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Implementation of the standard strategy for identification of Ig/TCR targets for minimal residual disease diagnostics in B-cell precursor ALL pediatric patients: Polish experience.

Dawidowska M, Jółkowska J, Szczepański T, Derwich K, Wachowiak J, Witt M - Arch. Immunol. Ther. Exp. (Warsz.) (2008)

Bottom Line: The U-Gauss test was used for statistical analysis of the Ig/TCR rearrangement pattern in Polish patients compared with relevant data on other nationalities.Statistically relevant differences were only found between the incidence of DH-JH in Polish (9%) and Dutch patients (24%; p<0.05) and Polish and Italian patients (19%; p<0.05), VH-JH in Polish (74%) and Chilean patients (100%; p<0.05), and TCRG in Polish (50%) and Brazilian patients (69%; p<0.05).The convergence of Ig/TCR patterns in Polish and European patients indicates that the strategy for Ig/TCR target identification based on standard primers and protocols might be directly used for the construction of Polish standards and recommendations for MRD diagnostics.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Clinical Genetics, Institute of Human Genetics, Polish Academy of Sciences, Strzeszyńska 32, 60-479 Poznań, Poland. ma.dawidowska@wp.pl

ABSTRACT

Introduction: Minimal residual disease (MRD), detected based on immunoglobulin and T-cell receptor (Ig/TCR) gene rearrangements as markers of residual leukemic cells, is currently the most reliable prognostic factor in acute lymphoblastic leukemia (ALL). A feasibility study is presented of the standard strategy for the identification of Ig/TCR targets for MRD diagnostics in Polish ALL patients by identifying Ig/TCR gene rearrangement pattern using standard primer sets and protocols.

Materials and methods: The PCR-heteroduplex approach based on BIOMED-1 and BIOMED-2 protocols (recommended as the European standard) was used to detect IGH, IGK-Kde, TCRD, TCRG, and TCRB rearrangements in 58 Polish B-cell precursor ALL patients. Sequencing and homology analysis between the obtained and germline Ig/TCR sequences enabled identification of the rearrangements. The U-Gauss test was used for statistical analysis of the Ig/TCR rearrangement pattern in Polish patients compared with relevant data on other nationalities.

Results: The following pattern was identified: IGH: 83% (VH-JH: 74%, DH-JH: 9%), IGK-Kde: 41%, TCRD: 78% (incomplete TCRD: 55%, Vdelta2-Ddelta3: 45%, Ddelta2-Ddelta3: 21%, Vdelta2-Jalpha: 35%), TCRG: 50%, and TCRB: 13%. Considerable convergence of the Ig/TCR pattern in Polish patients and those of other nationalities (mainly West Europeans) was demonstrated. Statistically relevant differences were only found between the incidence of DH-JH in Polish (9%) and Dutch patients (24%; p<0.05) and Polish and Italian patients (19%; p<0.05), VH-JH in Polish (74%) and Chilean patients (100%; p<0.05), and TCRG in Polish (50%) and Brazilian patients (69%; p<0.05).

Conclusions: The convergence of Ig/TCR patterns in Polish and European patients indicates that the strategy for Ig/TCR target identification based on standard primers and protocols might be directly used for the construction of Polish standards and recommendations for MRD diagnostics.

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Distribution of the number of rearrangements per patient detected in a total group of 58 patients (in a singleplex PCR approach) and in a subgroup of 23 patients (in a combined singleplex and multiplex PCR approach).
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Fig1: Distribution of the number of rearrangements per patient detected in a total group of 58 patients (in a singleplex PCR approach) and in a subgroup of 23 patients (in a combined singleplex and multiplex PCR approach).

Mentions: In the singleplex PCR approach, at least one clonal rearrangement was detected in 97% (56/58) of the BCP-ALL patients and at least two in 86% (50/58) of the patients. Insufficient quality of the DNA samples was excluded by amplification of the 100-, 200-, 400-, and 600-bp DNA fragments in multiplex PCR (BIOMED-2 Control Gene tube) in the case of the two patients in whom no PCR products were detected [30]. The number of rearrangements detected per patient ranged from 0 to 8, both cases occurring with a frequency of 3% (2/58), with 2 rearrangements as the most frequent number of rearrangements/patient (mode) occurring with a frequency of 24% (14/58). The mean number of rearrangements/patient was 3.6. In the case of the 23 patients (singleplex-multiplex approach), at least one clonal rearrangement was detected in 100% of the patients and at least two in 91% (21/23) of the patients. The number of rearrangements/patient ranged from 1 (2/23, 9%) to 8 (1/23, 4%), with a mode of rearrangements of 2 (6/23, 26%) and a mean of 4.2. The distributions of the number of rearrangements/patient detected in the groups of 58 and 23 patients are presented in Fig. 1.Fig. 1


Implementation of the standard strategy for identification of Ig/TCR targets for minimal residual disease diagnostics in B-cell precursor ALL pediatric patients: Polish experience.

