Haematological Malignancy Screening Flashcards
What are the 2 types of haematological malignancy?
- leukaemias
- lymphomas
What is a leukaemia? What are the 2 types?
they are tumours of the blood and the haematopoietic lineages
they are either myeloid (ML) or lymphoid (LL)
they are then defined as either acute (ALL/AML) or chronic (CML, CLL)
What cells are implicated in myeloid and lymphoid leukaemias?
myeloid:
- granulocytes
- monocytes
- erythrocytes
- platelets
lymphoid:
- B cells
- T cells
- NK cells
What is the difference between acute and chronic leukaemias?
acute:
aggressive disease with huge amounts of uncontrolled cell proliferation
chronic:
there is a much slower, steady increase in the number of cells
What is the definition of a lymphoma?
tumours of lymphocytes and the immune/lymphatic systems
these tend to be more solid, rather than circulating tumours
What are the 3 stages linked to the outcome of cancer treatment?
What are examples of determinants that affect the outcome?
Through evolving cancer diagnostics, how can each parameter be expanded?
complete the process of diagnosis and patient care
How does molecular diagnostics fit into the process of diagnosis and patient care?
How is molecular diagnostics involved in the diagnosis of haematological malignancies?
- it can identify disease causing translocations in leukaemia and lymphoma
- it can identify disease causing point mutations in myeloproliferative disorders
How is molecular diagnostics involved in the prognostication and treatment selection of haematological malignancies?
- It identifies DNA translocations and deletions which define prognostic groups and treatment choices
- it can idenitfy immunoglobulin gene mutations which loads defining risk
How is molecular diagnostics involved in disease monitoring, response assessment and relapse prevention?
tracking translocations in myeloid leukaemias
What are the 2 types of chromosomal translocations?
- fusion genes
- gene deregulation
What is involved in production of fusion genes through translocation?
2 chromosomes join together, leading to a fusion protein which causes problems downstream
e.g. the fusion protein may be involved in promoting cell division and the aberrant region means that it cannot be switched off
What is involved in gene deregulation as a form of translocation?
the promoter of a gene can be switched for the promoter of another gene
this leads to increased proliferation and lack of cell death as the gene is switched on in the wrong location
What is shown in this fluorescence in situ hybridisation image?
In a normal cell there are 2 copies of each gene (2 red dots and 2 green dots)
In a leukaemic cell, there is one normal copy of each gene and one translocation where the red and green dots have joined together
Which diagnostic technique is usually used in myeloid leukaemias?
Polymerase chain reaction for the fusion gene translocation
there is a predictable break point/fusion
mRNA is processed to remove introns
What are the characteristics defining the use of PCR for diagnostically defining a translocation?
- it is highly sensitive and specific
- it works only over a small size range (<1kb - 10kb)
- it only works at the gene level
- it needs to have a consistent junction and a precise sequence
Which diagnostic technique is usually used in lymphomas?
FISH for detecting promoter/enhancer substitution
There is an unpredictable break point/ fusion and no abnormal mRNA
What are the characteristics defining the use of FISH for diagnostically defining translocations?
- it is highly specific, but less sensitive
- it is suitable for large sizes of 100 kB +
- it is suitable at the chromosomal lvel
- it doesn’t need to have a consistent junction and the precise sequence doesn’t need to be known
What is the typical presentation of chronic myeloid leukaemia?
abdominal discomfort or pain:
- splenomegaly
- splenic infarction
fatigue:
- anaemia
gout:
- hyperuricaemia
What age group is CML typically seen in? What gender is more commonly affected?
median age is 55-60 with <10% of cases being < 20
the male to female ratio is 1.5:1
What would a full blood count in someone with CML look like?
there would be leucocytosis and anaemia
the leukocytes are pushing the red blood cells out of the blood vessels
What feature of CML is shown?
left-shifted immature myelocytes
What translocation is known for causing CML? How does it work?
the top part of chromosome 22 breaks off, exposing the BCR gene
the ABL gene breaks off of chromosome 9 and attaches to the exposed BCR gene
this creates the BCR-ABL fusion gene which codes an active tyrosine kinase
How does the BCR-ABL gene affect cell division?
the active tyrosine kinase gives signals for cells to divide and the signals cannot be switched off
How can the BCR-ABL gene be detected?
conventional reverse transcriptase (RT)-PCR detects the presence of the fusion transcript
What is the treatment for CML and how does it work?
