INCOMPLETE IGH GENE REARRANGEMENTS (DH-JH) AS A MINIMAL RESIDUAL DISEASE TARGET IN ADULT ACUTE LYMPHOBLASTIC LEUKEMIA PATIENTS
(Abstract release date: 05/19/16)
EHA Library. Sedlackova L. 06/09/16; 134494; PB1594
Ms. Lucie Sedlackova
Contributions
Contributions
Abstract
Abstract: PB1594
Type: Publication Only
Background
Monitoring of minimal residual disease (MRD) is an important prognostic tool in acute lymphoblastic leukemia (ALL) patients. Currently, one of the most widely used techniques to detect and quantify MRD is allele-specific real-time polymerase chain reaction (qPCR) based on clonal rearrangement of immunoglobulin heavy chain (IGH) or T-cell receptor genes. These molecular targets can be detected in approximately 90% of adult B-cell precursor ALL at initial diagnosis. In patients, where it is not possible to detect a clonal complete IGH rearrangement, additional MRD targets, such as immunoglobulin light chain (IGL) or incomplete IGH (DH-JH) rearrangements, could be also used. The first step in the recombination process of the IGH locus is a rearrangement of DH and JH segments which represents one of the earliest events in B-cell development. Incomplete DH-JH joints were observed in 20 – 25% of B-cell precursor ALLs. Therefore, in adult ALL patients without clonal complete IGH rearrangement, incomplete DH-JH rearrangements could be potentially used as MRD target.
Aims
The goal of our work was the application of clonal incomplete DH-JH rearrangements as MRD targets in routine laboratory practice.
Methods
Since 2010, we have performed clonal complete IGH rearrangement analysis in 45 patients with ALL suspicion in effort to obtain suitable MRD target. In 18 patients, where it was not possible to detect clonal complete IGH marker, screening by the multiplex PCR using BIOMED-2 primer sets for the detection of clonal DH-JH rearrangement was made as the next step. After sequencing of the clonal DH-JH rearrangement, allele-specific oligonucleotides were designed for each MRD target on the basis of the sequence data of the junctional regions. Tests for MRD were conducted by qPCR using TaqMan technology (Rotor-Gene Q, Qiagen).
Results
Clonal incomplete DH-JH rearrangements were detected and sequenced in 11 out of 18 ALL patients at initial diagnosis. Two out of 11 patients had another available molecular target for MRD monitoring (BCR/ABL, MLL/ENL) and for additional 3 patients MRD analysis was not requested. The qPCR assays were developed for 6 patients who were subsequently monitored for MRD. From February 2011 to January 2016 we examined 59 samples from bone marrow (n = 57) or peripheral blood (n = 2). The assay detection sensitivity achieved the threshold of 10-4 to 10-5 (1 leukemic cell in 10 000 cells to 1 leukemic cell in 100 000 cells). The MRD levels of residual leukemic cells correlated with clinical outcome. The clonal DH-JH markers found in these patients and used for the MRD analysis were the only option for residual leukemic cells monitoring with sensitivity of at least four orders of magnitude.
Conclusion
Clonal incomplete DH-JH gene rearrangement targets are applicable for routine MRD testing of ALL patients as a reliable sensitive method for MRD assessment and should be considered as a complementary target for PCR-based clonality assessment and MRD monitoring.
Session topic: E-poster
Keyword(s): ALL, IgH rearrangment, MRD, Quantitative RT-PCR
Type: Publication Only
Background
Monitoring of minimal residual disease (MRD) is an important prognostic tool in acute lymphoblastic leukemia (ALL) patients. Currently, one of the most widely used techniques to detect and quantify MRD is allele-specific real-time polymerase chain reaction (qPCR) based on clonal rearrangement of immunoglobulin heavy chain (IGH) or T-cell receptor genes. These molecular targets can be detected in approximately 90% of adult B-cell precursor ALL at initial diagnosis. In patients, where it is not possible to detect a clonal complete IGH rearrangement, additional MRD targets, such as immunoglobulin light chain (IGL) or incomplete IGH (DH-JH) rearrangements, could be also used. The first step in the recombination process of the IGH locus is a rearrangement of DH and JH segments which represents one of the earliest events in B-cell development. Incomplete DH-JH joints were observed in 20 – 25% of B-cell precursor ALLs. Therefore, in adult ALL patients without clonal complete IGH rearrangement, incomplete DH-JH rearrangements could be potentially used as MRD target.
