MATERIALS AND METHODS
We retrospectively analyzed patients with refractory or relapsed NB
treated at the GianninaGaslini Institute in Genoa (Italy) from 1996 to
2014. Those who had undergone treatment with
Th-131I-MIBG followed closely by ASCT after HD-MAT
with Busulfan and Melphalan after induction therapy were included in the
analysis, while those who had received Th-131I-MIBG
alone or in combination with a different schedule of HD-MAT were
excluded.
Figure 1 summarizes treatment schedules and disease response
evaluations.
All patients or their guardians signed a consent form allowing the use
of their data for clinical research purposes. We followed the procedures
in accordance with our institution’s ethical standards and according to
Italian guidelines. The Liguria Regional Ethics committee
(ComitatoEticoRegionale – Regione Liguria – IRCCS AOU San Martino –
IST) approved the collection for retrospective studies (360REG2014) of
data from medical records of patients admitted to our department.
All data were collected from our institutional database and
retrospectively analyzed. The main clinical characteristics of patients
and the types of treatment are summarized in Table 1 .
Criteria for diagnosis, staging and inclusion in the
study
The diagnosis of NB was established by histological examination of tumor
specimens or by bone marrow infiltration and/or elevated urinary
excretion of catecholamine metabolites. Disease extension on diagnosis
and response to treatment were evaluated by means of imaging
(ultrasonography/CT scan/MRI) of the primary tumor,123I-MIBG scintigraphy and study of bone marrow on at
least 2 aspirates and 2 trephine core biopsies. In the absence of MIBG
uptake, skeletal involvement was evaluated by means of technetium
99m-MDP scintigraphy or, more recently, 18F-FDG
PET/CT. However, in this specific case, the patient was not deemed
eligible for the study. All patients were staged according to INSS
criteria [10] . MYCN gene amplification was defined as a
copy number of 10 or more.
To be included in the study, patients had to have a poorly responding
HR-NB (MIBG-avid) after induction followed by further chemotherapy or,
alternatively, a residual HR-NB (MIBG-avid) after a salvage treatment
for relapse.
Treatments: Induction therapy, PBSC harvest and
Th-131I-MIBG
Induction therapy was administered according to the Italian NB97
protocol [14] before 2002 and the European SIOPEN NBAR-01
protocol [15] in subsequent years. Patients with refractory
disease after induction underwent salvage therapy with additional
courses of chemotherapy (ICE and/or TVD and/or TEMIRI ).
Peripheral blood stem cells (PBSC) were collected only if bone marrow
evaluation (at least 2 aspirates and 2 biopsies from different sites)
demonstrated complete remission; collection was performed after
stimulation with Granulocyte Colony-Stimulating Factor (G-CSF),
administered subcutaneously at a dosage of 10 μg/kg/day. The minimum
CD34+ cell-dose required in order to consider collection to be adequate
was 3 x 106/kg of the recipient’s weight (optimal dose
≥ 4 x 106/kg).
Before 131I-MIBG administration, patients underwent
thyroid blockade according to EANM procedure guidelines[16] and showed normal organ function and a value of WBC
> 2000/uL and of platelets > 75000/uL. They
were then admitted to the Nuclear Medicine Department, where
Th131I-MIBG was administered in a single intravenous
infusion over 2 hours (median dose 8.5mCi/Kg [IQR 6.5-12]). All
patients remained in radiation protective isolation for 5-7 days after131I-MIBG administration.
High-dose BU-Mel and autologous stem cell
transplantation
High-dose chemotherapy consisted of Busulfan, administered in 16 doses
from day -7 to day -3 before ASCT; administration was oral until 2011
(cumulative dose 16 mg/kg) and intravenous in the following years
(cumulative dose according to weight range: <9 Kg: 16
mg/Kg; 9-16 Kg: 19.2 mg/Kg; 16-23 Kg: 17.6 mg/Kg; 23-34 Kg: 15.2 mg/Kg;
>34 Kg: 12.8 mg/Kg ). Melphalan was administered on day -1
in a single intravenous dose of 140 mg/m2. On day 0,
patients underwent ASCT.
Disease response criteria and toxicity
Disease response was evaluated at the end of the induction phase, before
and after Th-131I-MIBG and HD-BuMel, at the end of
treatment and at the last follow-up examination.
Considering the disease response evaluated before
Th-131I-MIBG and HD-MAT BuMel, according to the
International Neuroblastoma Response Criteria (INRC) [10] ,
we defined 3 groups of response: i) good response (complete
response [CR], very good partial response [VGPR]); ii)partial response (PR); iii) poor response (no response
[NR], stable disease [SD] and progression of disease [PD]).
Toxicity was graded according to Common Terminology Criteria version
4.03 [11] , on considering mucosal, gastrointestinal,
pulmonary, hepatic and renal toxicities and veno-occlusive
disease/sinusoidal obstruction syndrome (VOD/SOS) according to the
Baltimore [12] and Seattle criteria [13] . Only
proven bacterial bloodstream infection or clinical sepsis and deep
mycosis were regarded as significant infective complications.
Engraftment was defined as the first of 3 consecutive days on which
neutrophil and platelet counts reached 500/mm3 and
50,000/mm3, respectively.
Statistical analysis
Descriptive statistics are reported in terms of absolute frequencies and
percentages for continuous data), and Pearson’s chi-square test was used
to compare proportions.
The overall survival (OS) probability and the cumulative risk of relapse
(CRR) were calculated by means of the Kaplan-Meier method, considering
time from diagnosis and from therapy (Th-131I-MIBG and
HD-MAT BuMel) to death from any cause (for OS) or to relapse (for CRR);
if neither event occurred, data were censored on the date of the last
follow-up examination. Differences among subgroups were assessed by
means of the Log Rank test. Cox modeling was adopted for multivariate
analysis, considering MYCN amplification, induction therapy, MIBG score
after induction, HD-CT with Thiotepa and disease status before
Th-131I-MIBG as variables.
Analyses were performed by means of STATA software (version 14.2, Stata
Corp., College Station, TX, USA). Two-tailed probabilities are reported;
a p-value of 0.05 was adopted to define nominal statistical
significance.