12th Congress of the European Hematology ... - Haematologica

enewal potential of X-RAR-positive HSC as revealed by a significantly
reduced replating efficiency. Conclusions. Here we provide first evidence
that it is possible to the exposure to therapeutically achievable dosages
of a NSAID revert the aberrant activation of the Wnt-signaling by LAFP.
The significant reduction of the aberrant self renewal potential of HSC
in the presence of X-RAR further support that the inhibition of the aberrantly
activated Wnt signaling in AML might be a valid molecular therapy
approach which has to further validated in in vivo leukemia models
and in a clinical setting.
0323
MTORC1 INHIBITION ACTIVATES PI3K/AKT BY UP-REGULATING THE IGF-R SIGNALLING
IN ACUTE MYELOID LEUKAEMIA: RATIONAL FOR INHIBITION OF BOTH PATHWAYS
J. Tamburini, 1 N. Chapuis, 1 V. Bardet, 1 S. Park, 1 N. Ifrah, 2 F. Dreyfus, 3
P. Mayeux, 1 C. Lacombe, 1 D. Bouscary1 1 Institut Cochin, PARIS; 2 Service des Maladies du Sang CHU, ANGERS; 3 Service
d'hématologie Hopital Cochin, PARIS, France
Background. The PI3K/Akt and mTORC1 pathways are frequently activated
in acute myeloid leukaemia (AML) cells and represent potential
therapeutic targets. Aims. In this report, we studied the interactions
between PI3K and mTORC1 pathways in primary blast cells from the
bone marrow of patients with AML at diagnosis after 4 hours of cytokine
and serum starvation. Methods. Bone marrow samples were obtained
from 22 newly diagnosed AML patients before induction of chemotherapy.
Blast cells were starved 4 hours in cytokine and serum free media.
Cells were then incubated with or without inhibitors: IC87114
(p110delta specific inhibitor), RAD001 (rapamycin derivative inhibitor
of mTORC1), LY294002 (multiple kinases inhibitor including mTORC1
and PI3K), ?IR3 (inhibitory anti-IGF-1 receptor monoclonal antibody)
and Western blot analysis and cell proliferation assays were performed.
To purify the blast cell population, cells were sorted according to CD45
expression and side scatter. Immunofluorescence on cytocentrifuge
preparations and quantitative RT-PCR for IGF-1 expression on CD45low
blast cells were performed. Results. We observed that specific inhibition
of mTORC1 activity with RAD001 upregulated PI3K activity, as evidenced
by an increased phosphorylation of AKT (Ser 473) on Western
blot analysis. The mean increase of AKT phosphorylation in the presence
of RAD001 was 186% (130-500%). The increase was maintained
after 24 hours of blast cell incubation with RAD001. This mTORC1mediated
Akt up-regulation was explained by an IGF-1/IGF-1 receptor
autocrine loop: a/ blast cells expressed a functional IGF-1 receptor and
IGF-1-induced Akt activation was increased by RAD001, b/ a neutralizing
anti-IGF-1R α-IR3 monoclonal antibody reversed the RAD001induced
Akt phosphorylation, c/ autocrine production of IGF-1 was
detected in highly purified CD45low blast cells from 8 patients by quantitative
RT-PCR and immunofluorescence. d/ Activation of the IGF-1
receptor paralleled an up-regulation of the IRS2 adaptor protein. Finally,
we observed, in AML blast cells, a dissociation of the mTORC1 and
PI3K pathways: IC87114, a specific p110delta PI3K inhibitor, had no
inhibitory effect on mTORC1 signalling. This was confirmed by the fact
that RAD001 and IC87114 induced additive anti-proliferative effects on
AML blast cells. Conclusions. Our results suggest that dual inhibition of
mTORC1 complex and IGF1/IGF-1R/PI3K/Akt axis may enhance the
efficacy of mTOR inhibitors in AML.
0324
DISCOVERY AND PRECLINICAL DEVELOPMENT OF SELECTIVE JAK INHIBITORS FOR THE
TREATMENT OF HEMATOLOGICAL MALIGNANCIES
S. Fridman, J. Li, P. Liu, E. Caulder, E. Crowgey, M. Favata, G. Guoen,
Y. Li, M. Covington, J. Rodgers, A. Combs, S. Yeleswaram,
R. Newton, P. Scherle, K. Vaddi
Incyte Corporation, WILMINGTON, USA
Background. Increased kinase activity is a common occurrence in malignant
cells and is associated with the activation of multiple downstream
pathways implicated in the pathobiology of various diseases. The identification
of activating genetic mutations in multiple members of the Janus
kinase (JAK) family of non-receptor tyrosine kinases has recently been
described in numerous malignancies, including the majority of Philadelphia
chromosome negative (Ph – ) myeloproliferative disorders (MPDs).
