Seventy-one autologous HSCT and non-myeloablative
allogeneic transplantation (NST) performed in 62 patients
at Institute of Oncology between January 2000 and
December 2002 were analyzed. During this period, 57
autologous HSCT and 14 NST were performed. Eligibility
criteria were: 1) Age 17 to 60 years (upper age limit
was not applied to patients undergoing NST and in
patients with myeloma); 2) normal hepatic, cardiac and
pulmonary functions based on echocardiography, pulmonary
function tests, and clinical and laboratory evaluations;
3) ECOG performance status less than or equal to 2.
Patients were excluded if they had a positive serologic
test for HIV or a serious coexisting illness. A written
informed consent was obtained from all patients. Patients
with chemoresistant disease except in patients with
myeloma were excluded.
From January 2000 to December 2002, 55 patients
with relapsed/refractory Hodgkins disease, relapsed/
refractory/high risk non-Hodgkins lymphoma,
newly diagnosed/ relapsed/refractory multiple myeloma,
acute myeloid leukemia (AML) without a matched sibling
and relapsed testicular carcinoma underwent autologous
HSCT following high-dose chemotherapy. Double autologous
HSCT was performed in 2 patients with myeloma.
Except in myeloma, patients resistant to salvage chemotherapy
were considered ineligible for HSCT. Response
to HSCT was determined on day +100.
Peripheral blood stem cell mobilization and preparative
Mobilization regimen consisted of cyclophosphamide
(4.5 g/m2) and granulocyte colony stimulating factor (GCSF)
(10 µg/kg/d) combination in lymphoma patients.
In patients with multiple myeloma, AML and testicular
carcinoma, G-CSF (10 µg/kg/d) alone was used prior to stem cell collection. Cells were collected with 2 or 3
aphereses when neutrophil count reached >5000x109/l.
Target dose was at least 2x106 CD34+ cells or 5x108
mononuclear cells per kg body weight.
In lymphoma patients, high-dose sequential therapy
(Figure 1) consisting of etoposide (2 g/m2), mitoxantrone
(60 mg/m2) and melphalan (180 mg/m2) was used as the
conditioning regimen following successful salvage therapy.
High dose melphalan at a dose of 200 mg/m2 was
administered to patients with myeloma patients prior to hematopoietic stem cell (HSC) infusion. Conditioning regimen was carboplatin (AUC=25)/melphalan (180
mg/m2) in patients with testicular carcinoma and mitoxantrone
(60 mg/m2)/melphalan (180 mg/m2) in patients
Figure 1: High-dose sequential therapy conditioning regimen administered to patients with Hodgkins and non-Hodgkins lymphoma
All patients received antifungal prophylaxis with
fluconazole 200 mg per day. Acyclovir was given at a
dose of 250 mg/m2 twice daily in patients with positive
HSV serology. Broad-spectrum combined intravenous
antibiotics were initiated for fever greater than 38.3°C
in the setting of neutropenia. Myeloma patients received
intravenous immunoglobulin 400 mg/kg on day +4. GCSF
10 µg/kg/day was administered intravenously beginning
1 day after hematopoietic stem cell (HSC)
infusion and continued until engraftment. Platelet transfusions
were routinely administered to keep the platelet
count above 20x106/l. Red cell transfusions were administered
to maintain the hematocrit level over 27%.
In lymphoma patients, post-transplant maintenance
therapy included interferon-alpha administered subcutaneously
at 1-5x106 units, three times weekly starting
after day +100 when platelet count reached >100x109/l.
Interferon therapy was usually continued for 18 months
until disease progression or patient intolerance was
observed. In myeloma patients, thalidomide was started
at a dose of 200 mg/day as the maintenance therapy for
Non-myeloablative allogeneic transplantation
The indications for NST were relapse following
autologous transplantation in Hodgkins disease, non-Hodgkins lymphoma and multiple myeloma, metastatic
renal cell carcinoma, relapsed AML, resistant CLL, CML,
and large granular leukemia.
HLA tissue typing
In the Department of Basic Oncology, Immunogenetics
and Molecular Tissue Typing Laboratory, procedures
for histocompatibility were designated according to the
regulations/criteria of European Federation for Immunogenetics
(EFI) and/or American Society for Histocompatibility
and Immunogenetics (ASHI). Strict precautions
for contamination control were taken such as dedicated
material and equipment as well as separated pre- and
Donor samples were collected from both siblings
and parents when/if available. DNA was extracted from
peripheral blood mononuclear cells using commercial
DNA extraction kits as described by the manufacturer
(Qiagen GmbH. Hilden, Germany). Back up of each
sample was obtained by using phenol-chlorophorm
extraction method. Both quantitative and qualitative
evaluation of DNA samples were done by using Perkin-
Elmer U.V. spectrophotometer. HLA alleles were assigned
employing low resolution Sequence Specific Primers
(SSP) method (Pel-Freez Clinical Systems LLC. Brown
Deer, WI, U.S.A.). PCR setup and thermocycling conditions
were performed as described by the manufacturer
(M.J Research PTC-100. Watertown, MA, U.S.A.). Postamplification
procedure was 2% agarose gel electrophoresis
followed by photography and visual evaluation. Allele
assignments, and family tree studies were controlled at
least by 3 observers and checked by using software
provided by the manufacturer. Results and personal data
were recorded on confidential basis by password in protected files and computers [8-10].
