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

12 th Congress of the European Hematology Association
activity was assessed in immunocompromised mice bearing established
human tumor xenografts. Results. Over 80 compounds were tested for
exposure following oral administration using the pharmcodynamic
assay and 17 were identified that achieve >80% inhibition of the proteasome
in blood and tissues. Pharmacokinetic and pharmacodynamic
analysis showed the bioavailability of the most active compounds to
range from 5-30%. In comparing the oral bioavailability determined in
mice with a variety of properties measured in vitro, we found that oral
bioavailability is associated with increased intrinsic solubility and metabolic
stability and reduced MDR1 sensitivity. The oral bioavailability of
a subset of these compounds was also determined in rats and dogs and
found to be comparable to mice both by pharmacodynamic and pharmacokinetic
analysis. Toxicity studies in mice showed that repeated
oral administration was well tolerated at doses that resulted in >80%
proteasome inhibition in most tissues. Anti-tumor activity of these orally
bioavailable inhibitors was assessed in two human tumor xenograft
models using cell lines derived from hematological malignancy: RL (non-
Hodgkin's lymphoma B cell) and HS-Sultans (Burkitt’s lymphoma). Oral
administration of several of these proteasome inhibitors resulted in antitumor
responses equivalent to intravenously administered PR-171 when
given on the same schedule. Summary. Based on these studies, a number
of potent, orally bioavailable proteasome inhibitors with potential
for the treatment of malignant diseases have been identified for further
pre-clinical and clinical development.
0740
MARKERS OF VIRAL INFECTIONS DURING RITUXIMAB TREATMENT OF PATIENTS WITH
RESISTANT AUTOIMMUNE HEMOLYTIC ANEMIA (AIHA)
T.A. Garanzha, N.V. Tsvetaeva, O.F. Nikulina, N.G. Yaroslavtseva,
D.S. Tikhomirov, T.A. Tupoleva, E.Yu. Varlamova, E.M. Gretsov,
I.A. Vorobyev, F.P. Filatov
National Hematology Research Center RAMS, MOSCOW, Russian Federation
Background. Standard AIHA treatment is based on application of corticosteroids.
Recently rituximab treatment was found to be quite efficient,
especially when AIHA is resistant to standard therapy. Earlier we
have found high incidence of EBV and HBV viral markers in AIHA
patients. Some publications report the cases where rituximab treatment
bear high risk of complications due to viral and bacterial infections. Aims.
to monitor markers of viral infections during treatment of AIHA patients,
in particular using Rituximab. Materials and Methods. EIA (IgM-VCA-
EBV, IgG-EA-EBV, IgG-NA-EBV, IgM-HCMV, IgG-HCMV, IgM-HSV1-
2,IgG-HSV1-2, HBsAg, a-HCV). PCR (DNA-HBV, DNA-EBV, DNA-
CMV, DNA-HSV1-2, DNA-HHV-6, RNA-HCV). Viral DNA(RNA) in
plasma and peripheral blood mononuclear cells (PBMC), saliva and broncheo-alveolar
lavage (BAL) fluid, spleen and liver bioptates was detected
by PCR. 41 AIHA patients were followed. Eight of forty one patients,
diagnosed with resistant AIHA were treated by rituximab and followed
more closely. Results. In 26 patients (including all 8 patients with resistant
AIHA) out of 41 DNA EBV was detected in PBMC. With standard
therapy DNA EBV was detected during all follow-up period (up to 6
years). In 8 patients treated with rituximab CD19, CD20 and CD22 cells
were not detectable about one week after injection. Gradual recovery of
this cell population took about 2-8 months. DNA EBV ceased to be
detectable within 4-8 month after rituximab injection. Patients are in the
state of clinical remission for a period from 6 months up to 3 years.
HCV viral load in plasma of one patient (genotype 1b) was not essentially
influenced by rituximab therapy. High titer of IgG-HSV1-2 (1:6400,
1:12800) was detected in all 8 patients. It was not changed during the
course of treatment. IgG-HCMV titer (initially in the range 1:400 to
1:12800 depending on the patient) was not changed as well. IgG-EA-EBV
was found in serum of 2 patients before, in the course and after rituximab
treatment. Conclusions. Rituximab treatment leads to clinical remission
in patients with resistant AIHA. In this group of patients we did not
detected any signs of activation of viral infections, moreover DNA EBV
was below detectable levels during remission period. Any considerable
reduction of viral antibodies levels was not detected.
276 | haematologica/the hematology journal | 2007; 92(s1)
0741
PHARMACEUTICAL DEVELOPMENT OF HIGH DOSE RADIOLABELLED RITUXIMAB AS
CONSOLIDATION TREATMENT FOR PATIENTS WITH RELAPSED OR REFRACTORY CD20
POSITIVE B-CELL NON-HODGKINS LYMPHOMA
T.H.L. Tran
The Netherlands Cancer Institute, AMSTERDAM, Netherlands
Background. Radioimmunotherapy is a promising target-oriented strategy
for treatment of B-cell non-Hodgkin´s lymphoma. In this study, the
CD20 targeted monoclonal antibody rituximab was coupled to the
radionuclide 131-Iodine. Rituximab induces antitumour activity through
antibody-mediated mechanisms, while 131-Iodine promotes cytotoxic
activity by the crossfire-effect to neighbouring tumour cells with none
or insufficient antigen expression. Disposition of this agent in the patient
can be visualised by gamma imaging. This study was performed to optimise
the pharmaceutical preparation of 131-Iodine labelled rituximab
and the subsequent quality control. Due to the required high dose of
radioactivity, employed for myeloablative treatment, the safety of the
involved personnel is of crucial importance. In addition, the integrity of
rituximab must be maintained. Aims. The aim of this study was to develop
clinical grade 131-Iodine labelled rituximab, maintaining its properties,
with minimum exposure to involved personnel. Methods. A previously
developed labelling method was used as a starting point. First, the
131-Iodine was pretreated to adjust the pH and to protect the radionuclide
from oxidation. The labelling was provoked by addition of 35 µg
of Iodogen in acetonitril (1 mg/mL) to a reaction vial with rituximab
and 131-Iodine. After 3 minutes, the reaction was terminated by addition
of ascorbic acid. Subsequently, the 131-Iodine rituximab conjugate
was purified. All labelling steps were carried out under aseptic conditions
and a sterile filtration was performed as final step. The labelling process
consisted mainly of a remote system, operated with underpressure to
promote the radiation safety. The labelling procedure was performed
under good manufacturing practice (GMP) conditions. Radiation exposure
was monitored by wearing a thermoluminescence detector (TLD)
and an electronic personal dosimeter. Exposure to the fingers was
assessed by attaching TLDs to the fingertips. The annual maximum permissible
dose to radiation workers is 20 mSv. The final product was
characterized by size exclusion chromatography (SEC-HPLC) and by
instant thin layer chromatography (iTLC). The immunoreactive fraction
was determined using a cell-binding assay at antigen excess using
Epstein Barr Virus-transformed human B-lymphocytes (EBV-LCL).
Results. This method resulted in a labelling yield of 80% and a radiochemical
purity of >99%. SEC-HPLC analysis showed that during the
labelling the integrity of the antibody was maintained, although a small
increase in formation of aggregates was observed (