Dawidowska M, Jółkowska J, Szczepański T, Derwich K, Wachowiak J, Witt M - Arch. Immunol. Ther. Exp. (Warsz.) (2008)

Distribution of the number of rearrangements per patient detected in a total group of 58 patients (in a singleplex PCR approach) and in a subgroup of 23 patients (in a combined singleplex and multiplex PCR approach).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2805919&req=5

Fig1: Distribution of the number of rearrangements per patient detected in a total group of 58 patients (in a singleplex PCR approach) and in a subgroup of 23 patients (in a combined singleplex and multiplex PCR approach).
Mentions: In the singleplex PCR approach, at least one clonal rearrangement was detected in 97% (56/58) of the BCP-ALL patients and at least two in 86% (50/58) of the patients. Insufficient quality of the DNA samples was excluded by amplification of the 100-, 200-, 400-, and 600-bp DNA fragments in multiplex PCR (BIOMED-2 Control Gene tube) in the case of the two patients in whom no PCR products were detected [30]. The number of rearrangements detected per patient ranged from 0 to 8, both cases occurring with a frequency of 3% (2/58), with 2 rearrangements as the most frequent number of rearrangements/patient (mode) occurring with a frequency of 24% (14/58). The mean number of rearrangements/patient was 3.6. In the case of the 23 patients (singleplex-multiplex approach), at least one clonal rearrangement was detected in 100% of the patients and at least two in 91% (21/23) of the patients. The number of rearrangements/patient ranged from 1 (2/23, 9%) to 8 (1/23, 4%), with a mode of rearrangements of 2 (6/23, 26%) and a mean of 4.2. The distributions of the number of rearrangements/patient detected in the groups of 58 and 23 patients are presented in Fig. 1.Fig. 1

Bottom Line: The U-Gauss test was used for statistical analysis of the Ig/TCR rearrangement pattern in Polish patients compared with relevant data on other nationalities.Statistically relevant differences were only found between the incidence of DH-JH in Polish (9%) and Dutch patients (24%; p<0.05) and Polish and Italian patients (19%; p<0.05), VH-JH in Polish (74%) and Chilean patients (100%; p<0.05), and TCRG in Polish (50%) and Brazilian patients (69%; p<0.05).The convergence of Ig/TCR patterns in Polish and European patients indicates that the strategy for Ig/TCR target identification based on standard primers and protocols might be directly used for the construction of Polish standards and recommendations for MRD diagnostics.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Clinical Genetics, Institute of Human Genetics, Polish Academy of Sciences, Strzeszyńska 32, 60-479 Poznań, Poland. ma.dawidowska@wp.pl

ABSTRACT

Introduction: Minimal residual disease (MRD), detected based on immunoglobulin and T-cell receptor (Ig/TCR) gene rearrangements as markers of residual leukemic cells, is currently the most reliable prognostic factor in acute lymphoblastic leukemia (ALL). A feasibility study is presented of the standard strategy for the identification of Ig/TCR targets for MRD diagnostics in Polish ALL patients by identifying Ig/TCR gene rearrangement pattern using standard primer sets and protocols.

Materials and methods: The PCR-heteroduplex approach based on BIOMED-1 and BIOMED-2 protocols (recommended as the European standard) was used to detect IGH, IGK-Kde, TCRD, TCRG, and TCRB rearrangements in 58 Polish B-cell precursor ALL patients. Sequencing and homology analysis between the obtained and germline Ig/TCR sequences enabled identification of the rearrangements. The U-Gauss test was used for statistical analysis of the Ig/TCR rearrangement pattern in Polish patients compared with relevant data on other nationalities.

Results: The following pattern was identified: IGH: 83% (VH-JH: 74%, DH-JH: 9%), IGK-Kde: 41%, TCRD: 78% (incomplete TCRD: 55%, Vdelta2-Ddelta3: 45%, Ddelta2-Ddelta3: 21%, Vdelta2-Jalpha: 35%), TCRG: 50%, and TCRB: 13%. Considerable convergence of the Ig/TCR pattern in Polish patients and those of other nationalities (mainly West Europeans) was demonstrated. Statistically relevant differences were only found between the incidence of DH-JH in Polish (9%) and Dutch patients (24%; p<0.05) and Polish and Italian patients (19%; p<0.05), VH-JH in Polish (74%) and Chilean patients (100%; p<0.05), and TCRG in Polish (50%) and Brazilian patients (69%; p<0.05).

Conclusions: The convergence of Ig/TCR patterns in Polish and European patients indicates that the strategy for Ig/TCR target identification based on standard primers and protocols might be directly used for the construction of Polish standards and recommendations for MRD diagnostics.

Show MeSH
Related in: MedlinePlus