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Northcott Lab: Research

Integrative genomic analysis of medulloblastoma

Genome-wide analyses of primary medulloblastoma tissue samples using various microarray platforms provided early clues into the genomic and transcriptional heterogeneity underlying medulloblastoma1,2. These prior efforts disclosed recurrent regions of DNA copy-number alteration involved in medulloblastoma pathogenesis and led to the identification and initial description of the four consensus molecular subgroups: WNT, SHH, Group 3, and Group 43. More recently, next-generation sequencing (NGS) of large medulloblastoma series has delineated recurrent somatic driver alterations in each of the four subgroups4. These studies have provided firm molecular evidence substantiating the concept of medulloblastoma as not a single disease, but more accurately, four distinct diseases consisting of discrete combinations of varied somatic alterations. Notable discoveries made possible through genomics include consistent targeting of epigenetic regulators across subgroups1,5, PVT1-MYC fusion transcripts in MYC-amplified Group 36, highly recurrent SNCAIP duplications in Group 46, and aberrant activation of GFI1 and GFI1B in Group 3 and Group 4 resulting from diverse structural variants that lead to enhancer hijacking7. Working closely with the Computational Biology Department at St. Jude and collaborators from around the globe, my lab continues to analyze large, multidimensional ‘omics datasets in an effort to fully annotate all genomic alterations contributing to medulloblastoma initiation, maintenance, progression, relapse, and metastases.

Figure 1. Recurrent somatically altered genes in medulloblastoma subgroups.

Recurrent  somatically altered genes in medulloblastoma subgroups

(a) Frequency and subgroup distribution of the most commonly mutated genes in medulloblastoma. Summarization is based on results reported by Jones et al., Pugh et al., and Robinson et al., that were independently published in Nature in 2012 and subsequently merged and published as shown in Nature Reviews Cancer4.

Summary of candidate medulloblastoma driver genes according to molecular subgroup

(b) Summary of candidate medulloblastoma driver genes according to molecular subgroup as published in Nature Reviews Cancer4.

Functional characterization of novel medulloblastoma driver genes

Recent NGS studies have revealed a wealth of novel candidate genes that are recurrently targeted by mutations or structural variants and presumed to contribute to medulloblastoma development4. Despite these insights, very few candidate driver genes have been functionally studied in the context of either normal cerebellar development or medulloblastoma. This is especially true for Group 3 and Group 4, two clinically challenging subgroups for which additional faithful genetic models of the disease are desperately needed8. My lab is harnessing insights gained through our NGS studies and linking this information with state-of-the-art in vitro and in vivo approaches to study candidate gene function in spatially and temporally relevant cellular environments. Our lab has considerable expertise with somatic perturbation of oncogenes and tumor suppressor genes in utero, giving us the capacity to rapidly screen candidates of interest in the early embryonic hindbrain. These efforts are focused on the generation of new subgroup-specific models of medulloblastoma that will improve our understanding of subgroup biology and enable urgent preclinical studies, especially for Group 3 and Group 4.

Figure 2. Perturbation strategies for studying candidate medulloblastoma driver genes in vivo. (a) Cartoon detailing the in utero lentiviral transduction method we employ for perturbation of candidate genes in the developing mouse hindbrain (e12.5-e13.5). (b) Imaging showing successful transduction of multiple cerebellar progenitor cell types with recombinant lentiviruses. Embryos (e12.5) were transduced with lentivirus expressing eGFP and imaged 24hrs post-transduction.

Perturbation strategies for studying candidate medulloblastoma driver genes

Cellular origins of medulloblastoma subgroups

During the past decade, an extensive amount of molecular, biological, and clinical data has emerged to support the vastly heterogeneous nature of medulloblastoma. As a community, we now recognize the existence of at least four unique molecular subgroups3, each characterized by distinct genetic, epigenetic, and transcriptional landscapes that suggest these subgroups arise from different cell populations in the developing hindbrain. Various, elegant genetically engineered mouse models have substantiated distinct cellular origins for the WNT and SHH subgroups, whereas the origins of Group 3 and Group 4 remain an open question. Our recent work describing transcriptional regulatory circuitry inherent to medulloblastoma subgroups has provided new clues into subgroup identity and implicated putative cells-of-origin for poorly characterized Group 3 and Group 4 subgroups (Lin, Erkek et al., In Press). We are now utilizing a combination of next-generation transcriptional and epigenetic analyses to prospectively ‘fingerprint’ spatiotemporally distinct stem/progenitor cell populations in the developing mouse hindbrain in order to confirm findings inferred from our human genomic studies. Identifying and confirming the cellular origins of medulloblastoma subgroups will enable us to generate improved preclinical models, identify tumor cell dependencies present in normal cell populations, and allow for dramatically improved tumor/normal cell comparisons with respect to transcriptional and epigenetic landscapes.

Figure 3. Medulloblastoma super-enhancers implicate subgroup-specific cellular origins.

Summary  of super-enhancers (SEs) discovered in Group 4 medulloblastoma. Notable  SE-associated genes in Group 4 are marked.

(a) Summary of super-enhancers (SEs) discovered in Group 4 medulloblastoma. Notable SE-associated genes in Group 4 are marked.

Regulatory circuitry in Group 3 and Group 4 medulloblastoma.
(b) Regulatory circuitry in Group 3 and Group 4 medulloblastoma. Master transcriptional regulators inferred to establish Group 3 and Group 4 subgroup identity and their connectivity are depicted.

Spatially localized expression of Group 4 master regulators in  the developing mouse cerebellum (e13.5).

(c) Spatially localized expression of Group 4 master regulators in the developing mouse cerebellum (e13.5). Lmx1a, Eomes, and Lhx2 are co-expressed in the nuclear transitory zone at this important timepoint in early cerebellar development.

References

  1. Northcott PA, et al. Multiple recurrent genetic events converge on control of histone lysine methylation in medulloblastoma. Nat Genet 41:465-472, 2009. doi:10.1038/ng.336.
  2. Northcott PA, et al. Medulloblastoma comprises four distinct molecular variants. J Clin Oncol 29, 1408-1414, 2011. doi:10.1200/JCO.2009.27.4324.
  3. Taylor MD, Northcott PA, et al. Molecular subgroups of medulloblastoma: the current consensus. Acta Neuropathol 123, 465-472, 2012. doi:10.1007/s00401-011-0922-z.
  4. Northcott PA, et al. Medulloblastomics: the end of the beginning. Nat Rev Cancer 12:818-834, 2012. doi:10.1038/nrc3410.
  5. Batora NV, Sturm D, Jones DT, Kool M, Pfister SM, Northcott PA. Transitioning from genotypes to epigenotypes: why the time has come for medulloblastoma epigenomics. Neuroscience 264:171-185, 2014. doi:10.1016/j.neuroscience.2013.07.030.
  6. Northcott PA*, Shih DJ*, et al. Subgroup-specific structural variation across 1,000 medulloblastoma genomes. Nature 488:49-56, 2012. doi:10.1038/nature11327.
  7. Northcott PA*, Lee C*, Zichner T,*et al. Enhancer hijacking activates GFI1 family oncogenes in medulloblastoma. Nature 511:428-434, 2014. doi:10.1038/nature13379.
  8. Northcott PA, Korshunov A, Pfister SM, Taylor MD. The clinical implications of medulloblastoma subgroups. Nat Rev Neurol 8:340-351, 2012. doi:10.1038/nrneurol.2012.78.

*Equal contribution.