Stem cell collection and conditioning therapy
All patients received unmanipulated HSC from
matched sibling donors. Donors were mobilized with GCSF at 10 µg/kg/d for administered 5 to 7 days. Cells
were collected by 2 to 3 aphereses performed on days 5-7 after initiation of G-CSF.
Conditioning regimen before infusion of allogeneic
stem cells mainly included immunosuppressive therapy
with fludarabine and single dose total body irradiation
(TBI). In patients with disease necessitating immediate cytoreductive therapy (patients no:1-4 and 14), cytarabine
was added to the conditioning regimen before emergence
of graft versus tumor effect. Two NST patients with
relapsed myeloma received melphalan only instead of
fludarabine/TBI regimen as they were already immunosuppressed
due to prior autologous HSCT. A patient with
renal cell carcinoma received cyclophosphamide/
fludarabine as the preparative regimen for NST.
Prophylaxis of Graft-versus-Host disease (GVHD)
GVHD prophylaxis consisted of cyclosporin-A (CsA)
administered as 24 hour infusion at 2.5 mg/kg and oral
mycophenolate mofetil (MMF) at 1500 mg/d starting
the day before allogeneic HSC infusion. CsA dose was
adjusted daily to maintain the target blood level between
475 and 525 ng/ml. CsA was switched to oral administration
at 5 mg/kg/d following engrafting and discontinuation
of TPN. CsA was slowly tapered according to
protocol schedule after day +60 and discontinued before
day +100. MMF was discontinued on day +180 if there
were no signs of GVHD.
All patients received antibacterial and antifungal
prophylaxis with levofloxacin 500 mg per day, ornidazole
200 mg twice daily and fluconazole 200 mg per day.
Acyclovir was given at 250 mg/m2 twice daily to patients
with positive HSV serology. Peripheral blood CMV
DNA titers were monitorized weekly by PCR assays.
Broad-spectrum intravenous antibiotics were initiated
when fever over 38.3°C was observed during neutropenia
and for documented infections. Prophylaxis against
Pneumocystis carinii included trimethoprim/sulfamethaxazole p.o. qd, thrice weekly, administered
during the period between engraftment and day +360.
Hematocrit and platelet counts were maintained with
irradiated blood products above >27% and >20x109/l,
respectively. G-CSF at 5 µg/kg/d was administered
between day +1 and time of engraftment.
Donor lymphocyte infusion (DLI)
DLIs were routinely performed on day +30 and day
+100 to achieve full chimerism. Then, decision for DLI
was made according to status of chimerism, disease
activity or presence of GVHD. DLI was infused at a
starting cell dose of 1x107 CD3+ cells/kg.
Analysis of chimerism
Chimerism was assessed by cytogenetic analysis
using fluorescent in situ hybridization (FISH) in sex
mismatched NSTs. PCR based cytogenetic analysis of
various number of tandem repeats (VNTR) was performed
in the setting of sex-mismatched transplants.
Ambulatory Nursing Care
Medical nurse coordinator monitored patients on outpatient
basis. She was responsible for administration of
blood products and routine intravenous medications,
routine care of central catheters, and education activities
for patient home care and self injections.
Dental care of the patients was carried out at the
Department of Oral Diagnosis in the Faculty of Dentistry
of Hacettepe University.
One social and one research coordinator organized
the maintenance of out-patient clinic, appointments and
financial reimbursement of patients with social security
civil public insurance services. Coordinators reported
all the patient data on a regular basis to EBMTR (CIC
code:292), ABMTR and IBMTR (Center code:589) every
Chronic GVHD and late effects of NST were followed
and recorded during periodic outpatient visits.
The primary end-point for this report is to determine
overall survival (OS) and disease-free survival (DFS).
OS was defined from the day of autologous HSCT until
death from any cause or latest contact with the patient.
DFS was defined as the time from the day of transplantation
until disease progression, death from any cause
or latest follow-up. Transplant-related mortality (TRM)
was the secondary end-point and was defined as death
from any cause except relapse during the first 100 days
of transplantation. Kaplan-Meier method was used to
calculate estimated DFS and OS .