Contributions
Abstract: PB1514
Type: Publication Only
Session title: Chronic myeloid leukemia - Clinical
Background
CML is the result of the chromosome translocation t(9;22), which leads to the fusion gene between ABL1 proto-oncogen and BCR gene [1]. The resulting product is an active tyrosine kinase leading to uncontrolled proliferation of different cell lineages and eventually to leukaemia.The crude annual incidence of CML in Europe ranges between 0.7–1.0/100,000 with a median age at diagnosis of 57-60 years [2]. WHO suggests no association with race or ethnicity [3]. Due to lack of reliable data for Low-and-middle-income countries (LMIC), numbers remain an estimation, but taking existing data for extrapolation, the global annual incidence would be above 100,000 [3]. Few studies on treatment outcome in LMIC suggest a lower response rate to TKI [4,5], despite following international treatment guidelines. Assessment of additional chromosomal aberrations (ACA) is no criterion according to the ELN staging recommendation, but one criterion for accelerated phase in the WHO recommendations. However, it is not routinely assessed in LMIC due to unavailability of karyotyping diagnostics, challenges in timely transportation of samples and costs.
Aims
To assess the possibility of obtaining high quality results in karyotyping of CML patients by using dry spot cards for sample collection and storage in a resource poor setting.
Methods
9 peripheral blood samples of newly diagnosed CML patients have been collected and stored between 1 and 5 months before analysed by array-CGH (aCGH). Storage and transportation of the samples were performed on dry spot cards. ELN and WHO criteria for CML phases were applied to all patients. aCGH analyses were carried out for all cases (4x180K microarray slides, Agilent Technologies, Santa Clara, CA). Images were analyzed using the DEVA Software v1.2.1 (Roche Nimblegen) and Nexus Copy Number 6.1 (Biodiscovery, Inc., El Segundo, CA). ACA were evaluated in each sample using BioDiscovery’s Fast Adaptive States Segmentation Technique (FASST2) algorithm. Targeted NGS sequencing was performed with the TruSeq DNA library Prep (Illumina, San Diego, USA) followed by a hybridization capture workflow (IDT, Coralville, USA) for enrichment of a myeloid 34 gene panel including a CNV spike-in panel.
Results
8 patients were in CP, 1 in AP (ELN criteria) and all 9 patients in AP (WHO criteria). All specimen for karyotyping were evaluable and 7/9 showed ACA, mainly small deletions which are cytogenetically cryptic and thus only detectable using aCGH. 2 cases showed larger chromosomal aberrations, one patient showed a trisomy 8, while another one showed a deletion 7q and gain of 8q, resulting for both in an unfavourable prognosis. Interestingly, no gene mutation was identified within the 34 analysed genes. The NGS CNV panel confirmed all larger chromosomal changes.
Conclusion
Applying ELN criteria for CML phases without karyotyping might lead to underdosing and eventually treatment failure. Our small cohort shows that 7 out of 8 patients diagnosed in CML-CP by ELN criteria, have ACA and need higher dosages of Imatinib or 2nd line TKI (CML-AP by WHO criteria). The results provide a likely explanation for the general poorer outcome of standard dose treatment in LMIC. Using dry spot cards for storage and transportation of blood samples is a simple to apply technique for clinicians and delivers DNA for standard molecular genetic diagnostics like information about chromosomal copy number changes as well as mutation analyses. These information allow further risk stratification of these patients. This approach can overcome current challenges of sample shipment and storage in LMIC.
Keyword(s): Chronic myeloid leukemia, Diagnosis, Gene array, Treatment
Abstract: PB1514
Type: Publication Only
Session title: Chronic myeloid leukemia - Clinical
Background
CML is the result of the chromosome translocation t(9;22), which leads to the fusion gene between ABL1 proto-oncogen and BCR gene [1]. The resulting product is an active tyrosine kinase leading to uncontrolled proliferation of different cell lineages and eventually to leukaemia.The crude annual incidence of CML in Europe ranges between 0.7–1.0/100,000 with a median age at diagnosis of 57-60 years [2]. WHO suggests no association with race or ethnicity [3]. Due to lack of reliable data for Low-and-middle-income countries (LMIC), numbers remain an estimation, but taking existing data for extrapolation, the global annual incidence would be above 100,000 [3]. Few studies on treatment outcome in LMIC suggest a lower response rate to TKI [4,5], despite following international treatment guidelines. Assessment of additional chromosomal aberrations (ACA) is no criterion according to the ELN staging recommendation, but one criterion for accelerated phase in the WHO recommendations. However, it is not routinely assessed in LMIC due to unavailability of karyotyping diagnostics, challenges in timely transportation of samples and costs.
Aims
To assess the possibility of obtaining high quality results in karyotyping of CML patients by using dry spot cards for sample collection and storage in a resource poor setting.
Methods
9 peripheral blood samples of newly diagnosed CML patients have been collected and stored between 1 and 5 months before analysed by array-CGH (aCGH). Storage and transportation of the samples were performed on dry spot cards. ELN and WHO criteria for CML phases were applied to all patients. aCGH analyses were carried out for all cases (4x180K microarray slides, Agilent Technologies, Santa Clara, CA). Images were analyzed using the DEVA Software v1.2.1 (Roche Nimblegen) and Nexus Copy Number 6.1 (Biodiscovery, Inc., El Segundo, CA). ACA were evaluated in each sample using BioDiscovery’s Fast Adaptive States Segmentation Technique (FASST2) algorithm. Targeted NGS sequencing was performed with the TruSeq DNA library Prep (Illumina, San Diego, USA) followed by a hybridization capture workflow (IDT, Coralville, USA) for enrichment of a myeloid 34 gene panel including a CNV spike-in panel.
Results
8 patients were in CP, 1 in AP (ELN criteria) and all 9 patients in AP (WHO criteria). All specimen for karyotyping were evaluable and 7/9 showed ACA, mainly small deletions which are cytogenetically cryptic and thus only detectable using aCGH. 2 cases showed larger chromosomal aberrations, one patient showed a trisomy 8, while another one showed a deletion 7q and gain of 8q, resulting for both in an unfavourable prognosis. Interestingly, no gene mutation was identified within the 34 analysed genes. The NGS CNV panel confirmed all larger chromosomal changes.
Conclusion
Applying ELN criteria for CML phases without karyotyping might lead to underdosing and eventually treatment failure. Our small cohort shows that 7 out of 8 patients diagnosed in CML-CP by ELN criteria, have ACA and need higher dosages of Imatinib or 2nd line TKI (CML-AP by WHO criteria). The results provide a likely explanation for the general poorer outcome of standard dose treatment in LMIC. Using dry spot cards for storage and transportation of blood samples is a simple to apply technique for clinicians and delivers DNA for standard molecular genetic diagnostics like information about chromosomal copy number changes as well as mutation analyses. These information allow further risk stratification of these patients. This approach can overcome current challenges of sample shipment and storage in LMIC.
Keyword(s): Chronic myeloid leukemia, Diagnosis, Gene array, Treatment