Mouse experiments have suggested a causal role for these mutated JAKs
implicating them in the pathogenesis of the human conditions. This suggests
that JAK inhibition may be a promising therapeutic strategy for the
treatment of diseases associated with elevated JAK kinase activity. Aims.
To identify and characterize potent, selective, orally bioavailable inhibitors
12 th Congress of the European Hematology Association
of the JAK kinases. Methods. A compound library was screened for inhibition
of JAK kinase activity. Structure-activity relationship based lead
optimization was conducted to identify novel, potent and selective JAK
inhibitors. Further characterization of in vitro and in vivo pharmacokinetic,
pharmacological and toxicological properties was performed to identify
molecules suitable for further advancement into animal models of cancer
and MPDs. Results. Incyte has identified multiple novel, potent, and selective
inhibitors of the JAK kinases, including the recently described JAK2
V617F mutant common to the majority of the Ph- MPDs. These compounds
are active against JAKs at nanomolar concentrations while demonstrating
excellent selectivity against a broad panel of unrelated kinases. In
cell-based assays, these compounds retain their nanomolar potency and
selectivity and reduce JAK-mediated tumor cell growth and survival. In
contrast, these compounds do not impact the growth or survival of cells
dependent on alternative signaling pathways (e.g. Bcr-Abl), even at micromolar
concentrations. in vivo, in a mouse model of MPD-associated mutant
JAK2 driven organomegaly (BaF/3-JAK2V617F cells), Incyte JAK inhibitors
markedly reduce the splenomegaly common to both the model and the
human disease (Figure 1). A detailed characterization of the activity of
these molecules will be presented. Summary. Increased JAK signaling has
been associated with various malignancies, including the majority of Ph-
MPDs. Incyte has identified multiple potent and selective inhibitors of the
JAK kinases. These compounds are orally bioavailable and are efficacious
in vivo at well tolerated doses. As such, these compounds may be promising
new therapeutics for the treatment of MPDs and other disease states
associated with elevated JAK activity.
Figure 1. Normalization of MPD-related splenomegaly.
0325
GEMTUZUMAB OZOGAMICIN (MYLOTARG ) AS MAINTENANCE THERAPY AFTER
AUTOLOGOUS STEM CELL TRANSPLANTATION IN ELDERLY PATIENTS WITH ACUTE
MYELOID LEUKEMIA
N. Cascavilla, C. Bodenizza, A.M. Carella, M. Dell'Olio, A.P. Falcone,
M.M. Greco, A. La Sala, S. Mantuano, L. Melillo, E. Merla, M. Nobile,
G. Sanpaolo, P. Scalzulli
Hematology CSS Hospital, SAN GIOVANNI ROTONDO, Italy
Background. Acute Myeloid Leukemia (AML) is a disease predominantly
affecting older adults, with a median age of 65 years. These patients represent
a poor risk population with a high chemotherapy-related mortality.
Standard of care in the management of elderly patients with AML
includes combination therapy with cytarabine plus fludarabine and an
anthracycline and can result in complete remission (CR) rate of 40% to
60%. Even if successful, patients relapse quickly. Aggressive postremission
therapy does not appear to improve survival. Future directions include
therapies targeted at immunomodulation, and, among newer treatments,
Gemtuzumab ozogamicin (GO) has given promising results in relapsed,
refractory and untreated CD33 + AML as monotherapy or combination
regimens or adjunct to conditioning regimens for Stem Cell Transplantation
(SCT). Patients and Methods. In this study we analyzed the efficacy and
the safety of GO as maintenance treatment after Autologous-SCT in three
elderly patients (2 males and 1 female; 64, 67 and 69 years, respectively)
with CD33 + AML in 1st (2 patients) or in 2nd (1 patient) CR. After a complete
hematopoietic engraftment (at least 6 weeks from ASCT), GO was
administered alone for 4 doses with 28 days between doses (two patients
received 6 mg/mq for the two first doses and 3 mg/mq for the two last doses;
the last received 3 mg/mq for four doses). Patients were evaluated for
Continue Complete Remission (CCR) and therapy-related adverse events.
Results. All patients are in CCR at +17, +13, +11 months, respectively.
Neutropenia (between 500 and 1000/microl) and thrombocytopenia
(between 50.000 and 100.000/microl) were observed in two (when GO
was given at a dose of 6 mg/mq) and in three patients, respectively. No
grade 3 or 4 liver toxicity was observed. Conclusions. GO administered in
fractionated doses as maintenance treatment after ASCT in older patients
with CD33 + AML in first or second CR demonstrated a good efficacy and
an acceptable safety profile.
haematologica/the hematology journal | 2007; 92(s1) | 117