Cytogentic Services in Leukaemia - Myeloid Malignancies Flashcards
What is the purpose of cytogenetic testing for myeloid malignancies?
1) . DIAGNOSIS - t(15;17) in Acute Promyelocytic Leukaemia (APML).
2) . PROGNOSIS - inv(16) in AML is good prognosis, but a complex karyotype in AML is poor.
3) . MONITORING - response to treatment, remission/relapse statues, loss or return of abnormalities seen at diagnosis, post BMT look for recipient/donor cells.
4) . TRIAL REQUIREMENTS - may ask for particular levels of analysis.
Discuss CML
- CML is Chronic Myeloid Leukaemia.
- The acquired chromosome translocation t(9;22)(q34;q11) is an example which leads to a new fusion protein BCR-ABL1.
- Can detect this abnormality by karyotype, FISH, PCR.
- ABL1 on chromosome 9, BCR on chromosome 22.
- BCR-ABL1 gene fusion formed on the the derivative chromosome 22 (Philadelphia chromosome).
- The reciprocal ABL1-BCR gene fusion is formed on the derivative chromosome 9.
- Can have variants.
What chromosome is ABL1 on?
- ABL1 is on chromosome 9
What chromosome is BCR on?
- BCR is on chromosome 22
What translocations leads to the formation of the BCR-ABL1 fusion protein? What chromosome is this formed on?
- BCR-ABL1 gene fusion formed on the the derivative chromosome 22 (Philadelphia chromosome).
- The reciprocal ABL1-BCR gene fusion is formed on the derivative chromosome 9.
What fusion protein will always be found in CML?
- BCR-ABL1 gene fusion formed on the the derivative chromosome 22 (Philadelphia chromosome).
How can we detect the BCR-ABL1 fusion using FISH?
- Using probes which span the breakpoints of ABL1 in the 9 and BCR on the 22 we will be able to see both probes present in on the derivative chromosome in which the BCR-ABL1 and ABL1-BCR gene fusions have formed.
What the consequence of the BCR-ABL1 fusion gene that is present in CML cases? How can the effects of this gene fusion be treated?
- The BCR-ABL1 fusion gene produces a chimeric abnormal protein which has enhanced tyrosine kinase activity.
- Tyrosine Kinase Inhibitors (TKI) are used to treat CML.
- The drug Imatinib (Glivec, Gleevec, STI571) is designed to fit into the ATP binding site of this chimeric protein.
- If the binding site is blocked the BCR-ABL1 protein is effectively switched off.
What is Imatinib?
- The BCR-ABL1 fusion gene produces a chimeric abnormal protein which has enhanced tyrosine kinase activity.
- Tyrosine Kinase Inhibitors (TKI) are used to treat CML.
- The drug Imatinib (Glivec, Gleevec, STI571) is designed to fit into the ATP binding site of this chimeric protein.
- If the binding site is blocked the BCR-ABL1 protein is effectively switched off.
What guidelines are available for monitoring the response to treatment in CML?
- There are British Committee for Standards in Haematology (BCSH) guidelines to monitor response, updated in Baccarani (2009), on belhalf of European Leukaemia net (ELN).
- ACC BPGs.
- May have local policies.
- After diagnosis, follow up samples at intervals.
What are the definitions for how CML patients are responding to treatment in cytogenetic terms?
- Definitions of cytogenetic an molecular responses (Baccarani et al 2009).
- Cytogenetic Response is measured by (CgR) Ph+ve (Philadelphia positive) cell levels.
- Based on Ph positive metaphase cells in marrow.
- If someone still has 96-100% Ph+ve levels after Imatinib treatment they are considered to have no cytogenetic response to treatment (noCgR).
- If someone still has 66-95% Ph+ve levels after Imatinib treatment they are considered to have minimal cytogenetic response to treatment (minCgR).
- If someone still has 36-65% Ph+ve levels after Imatinib treatment they are considered to have minor cytogenetic response to treatment (mCgR).
- If someone still has 1-35% Ph+ve levels after Imatinib treatment they are considered to have partial cytogenetic response to treatment (PCgR).
- If someone still has 0% Ph+ve levels after Imatinib treatment they are considered to have complete cytogenetic response to treatment (CCgR).
- Once you have achieved a complete cytogenetic response it is recommended that you then detect biomolecular means from then on.
In cytogenetic terms what is considered to be no response to Imatinib treatment for CML?
- If someone still has 96-100% Ph+ve levels after Imatinib treatment they are considered to have no cytogenetic response to treatment (noCgR).
In cytogenetic terms what is considered to be minimal response to Imatinib treatment for CML?
- If someone still has 66-95% Ph+ve levels after Imatinib treatment they are considered to have minimal cytogenetic response to treatment (minCgR).
In cytogenetic terms what is considered to be minor response to Imatinib treatment for CML?
- If someone still has 36-65% Ph+ve levels after Imatinib treatment they are considered to have minor cytogenetic response to treatment (mCgR).
In cytogenetic terms what is considered to be partial response to Imatinib treatment for CML?
- If someone still has 1-35% Ph+ve levels after Imatinib treatment they are considered to have partial cytogenetic response to treatment (PCgR).
