 2012)

Refractory SAA (n=27) : OR 37%

Relapsed disease (n=25) : OR 56%

Treatment-naïve (n=16) : OR 19%

Moderate AA

● IST– Better survival with ATG + CyA than CyA alone

(Marsh, 1999)

● Daclizumab (humanized monoclonal Ab of IL-2 receptor)

- 16 AA : response 6/16 (38%) with little toxicity (Maciejewski, 2003)

● Androgen

- stimulate TERT expression via estrogen receptor

increase telomerase activity in lymphocytes and CD34+ cells

(Calado, 2005)

Various ATG preparations

● Horse ATG

- ATGAM® (40 mg/kg per day for 4 d)

- Lymphoglobuline® (15 mg/kg per day for 5 d)

● Rabbit ATG

- Thymoglobuline® (2.5~3.75mg/kg per day for 5 D)

- ATG-Fresenius® (5mg/kg/d for 5 D)

: inferior response to hATG (53% vs 78%, p=0.03),

5 yr-OS (66% vs 80%, p=0.002)

(Zheng et al, Exp Hematol 34:826, 2006)

Phase II prospective study from Cleveland

SAA treated with rabbit ATG (n=21) versus matched historical control treated with horse ATG (n=67).

Rabbit ATG : 3.5 mg/kg/day for 5 days Horse ATG: 40mg/kg/day for 4 days

Cyclosporine – maintain until response or developing renal complications.

Afable et al, Haematologica 96:1269, 2011

Type of ATG

n Early mortality

Response (%) Relapse (%)

3 months 6 months 12 months

Rabbit ATG 20 2 40 (CR: 0) 45 (CR: 0) 50 (CR: 0) 5

Horse ATG 67 6 55 (CR: 5) 58 (CR: 6) 58 (CR: 8) 16

p value 1.0 0.43 0.44 0.61 0.28

Atta et al, Annals of hematology. 89:851, 2010

Horse ATG Rabbit ATG p value

N 42 29

Partial response at 6 months 20 (47.6%) 8 (27.6%) 0.14

Complete response at 6 months 5 (11.9%) 2 (6.9%) 0.69

Overall response at 6 months 25 (59.5%) 10 (34.5%) 0.05

Interval between ATG and response (days) 99 (39-166) 88.5 (39-161)

0.98

Overall survival at 2 years 78.4% 55.4% 0.03

Relapse 9 (36%) 3 (30%) 1

A retrospective comparative study from Brazil

Rabbit ATG : 2.5 mg/kg (range, 1.5-3 mg/kg),

Horse ATG : 15 mg/kg (range, 10-21.4 mg/kg)

Randomized prospective study in NIH (n=120)

Rabbit ATG (3.5mg/kg for 5 days) vs Horse ATG (40mg/kg for 4 days)

Cyclosporine – continued for at least for 6 months

Scheinberg et al, NEJM 365:430, 2011

Response Horse ATG

Rabbit ATG

p

3 months 37 (62) 20 (33%) 0.002

6 months 41 (68%) 22 (37%) <0.001

Comparison of response between horse ATG (n=53) and rabbit ATG group (n=46) in SAA

Horse ATG (OR)Rabbit ATG (OR)Horse ATG (CR)Rabbit ATG (CR)

p value OR 0.319 0.834 0.925 0.757

CR 0.123 0.048 0.387 0.905

Shin, et al 2011 ASH #3430

Similar OS between rATG and hATG

P=0.46

Shin, et al 2011 ASH #3430

Lymphocytes depletion after ATG treatment

Scheinberg et al, NEJM 365:430, 2011

Lymphocytes depletion after ATG treatment

Scheinberg et al, NEJM 365:430, 2011

Comparison between horse and rabbit ATG

2.5mg/kg/d Х 5 days

3.5mg/kg/d Х 5 days

3.5mg/kg/d Х 5 days

3.5mg/kg/d Х 5 days

2.5mg/kg/d Х 5 days

n=22 n=67n=29 n=42 n=60 n=60 n=33 n=29 n=53 n=46

(1) Atta E, et al. Ann Hematol. 2010 89(9):851-9. (2) Sheinberg, et al. N Engl J Med. 2011 365(5):430-8.

(3) Chang M, et al. Eur J Haematol. 2010 84(2):154-9. (4) Afable M, et al. Haematologica. 2011 96(9):1269-75

(5) Shin, et al 2011 ASH #3430.

International Clinical Trial

● A prospective randomized multicenter study comparing different dosages of rabbit antithymocyte globulin (Thymoglobuline) in patients with severe aplastic anemia

● A total of 256 SAA patients will be enrolled, with 128 patients randomized to arm A (2.5mg/kg/d) and 128 patients to arm B (3.5mg/kg/d).

● Korea, Japan, China

Conclusions

1. For the last 20 years, the standard immunosuppressive regimen for AA patients has been ATG plus CsA, which results in response rates ranging between 50% and 60%, and even higher overall survival.

2. Response rates of rabbit ATG seem to be similar to horse ATG.

3. Thymoglobuline might need more time to reach the best response after IST in comparison to horse ATG.

4. CsA should be continued for at least 12 months after achieving

maximal hematological response, followed by a very slow tapering,

to reduce the risk of relapse.

5. Optimal dose of rabbit ATG to be determined.