Clinical Utility of Fluorescence in-situ Hybridization Profile Test in Detecting Genetic Aberrations in Acute Leukemia

Suk Ran Kim; Hee-Jin Kim; Sun-Hee Kim

doi:10.3343/kjlm.2009.29.5.371

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Kim, Kim, and Kim: Clinical Utility of Fluorescence in-situ Hybridization Profile Test in Detecting Genetic Aberrations in Acute Leukemia

Original Article

Korean J Leg Med 2009;29(5):371-378.

Published online: 14 January 2009

DOI: https://doi.org/10.3343/kjlm.2009.29.5.371

Clinical Utility of Fluorescence in-situ Hybridization Profile Test in Detecting Genetic Aberrations in Acute Leukemia

Suk Ran Kim, M.D., Hee-Jin Kim, M.D., Sun-Hee Kim, M.D.

Department of Laboratory Medicine & Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

Corresponding author: Sun-Hee Kim, M.D. Department of Laboratory Medicine & Genetics, Samsung Medical Center, Sungkyunkwan University School of Medicine, 50 Irwon-dong, Gangnam-gu, Seoul 135-710, Korea Tel: +82-2-3410-2704, Fax: +82-2-3410-2719 E-mail: sunnyhk@skku.edu

Revised 24 April 20095 September 2009 Accepted 10 September 2009

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background:

Cytogenetic abnormalities are one of the most reliable prognostic factors in acute leukemia. Combination of conventional chromosome analysis (CCA) and FISH provides higher sensitivity in detecting these genetic abnormalities, and it is effective to apply several FISH probes as a profile test. The objective of this study was to investigate the utility of FISH profile analyses in the initial diagnosis of acute leukemia.

Methods:

Two hundred and forty one de novo acute leukemia patients diagnosed from January, 2002 to November, 2007 were included. For acute lymphoblastic leukemia profile test, FISH probes for BCR/ABL, TEL/AML1, MLL gene rearrangement and CDKN2A deletion were used. For acute myeloid leukemia profile test, probes for AML1/ETO, MLL and CBFβ gene rearrangement were used. The results of CCA and FISH profile tests were collected, and the positive rates were compared.

Results:

ALL FISH profile tests revealed additional genetic aberrations not detected by chromosome analysis in 48.6% (67/138) of cases, including those with normal karyotypes or no mitotic cells (37%, 51/138). Among these 51 cases, TEL/AML1 abnormalities were detected in 44.3%, followed by the abnormal CDKN2A signal (24.6%) and hyperdiploidy (18.0%). AML FISH profile tests revealed additional genetic abnormalities in 7.8% (8/103) of cases.

Conclusions:

FISH analysis as a profile test detected additional genetic aberrations in a significant proportion of acute leukemia, and was effective especially in detecting cryptic translocations, submicroscopic deletions and complex karyotypes. Our study supports the need to incorporate FISH profile test at initial work up in acute leukemia.

Keywords: Acute leukemia, Cytogenetic analysis, Fluorescence in-situ hybridization

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Fig. 1.

Karyotype by conventional chromosome analysis (A) and the FISH patterns showing CBFβ break-apart signals (B) shown in case 1.

Table 1.

List of the probes used for fluorescence in-situ hybridization analysis in this study

Aberration	Localization	Gene/locus	Probe type
t(9;22)	9q34/22q11.2	ABL/BCR	Dual fusion translocation probe
t(12;21)	12p13/21p22	TEL/AML1	ES translocation probe
t(11q23;v)	11q23	MLL	Break apart probe
p16/CEP 9	9p21/9p11-q11	CNKN2A/alpha satellite	Enumeration probe
t(8;21)	8q22/21q22	AML1/ETO	Dual fusion translocation probe
inv(16)	16q22	CBFβ	Break apart probe

All aberration was detected by dual color set and Abbott Vysis.

Abbreviations: ES, extra signal; CEP 9, chromosome 9 enumeration probe.

Table 2.