Aims
The goal of our work was the application of clonal incomplete DH-JH rearrangements as MRD targets in routine laboratory practice.
Methods
Since 2010, we have performed clonal complete IGH rearrangement analysis in 45 patients with ALL suspicion in effort to obtain suitable MRD target. In 18 patients, where it was not possible to detect clonal complete IGH marker, screening by the multiplex PCR using BIOMED-2 primer sets for the detection of clonal DH-JH rearrangement was made as the next step. After sequencing of the clonal DH-JH rearrangement, allele-specific oligonucleotides were designed for each MRD target on the basis of the sequence data of the junctional regions. Tests for MRD were conducted by qPCR using TaqMan technology (Rotor-Gene Q, Qiagen).
Results
Clonal incomplete DH-JH rearrangements were detected and sequenced in 11 out of 18 ALL patients at initial diagnosis. Two out of 11 patients had another available molecular target for MRD monitoring (BCR/ABL, MLL/ENL) and for additional 3 patients MRD analysis was not requested. The qPCR assays were developed for 6 patients who were subsequently monitored for MRD. From February 2011 to January 2016 we examined 59 samples from bone marrow (n = 57) or peripheral blood (n = 2). The assay detection sensitivity achieved the threshold of 10-4 to 10-5 (1 leukemic cell in 10 000 cells to 1 leukemic cell in 100 000 cells). The MRD levels of residual leukemic cells correlated with clinical outcome. The clonal DH-JH markers found in these patients and used for the MRD analysis were the only option for residual leukemic cells monitoring with sensitivity of at least four orders of magnitude.
Conclusion
Clonal incomplete DH-JH gene rearrangement targets are applicable for routine MRD testing of ALL patients as a reliable sensitive method for MRD assessment and should be considered as a complementary target for PCR-based clonality assessment and MRD monitoring.
Session topic: E-poster
Keyword(s): ALL, IgH rearrangment, MRD, Quantitative RT-PCR
Abstract: PB1594
Type: Publication Only
Background
Monitoring of minimal residual disease (MRD) is an important prognostic tool in acute lymphoblastic leukemia (ALL) patients. Currently, one of the most widely used techniques to detect and quantify MRD is allele-specific real-time polymerase chain reaction (qPCR) based on clonal rearrangement of immunoglobulin heavy chain (IGH) or T-cell receptor genes. These molecular targets can be detected in approximately 90% of adult B-cell precursor ALL at initial diagnosis. In patients, where it is not possible to detect a clonal complete IGH rearrangement, additional MRD targets, such as immunoglobulin light chain (IGL) or incomplete IGH (DH-JH) rearrangements, could be also used. The first step in the recombination process of the IGH locus is a rearrangement of DH and JH segments which represents one of the earliest events in B-cell development. Incomplete DH-JH joints were observed in 20 – 25% of B-cell precursor ALLs. Therefore, in adult ALL patients without clonal complete IGH rearrangement, incomplete DH-JH rearrangements could be potentially used as MRD target.
Aims
The goal of our work was the application of clonal incomplete DH-JH rearrangements as MRD targets in routine laboratory practice.
Methods
Since 2010, we have performed clonal complete IGH rearrangement analysis in 45 patients with ALL suspicion in effort to obtain suitable MRD target. In 18 patients, where it was not possible to detect clonal complete IGH marker, screening by the multiplex PCR using BIOMED-2 primer sets for the detection of clonal DH-JH rearrangement was made as the next step. After sequencing of the clonal DH-JH rearrangement, allele-specific oligonucleotides were designed for each MRD target on the basis of the sequence data of the junctional regions. Tests for MRD were conducted by qPCR using TaqMan technology (Rotor-Gene Q, Qiagen).
