AGBT 2010 - Keynote speaker: James Downing - St. Jude Children's Hospital
The Molecular Pathology of Acute Leukemia
Was head of pathology at St. Jude for many years - doing cancer genomics before it was called cancer genomics.
First time at AGBT
No methodology or technology - focus on biology and clinical relevance. Not going to present NGS data! Using completely outdated technology - and all of it was published in the last 12 months.
The cancer he's focused on is the best characterized of all the cancers.
What leukemia really is: Proliferating B-cells that rapidly take over the whole body. Highest tumor lode of all the cancers. In his generation, 95% of children died within 12 months of diagnosis. Now have 80-85% cure... but relapses happen in 30%.
[Classical diagram of immune system lineage]
Mutations in early progenitors generate leukemias. Two types: ALL and AML. They are not homogeneous diseases, however. Distinct biological subtypes are characterized by translocations. - They contribute to the leukemia: Necessary, but not sufficient.
What are the biologic processes that need to be altered to generate leukemia:
1. Alteration in self-renewal capacity - need to become "immortal" (unlimited self-renewal) (eg AML1-ETO)
2. Need to have an altered response to growth signals - contnued growth (eg. BCR-ABL1)
3. Block in apoptosis (eg PML-RAR alpha)
4. Block in differentiation
Doing "routine molecular diagnosis":
* CNV, expression, etc
* Use Affy Chips
What have they found? (using 242 diagnostic ALLs with matched germ line DNA.)
* there are a small number of copy number changes per casee... vary markedly across the different subtypes. (eg, MLL: ~1, other has ~11)
* more Deletions that Amplifications
* 60% of b lineage all have a genetic lesion in a gene regulating B-cell differentiantion (PAC5, Ikaros, EVF, LDF1, BNK)
PAX5 deletions most common.
* 10 exons...
* Half of deletions deleted half of the genes
* Others delete required domains
* some were homozygous, but not all.
* Lots of fusions with this gene occurs as well.
* Point mutations were also seen in binding domains...
[Ok, so this gene can be deleted in many ways... got it. The cells find ways to kill off this gene.]
Haploinsufficiency in PAX5 deficient mice
* Was not sufficient to cause lymphoma.
* cooperates with BCR-ABL1 to cause lymphoma. (Mouse Model)
* strong driving pressure for diabling the b-cell differentiation genes in Leukemia.
60% of B-progenitors ALL have Mutations in B-cell regulatory Genes
Look at Ikaros
* entire literature about altered isoforms.
* saw a high frequency of mutations in BCR-ABL1 ALL,
* 85% of BCR-ABL ALL have deletions of Ikaros: Almost never see the deletions in Ikaros.
* mapping deletions of Ikaros: Some are complete, but there is a subset of deletions that commonly knock out all 4 zinc fingers (exons 3-6).
* Never see Ikaros "isoforms" without these deleitons. There probably are no isoforms - it's always genetic lesions.
* Deletions typically happen within a few bases of each other - result from aberrant RAG-mediated recombinations.
Start putting the lesions together. [Nice lists of genes for each of the 3 pathways]
Clinical relevance:
* looking for markers in a new cohort. Remove two types of ALL (BCR-ABL1 + infant), look at 221 samples: Are there new markers?
* Yes, it was Ikaros: 75% of relapse if you have Ikaros deletions.
Compare BCR-ABL1- and Ikaros- (Bad outcome) with BCR-ABL1+ ALL (Also has Ikaros deletions)
* Significant expression similarity
* Look at the Kinases: JAK family, which have a high rate of mutations in ALLs.
JAK mutations:
* not seen in other types of cancers - unique to JH2 domain, clustering in a single spot. (R683)
* Turns out that high risk ALL have JAK deletions.
CRLF2 = TSLPR, IL-7/IL07R
* Over expression of this receptor (compensating for Jak Mutations and lack of signaling), combine to cause a proliferative signal. [I didn't get everything here.]
Looking at high risk again:
* Ikaros deletions
* Jak Mutations
* CRLF2 (cytokine receptor mutations)
What other kinases are activated in this subset of patients?
* Work in progress
* quick review of other genes they're now finding... [too fast to get that down.]
Genetic Alterations Acquired at Relapse
* Relapsing is only 20% blast population.
* Need to Flow sort.
* CDKN2A/B mutations
* [list of genes, including ikaros... ]
* No common mechanism of relapse - variety of pathways
* Varieties do not include drug target mutations. It's always in signalling, etc.
* 7% of relapse is "unrelated" (secondary leukemia)
* 8% same as diagnosis
* 34% clonal evolution from diagnosis
* 51% clonal evolution from pre-leukemic clone
Summary:
* small number of variation
* Ikaros mutations
* Aberant RAG-mdeidated recombination
* JAK mutations
* ...
This disease "begs for NGS" - Get a complete picture of what's going on.
* Collaborating with WashU. (Mardis, Wilson, Ley)
* Doing the "Bad" leukemias (infant, high risk, CBF)
* also doing brain and solid tumours (neuroblastoma osteosarcoma, retinoblastoma)
* Started Feb 1st - already have 5 genomes and matched normals.
