New Orleans, LA—Genomic sequencing and protein analysis are providing new insights into hematologic malignancies, paving the way for treatment advances and personalized medicine. At the Presidential Symposium at ASH 2013, the potential use of genomics in clinical decision-making was explored.
Genomics and Childhood Leukemias
Next-generation DNA sequencing has retrospectively identified new genetic subtypes of leukemia, said James R. Downing, MD, Scientific Director, St Jude Children’s Research Hospital, Memphis, TN. Dr Downing described the Pediatric Cancer Genome Project, a 3-year $65-million project to sequence the complete normal and cancer genomes of 600 children and adolescents with some of the most aggressive and least understood cancers. Over the 3 years, 700 tumors have been sequenced.
“I can say today that this has been the most successful project that I have ever been associated with,” said Dr Downing. “For every single pediatric cancer that we looked at, we gained fundamental new insights into the pathogenesis of these diseases, and we gained new prognostic markers and new therapeutic targets.”
He described 3 examples of whole-genome sequencing and their clinical implications.
The first is the discovery of a fusion gene that is responsible for almost 30% of a rare subtype of pediatric acute myeloid leukemia (AML) called acute megakaryoblastic leukemia (AMKL). It accounts for approximately 4% to 15% of pediatric AML, and is associated with poor prognosis and a high rate of treatment failure. The fusion product detected by sequencing 14 cases of pediatric non–Down syndrome AMKL involved 2 genes on chromosome 16—GLIS2 (a protein normally expressed only in renal cells) and CBFA2T3. Half of the patients carried the CBFA2T3-GLIS2 fusion, which enhances the self-renewal of megakaryocyte progenitors.
Gene-expression profiling showed that bone morphogenetic protein was markedly upregulated in the cases that expressed the fusion product.
Patients with the CBFA2T3-GLIS2 fusion gene had worse overall survival than patients without the CBFA2T3-GLIS2 fusion. “From this discovery effort, we have a new prognostic marker; one that I would argue we would probably want to know at diagnosis, so that patients that don’t respond can be funneled into therapeutic protocols that provide a more aggressive treatment approach than the standard AML therapy,” Dr Downing suggested.
Two other examples are the discovery of a new high-risk acute lymphoblastic leukemia (ALL) genetic subtype referred to as BCR-ABL1–like ALL, and hypodiploid ALL.
Of the patients in the project, 50% were found to have CRLF2 rearrangements, and half of those mutations have activating mutations in the Janus kinase family. RNA sequencing, whole-genome sequencing, and whole-exome sequencing of 165 pediatric and adult cases of BCR-ABL1–like ALL revealed unique fusion products that led to activation of the tyrosine kinase.
Hypodiploid ALL is a rare aggressive subtype of pediatric ALL. Genetic alterations involving TP53, RB1, and IKZF2 are common in patients with hypodiploid ALL.
Digital Sequencing and Clonality in Myelodysplastic Syndromes
Matthew J. Walter, MD, Associate Professor of Medicine, Washington University, St Louis, MO, discussed the use of whole-genome sequencing of patients’ DNA to study myelodysplastic syndromes (MDS). His group has undertaken a large-scale sequencing effort to identify gene mutations in MDS, focusing on the transition to secondary AML. “Along the way, we realized that using digital sequencing, we could study clonality,” Dr Walter said.
Only 50% of patients with MDS have a cytogenetic abnormality. Using whole-genome sequencing, it was discovered that all patients with MDS carry genetic abnormalities. Digital sequencing has been able to identify additional patients with genetic mutations, “above and beyond cytogenetics,” he noted.
Sequencing has found an abnormal karyotype or mutation in up to 90% of patients with MDS, and has identified approximately 20 genes that are consistently mutated in >2% of these patients. There are 6 categories of frequently mutated genes in MDS: TP53, spliceosome genes, epigenetic modifiers, cohesin genes, transcription factor genes, and activated signaling genes. In a gene category, each patient tends not to have more than 1 mutation.
These mutations impact prognosis, Dr Walter said. Patients with TP53 mutations tend to have a complex karyotype and deletions on chromosomes 5 and 7, and this group has a poor prognosis. Mutations in 4 other genes—EZH2, ETV6, RUNX1, and ASXL1—have independent prognostic significance for overall survival. Patients categorized as intermediate risk-1 who have a mutation in 1 of these genes have overall survival that is as poor as patients with intermediate risk–2 MDS. “This is a tipping point for recommendation of allogeneic hematopoietic stem-cell transplantation—patients that are intermediate risk-2,” said Dr Walter.
Clonal heterogeneity in MDS is common. The clonal evolution model suggests that the progression of MDS to secondary AML is characterized by the persistence of a single founder clone, defined by hundreds of mutations, and the outgrowth of at least 1 new subclone that contains additional mutations. A single population of MDS cells, therefore, undergoes several rounds of mutation, engendering multiple subpopulations that are present in secondary AML.
To be useful clinically, “we need to know if a mutation is in the founding clone or a subclone,” said Dr Walter. Without targeting all clones in a tumor, it is unlikely that a durable response can be achieved.