title: Evaluation of multiplex ligation dependent probe amplification (MLPA) for identification of acute lymphoblastic leukemia with an intrachromosomal amplification of chromosome 21 (iAMP21) in a Brazilian population.
abstract: BACKGROUND
  An intrachromosomal amplification of chromosome 21 (iAMP21) defines a unique subgroup of B-cell precursor acute lymphoblastic leukemia (BCP-ALL). The finding of three or more extra copies of the RUNX1 gene by fluorescence in situ hybridization (FISH) is internationally used to define an iAMP21. Genomic profiling of chromosome 21 has been suggested for assisting diagnostic case identification. Due to limitations of comparative genomic hybridization, in terms of a routine application as first line-screening tests we evaluated the multiplex ligation-dependent probe amplification (MLPA) SALSA P327_A1 and P327_B1 probe sets for detecting chromosome 21 copy number alterations in Brazilian childhood BCP-ALL.
  
  
  RESULTS
  In 74 out of 368 patients gain of genetic material was detected. For data confirmation RUNX1 directed FISH was performed. Cells with ≥5 RUNX1 signals (n = 9) were considered as "true iAMP21" while <5 RUNX1 signals (n = 41) were counted as evidence for additional copies of intact chromosomes 21. All patients with an iAMP21 had high MLPA peak ratios (≥1.8), while the majority of patients with <5 RUNX1 presented low MLPA peak ratios (<1.8). Observed differences gained statistical strength by comparing probes located within the common region of amplification. Next, a principal component analysis was performed in order to illustrate distribution of cases according to their MLPA peak profile in two dimensions. Cases with an iAMP21 mostly clustered together, however additional cases with <5 RUNX1 signals or no available FISH data located in proximity.
  
  
  CONCLUSIONS
  MLPA qualified as a high throughput technique that could be employed in future studies for a critical comparison with data obtained by FISH, especially in cases where metaphase nuclei are not available. Taking submicroscopic aberrations into account examined by MLPA, cases exhibiting an "iAMP21 like" peak ratio profile but <5 RUNX1 signals should be considered as candidates for this chromosomal abnormality.
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Fuka
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Tállita M
authors.initials: TM
authors.lastname: Farias-Vieira
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Leticia
authors.initials: L
authors.lastname: Hummel
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Caroline B
authors.initials: CB
authors.lastname: Blunck
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Júlio C
authors.initials: JC
authors.lastname: Santoro
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Eugênia
authors.initials: E
authors.lastname: Terra-Granado
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Thayana Conceição
authors.initials: TC
authors.lastname: Barbosa
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Mariana
authors.initials: M
authors.lastname: Emerenciano
authors.affiliation: Pediatric Hematology-Oncology Program, Research Center, Instituto Nacional de Câncer (INCA), Rua André Cavalcanti, 37, Rio de Janeiro, RJ 20231-050 Brazil.
authors.forename: Maria S
authors.initials: MS
authors.lastname: Pombo-de-Oliveira
keywords: B-cell precursor acute lymphoblastic leukemia (BCP-ALL); Fluorescence in situ hybridization (FISH); Intrachromosomal amplification of chromosome 21 (iAMP21); Multiplex ligation dependent probe amplification (MLPA)