In cytogenetic terms what is considered to be complete response to Imatinib treatment for CML?
- If someone still has 0% Ph+ve levels after Imatinib treatment they are considered to have complete cytogenetic response to treatment (CCgR).
What are the definitions for how CML patients are responding to treatment in molecular terms?
- Can do molecular testing instead of cytogenetic to see how much BCR-ABL transcription you can detect.
- Once you have achieved a complete cytogenetic response it is recommended that you then detect biomolecular means from then on.
- Molecular response is based upon measuring the log reduction (LR) BCR-ABL protein in the marrow.
- A less than major molecular response is defined as a log reduction less than 3.
- A major molecular response is defined as a log reduction of more than 3 in 2 consecutive samples.
- A complete molecular response / molecular remission is BCR-ABL1 negative in 2 consecutive samples of adequate sensitivity (>=4 LR).
In molecular terms what is considered to be a less than major molecular response to Imatinib treatment for CML?
- Molecular response is based upon measuring the log reduction (LR) BCR-ABL protein in the marrow.
- A less than major molecular response is defined as a log reduction less than 3.
In molecular terms what is considered to be a major molecular response to Imatinib treatment for CML?
- Molecular response is based upon measuring the log reduction (LR) BCR-ABL protein in the marrow.
- A major molecular response is defined as a log reduction of more than 3 in 2 consecutive samples.
In molecular terms what is considered to be complete
molecular response / molecular remission for CML in response to Imatinib?
- Molecular response is based upon measuring the log reduction (LR) BCR-ABL protein in the marrow.
- A complete molecular response / molecular remission is BCR-ABL1 negative in 2 consecutive samples of adequate sensitivity (>=4 LR).
How do we evaluate the response to treatment for CML?
- Definitions of cytogenetic an molecular responses (Baccarani et al 2009).
- Cytogenetic Response is measured by (CgR) Ph+ve (Philadelphia positive) cell levels. Based on Ph positive metaphase cells in marrow.
- Molecular response is based upon measuring the log reduction (LR) BCR-ABL protein in the marrow.
- There are recommendations for evaluating response to Imatinib by testing at regular time intervals of 3, 6, 12, 18 months.
- The response is graded as optimal, suboptimal, or failure in defined terms of haematological response, cytogenetic response, and molecular response.
- There are also definitions of response to 2nd generation TKIs Dasatinib and Nilotinib as second line therapy - also trials of first line therapy.
- 3rd generation Bosutinib is being evaluated.
What treatments are currently available or being evaluated for the treatment of CML?
- Currently used treatment is an TKI called Imatinib.
- There are also definitions of response to 2nd generation TKIs Dasatinib and Nilotinib as second line therapy - also trials of first line therapy.
- 3rd generation Bosutinib is being evaluated.
Discuss AML.
- AML is Acute Myeloid Leukaemia.
- Acquired abnormalities at the chromosome level can give you information regarding diagnosis, prognosis, and response to treatment e.g. t(15;17), t(8;21), inv(16) are all diagnostic.
- Cytogenetic results from previous adult and childhood treatment trials have been used to contribute to the assessment of the prognostic risk.
AML can be classified into different risk groups based upon cytogenetic abnormalities present. Outline these.
- Grimwade ‘98 - cytogenetics give quite reliable information with regards to the risk associated and the outcome of the CML.
- Favourable risk = t(8;21), t(15;17), inv(16).
- Intermediate risk - normal cytogenetic abnormalities not associated as favourable or adverse.
- Adverse risk = -5, -7, del(5q), Abn 3q, Complex.
- Updated in 2010 but only in adults.
- Child reports in 2010 reported slightly different risk associations than in both adult and children combined.
What CML abnormalities are in the favourable risk group?
- Favourable risk = t(8;21), t(15;17), inv(16).
What CML abnormalities are in the intermediate risk group?
- Intermediate risk - normal cytogenetic abnormalities not associated as favourable or adverse.
What CML abnormalities are in the adverse risk group?
- Adverse risk = -5, -7, del(5q), Abn 3q, Complex.
AML can be classified into different risk groups based upon cytogenetic abnormalities present. Outline the abnormalities reported in these risk groups for children only.
- Favourable risk = t(8;21), inv(16).
- Intermediate risk - 11q23
- Adverse risk = -7, Abn 5q, t(6;9)
Discuss MDS.
- MDS is myelodysplasia..
- Chromosome abnormalities contribute to the International Prognostic Scoring (IPSS).
- Good = normal, -Y, del(5q), del(20q).
- Intermediate = other abnormalities.
- Poor = complex (>3 abnormalities) or chromosome 7 anomalies.
What cytogenetic abnormalities are associated with a good prognosis in MDS?
- Good = normal, -Y, del(5q), del(20q).
What cytogenetic abnormalities are associated with an intermediate prognosis in MDS?
- Intermediate = other abnormalities.
What cytogenetic abnormalities are associated with a poor prognosis in MDS?
- Poor = complex (>3 abnormalities) or chromosome 7 anomalies.
What are the 3 main myeloid malignancies?
- CML = Chronic Myeloid Leukaemia
- AML = Acute Myeloid Leukaemia
- MDS = Myelodysplasia