Characteristics of acute leukemia cases where fluorescence in-situ hybridization profile test were applied

ALL FISH profile test were applied				AML FISH profile test were applied
Parameter/immunophenotype	B-cell type (109)∗	T-cell type (14)∗	Biphenotype/mixed lineage (15)∗	Parameter/classification	AML with recurrent genetic abnormalities (29)∗	AML, not otherwise specified (65)∗	Others (AML with MD, etc.) (9)∗
Age (yr)				Age (yr)
Children (age<18)				Children (age<18)	14	10 (M7:4)	1
<1	3	0	0	Adult
1-10	61	2	5	<55	9	36	6
>10	24	3	2	>55	6	19	2
Adult				Gender
<65	21	9	8	Male	16	39	7
Gender				Female	13	26	2
Male	53	9	8
Female	56	5	7

^∗ () Number of patients with the specific immunophenotypic classification.

Abbreviations: AML with MD, acute myeloid leukemia with multilineage dysplasia; M7, acute megakaryoblastic leuekemia.

Table 3.

Frequencies of the genetic abnormalities according to the detection methods, conventional cytogenetic analysis (CCA) and FISH profile test

Applied test	Phenotype	N of patients	I			II
Applied test	Phenotype	N of patients	Category 1∗	Category 2^†	Category 3^‡	Category 4^§	Category 5^‖
ALL profile test	Precursor B-cell ALL	109	15	32	9	14	39
	Precursor T-cell ALL	14	6	1	1	1	5
	Acute leukemias of ambiguous lineage	15	1	5	1	1	7
	Total (%)	138	22 (15.9%)	38 (27.5%)	11 (8.0%)	16 (11.6%)	51 (37.0%)
AML profile test	AML with AML1/ETO	15		14			1
	AML with MLL abnormalities	9		8			1
	AML with CBFβ/MYH11	5		3			2
	Minimally differentiated	2	1	1
	AML without maturation	25	18	1	5		1
	AML with maturation	17	12	1	2		2
	Acute myelomonocytic leukemia	4	3	1
	Acute monoblastic/monocytic leukemia	11	7	1	2		1
	Acute megakaryoblastic leukemia	4		2	2
	Acute panmyelosis and myelofibrosis	2	1		1
	AML with multilineage dysplasia	8	6	2
	Biphenotypic acute leukemia	1			1
	Total	103	48 (46.6%)	34 (33.0%)	13 (12.6%)	0	8 (7.8%)

Categories: 1∗, CCA: normal or ‘no mitotic cells’/FISH profile test: normal; 2^†, Compatible results of cytogenetic abnormalities between the two detection methods; 3^‡, CCA: abnormal / FISH profile test: normal; 4^§, Additional cytogenetic abnormalities were detected only by FISH profile test; 5^‖, CCA: normal or ‘no mitotic cells’/FISH profile test: abnormal.

Table 4.

Frequencies of the abnormalities identified by ALL FISH profile in 51 acute leukemia cases with normal karyotypes or without cytogenetic results (category 5)

Analyzed gene	N of abnormal cases (%)	Rearrangement	Deletion	Amplification	Extra copies related to aneuploidy
CDKN2A(p16)	15 (24.6)		14		1
TEL/AML1	27 (44.3)	13	10(TEL), 1(AML1)	3 (AML1)	7(AML1)
BCR/ABL	5 (8.2)	4			1(ABL)
MLL	3 (4.9)	3
Hyperdiploidy	11 (18.0)
Total	61∗ (100)

^∗ Number of patients harboring 2 abnormalities, 8; number of patients harboring 3 abnormalities, 1.

Table 5.

Summary of the results of conventional chromosome analysis and FISH profile test of 4 cases of “AML with recurrent genetic abnormalities” and category 5

Case No.	Sex	Age	Initial diagnosis	Chromosome analysis	FISH result	Final diagnosis
1	M	55	AML with maturation	45,X,-Y, t(2;21;8)(q37;q22;q22)[14]/46,XY[6]	AML1/ETO	AML with AML1/ETO
2	F	1 month	Acute monoblastic leukemia	46,XX[20]	Break-apart MLL signals (31.5%)	AML with 11q23(MLL) abnormalities
3	M	6	Acute myelomonocytic leukemia	47,XY,+22[3]/46,XY[1]	Break-apart CBFβ signals (83%)	AML with CBFβ/MYH11
4	M	15	Acute myelomonocytic leukemia	46,XY[20]	Break-apart CBFβ signals (89%)	AML with CBFβ/MYH11

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