* over $50M invested in this project
Was head of pathology at St. Jude for many years - doing cancer genomics before it was called cancer genomics.
First time at AGBT
No methodology or technology - focus on biology and clinical relevance. Not going to present NGS data! Using completely outdated technology - and all of it was published in the last 12 months.
The cancer he's focused on is the best characterized of all the cancers.
What leukemia really is: Proliferating B-cells that rapidly take over the whole body. Highest tumor lode of all the cancers. In his generation, 95% of children died within 12 months of diagnosis. Now have 80-85% cure... but relapses happen in 30%.
[Classical diagram of immune system lineage]
Mutations in early progenitors generate leukemias. Two types: ALL and AML. They are not homogeneous diseases, however. Distinct biological subtypes are characterized by translocations. - They contribute to the leukemia: Necessary, but not sufficient.
What are the biologic processes that need to be altered to generate leukemia:
1. Alteration in self-renewal capacity - need to become "immortal" (unlimited self-renewal) (eg AML1-ETO)
2. Need to have an altered response to growth signals - contnued growth (eg. BCR-ABL1)
3. Block in apoptosis (eg PML-RAR alpha)
4. Block in differentiation
Doing "routine molecular diagnosis":
* CNV, expression, etc
* Use Affy Chips
What have they found? (using 242 diagnostic ALLs with matched germ line DNA.)
* there are a small number of copy number changes per casee... vary markedly across the different subtypes. (eg, MLL: ~1, other has ~11)
* more Deletions that Amplifications
* 60% of b lineage all have a genetic lesion in a gene regulating B-cell differentiantion (PAC5, Ikaros, EVF, LDF1, BNK)
PAX5 deletions most common.
* 10 exons...
* Half of deletions deleted half of the genes
* Others delete required domains
* some were homozygous, but not all.
* Lots of fusions with this gene occurs as well.
* Point mutations were also seen in binding domains...
[Ok, so this gene can be deleted in many ways... got it. The cells find ways to kill off this gene.]
Haploinsufficiency in PAX5 deficient mice
* Was not sufficient to cause lymphoma.
* cooperates with BCR-ABL1 to cause lymphoma. (Mouse Model)
* strong driving pressure for diabling the b-cell differentiation genes in Leukemia.
60% of B-progenitors ALL have Mutations in B-cell regulatory Genes
Look at Ikaros
* entire literature about altered isoforms.
* saw a high frequency of mutations in BCR-ABL1 ALL,
* 85% of BCR-ABL ALL have deletions of Ikaros: Almost never see the deletions in Ikaros.
* mapping deletions of Ikaros: Some are complete, but there is a subset of deletions that commonly knock out all 4 zinc fingers (exons 3-6).
* Never see Ikaros "isoforms" without these deleitons. There probably are no isoforms - it's always genetic lesions.
* Deletions typically happen within a few bases of each other - result from aberrant RAG-mediated recombinations.
Start putting the lesions together. [Nice lists of genes for each of the 3 pathways]
Clinical relevance:
* looking for markers in a new cohort. Remove two types of ALL (BCR-ABL1 + infant), look at 221 samples: Are there new markers?
* Yes, it was Ikaros: 75% of relapse if you have Ikaros deletions.
Compare BCR-ABL1- and Ikaros- (Bad outcome) with BCR-ABL1+ ALL (Also has Ikaros deletions)
* Significant expression similarity
* Look at the Kinases: JAK family, which have a high rate of mutations in ALLs.
JAK mutations:
* not seen in other types of cancers - unique to JH2 domain, clustering in a single spot. (R683)
* Turns out that high risk ALL have JAK deletions.
CRLF2 = TSLPR, IL-7/IL07R
* Over expression of this receptor (compensating for Jak Mutations and lack of signaling), combine to cause a proliferative signal. [I didn't get everything here.]
Looking at high risk again:
* Ikaros deletions
* Jak Mutations
* CRLF2 (cytokine receptor mutations)
What other kinases are activated in this subset of patients?
* Work in progress
* quick review of other genes they're now finding... [too fast to get that down.]
Genetic Alterations Acquired at Relapse
* Relapsing is only 20% blast population.
* Need to Flow sort.
* CDKN2A/B mutations
* [list of genes, including ikaros... ]
* No common mechanism of relapse - variety of pathways
* Varieties do not include drug target mutations. It's always in signalling, etc.
* 7% of relapse is "unrelated" (secondary leukemia)
* 8% same as diagnosis
* 34% clonal evolution from diagnosis
* 51% clonal evolution from pre-leukemic clone
Summary:
* small number of variation
* Ikaros mutations
* Aberant RAG-mdeidated recombination
* JAK mutations
* ...
This disease "begs for NGS" - Get a complete picture of what's going on.
* Collaborating with WashU. (Mardis, Wilson, Ley)
* Doing the "Bad" leukemias (infant, high risk, CBF)
* also doing brain and solid tumours (neuroblastoma osteosarcoma, retinoblastoma)
* Started Feb 1st - already have 5 genomes and matched normals.
* over $50M invested in this project
Labels: AGBT 2010
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