Gleevec (imatinib mesylate)
it is a small molecule that is designed to block the active site in the BCR-ABL tyrosine kinase
How is the treatment response to Gleevec assessed?
the number of cells that still have the BCR-ABL transformation after treatment are counted
this is done through real-time PCR (qPCR)
Why can normal PCR not be used to assess response to Gleevec in CML?
PCR is a qualitative assessment meaning that it can say whether or not CML is present, but cannot show how many cells are left
How does qPCR work?
it is the same as PCR but with a fluorescent probe
each time a copy of the BCR-ABL transcript is made, the probe is incorporated and light is emitted
the light can be measured to know how many copies of a particular gene are present
What is one of the main benefits in using qPCR rather than cytogenetics?
it is more sensitive and can identify a relapse early
if a patient becomes resistant to drug treatment then a second generation molecule can be used (dasatinib)
What is shown in this image?
bone marrow in a patient with a chronic myeloproliferative disorder (CMPD)
What mutations are implicated in CMPD?
mutations in the JAK2 gene
this also codes for a tyrosine kinase that signals for cells to proliferate
What are the 3 mutations that may interact to cause varying severities of myeloproliferative neoplasms?
- JAK2
- MPL
- CALR
What is meant by next generation sequencing?
sequencing hundreds of millions of pieces of DNA in a single run
What procedures does NGS allow for?
- the sequencing of an entire genome at low depth of coverage (WGS)
- sequencing the entire exome of the transcribed genes at a higher depth (WES)
- can sequence large panels of genes
When looking at chronic lymphocytic leukaemia (CLL), what is it important to distinguish?
- indolent disease with little risk
- aggressive disease with high risk
What cells are implicated in CLL? What ages and genders does it most commonly affect?
It is a disease of B cells
It has a male to female ratio of 2:1
the median age is 70, with the majority of cases > 50
What are the typical symptoms of B-CLL?
it is usually asymptomatic for many years and discovered by accident through a full blood count
- fatigue
- autoimmune anaemia
- splenomegaly
- lymphadenopathy
What would be seen on a full blood count of someone with B-CLL?
lymphocytosis
What is meant by monoclonal B cell lymphocytosis and how does it affect B-CLL?
around 5% of the population harbour small monoclonal B cell expansions (MBL)
they are phenotypically similar to CLL, but with a number of cells lower than 5 x 10^9
around 1% of MBLs will progress to CLL each year
What are the stages in the B cell lifecycle?
What is involved in somatic hypermutation?
The B cell receptor is mutated by the body to give a better immune response through antigen recognition
The B cells that are better at responding to the antigen are proliferated
The B cells that are worse at responding to the antigen are eradicated
label the stages in the immunoglobulin VH gene sequencing pathway that is a prognostic factor for CLL
this process tells you where the B cell came from
Complete the CLL cytogenetics by FISH for specific genes
you would expect to see 2 spots in the nucleus of every cell as there should be 2 copies of every gene
what is involved in early stage CLL stratification?
patients with a mutated immunoglobulin gene have a low risk
patients with an unmutated gene have a severe risk
patients with p53 deletion have a very poor outcome
Why is chemotherapy not suitable in CLL?
any chemotherapy agent will damage DNA
this damage is detected by ATM that switches on p53 (this is the gene that causes cell death)
If there is no p53 present, chemotherapy will not work
How is a risk assessment carried out and CLL patients stratified?
What is the treatment for someone with mutated Ig and no deletions?
watch and wait
minimally toxic therapy with:
- rituximab
- chlorambucil
- fludarabine
What is the treatment for someone with unmutated Ig but no 11q/17p deletion?
nucleoside analogue combination regimens
- fludarabine and cyclophosphamide
- fludarabine and rituximab
What is the treatment for someone with CLL who has the 11q/17p deletion?
a high risk clinical trial involving CAMPATH anti-CD52 antibody
Why is the B cell receptor a target for drug treatment in CLL?
B cell receptor signalling is vital to CLL cells as it is involved in many processes
What are the stages in the cellular signal transduction pathway?
Why is B-cell receptor signalling crucial to CLL?
it activates vital downstream pathways that lead to cell proliferation and resistance to apoptosis