Results
Clonal incomplete DH-JH rearrangements were detected and sequenced in 11 out of 18 ALL patients at initial diagnosis. Two out of 11 patients had another available molecular target for MRD monitoring (BCR/ABL, MLL/ENL) and for additional 3 patients MRD analysis was not requested. The qPCR assays were developed for 6 patients who were subsequently monitored for MRD. From February 2011 to January 2016 we examined 59 samples from bone marrow (n = 57) or peripheral blood (n = 2). The assay detection sensitivity achieved the threshold of 10-4 to 10-5 (1 leukemic cell in 10 000 cells to 1 leukemic cell in 100 000 cells). The MRD levels of residual leukemic cells correlated with clinical outcome. The clonal DH-JH markers found in these patients and used for the MRD analysis were the only option for residual leukemic cells monitoring with sensitivity of at least four orders of magnitude.
Conclusion
Clonal incomplete DH-JH gene rearrangement targets are applicable for routine MRD testing of ALL patients as a reliable sensitive method for MRD assessment and should be considered as a complementary target for PCR-based clonality assessment and MRD monitoring.
Session topic: E-poster
Keyword(s): ALL, IgH rearrangment, MRD, Quantitative RT-PCR
Type: Publication Only
Background
Monitoring of minimal residual disease (MRD) is an important prognostic tool in acute lymphoblastic leukemia (ALL) patients. Currently, one of the most widely used techniques to detect and quantify MRD is allele-specific real-time polymerase chain reaction (qPCR) based on clonal rearrangement of immunoglobulin heavy chain (IGH) or T-cell receptor genes. These molecular targets can be detected in approximately 90% of adult B-cell precursor ALL at initial diagnosis. In patients, where it is not possible to detect a clonal complete IGH rearrangement, additional MRD targets, such as immunoglobulin light chain (IGL) or incomplete IGH (DH-JH) rearrangements, could be also used. The first step in the recombination process of the IGH locus is a rearrangement of DH and JH segments which represents one of the earliest events in B-cell development. Incomplete DH-JH joints were observed in 20 – 25% of B-cell precursor ALLs. Therefore, in adult ALL patients without clonal complete IGH rearrangement, incomplete DH-JH rearrangements could be potentially used as MRD target.
Aims
The goal of our work was the application of clonal incomplete DH-JH rearrangements as MRD targets in routine laboratory practice.
Methods
Since 2010, we have performed clonal complete IGH rearrangement analysis in 45 patients with ALL suspicion in effort to obtain suitable MRD target. In 18 patients, where it was not possible to detect clonal complete IGH marker, screening by the multiplex PCR using BIOMED-2 primer sets for the detection of clonal DH-JH rearrangement was made as the next step. After sequencing of the clonal DH-JH rearrangement, allele-specific oligonucleotides were designed for each MRD target on the basis of the sequence data of the junctional regions. Tests for MRD were conducted by qPCR using TaqMan technology (Rotor-Gene Q, Qiagen).
Results
Clonal incomplete DH-JH rearrangements were detected and sequenced in 11 out of 18 ALL patients at initial diagnosis. Two out of 11 patients had another available molecular target for MRD monitoring (BCR/ABL, MLL/ENL) and for additional 3 patients MRD analysis was not requested. The qPCR assays were developed for 6 patients who were subsequently monitored for MRD. From February 2011 to January 2016 we examined 59 samples from bone marrow (n = 57) or peripheral blood (n = 2). The assay detection sensitivity achieved the threshold of 10-4 to 10-5 (1 leukemic cell in 10 000 cells to 1 leukemic cell in 100 000 cells). The MRD levels of residual leukemic cells correlated with clinical outcome. The clonal DH-JH markers found in these patients and used for the MRD analysis were the only option for residual leukemic cells monitoring with sensitivity of at least four orders of magnitude.
Conclusion
Clonal incomplete DH-JH gene rearrangement targets are applicable for routine MRD testing of ALL patients as a reliable sensitive method for MRD assessment and should be considered as a complementary target for PCR-based clonality assessment and MRD monitoring.
Session topic: E-poster
Keyword(s): ALL, IgH rearrangment, MRD, Quantitative RT-PCR
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