title: Identification of Variant-Specific Functions of PIK3CA by Rapid Phenotyping of Rare Mutations.
abstract: Large-scale sequencing efforts are uncovering the complexity of cancer genomes, which are composed of causal "driver" mutations that promote tumor progression along with many more pathologically neutral "passenger" events. The majority of mutations, both in known cancer drivers and uncharacterized genes, are generally of low occurrence, highlighting the need to functionally annotate the long tail of infrequent mutations present in heterogeneous cancers. Here we describe a mutation assessment pipeline enabled by high-throughput engineering of molecularly barcoded gene variant expression clones identified by tumor sequencing. We first used this platform to functionally assess tail mutations observed in PIK3CA, which encodes the catalytic subunit alpha of the phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) frequently mutated in cancer. Orthogonal screening for PIK3CA variant activity using in vitro and in vivo cell growth and transformation assays differentiated driver from passenger mutations, revealing that PIK3CA variant activity correlates imperfectly with its mutation frequency across breast cancer populations. Although PIK3CA mutations with frequencies above 5% were significantly more oncogenic than wild-type in all assays, mutations occurring at 0.07% to 5.0% included those with and without oncogenic activities that ranged from weak to strong in at least one assay. Proteomic profiling coupled with therapeutic sensitivity assays on PIK3CA variant-expressing cell models revealed variant-specific activation of PI3K signaling as well as other pathways that include the MEK1/2 module of mitogen-activated protein kinase pathway. Our data indicate that cancer treatments will need to increasingly consider the functional relevance of specific mutations in driver genes rather than considering all mutations in drivers as equivalent.
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, Texas.
authors.forename: Turgut
authors.initials: T
authors.lastname: Dogruluk
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, Texas.
authors.forename: Yiu Huen
authors.initials: YH
authors.lastname: Tsang
authors.affiliation: Department of Systems Biology, University of Texas M.D. Anderson Cancer Center, Houston, Texas.
authors.forename: Maribel
authors.initials: M
authors.lastname: Espitia
authors.affiliation: Dan L. Duncan Cancer Center, Baylor College of Medicine, One Baylor Plaza, Houston, Texas. Department of Medicine, Baylor College of Medicine, One Baylor Plaza, Houston, Texas.
authors.forename: Fengju
authors.initials: F
authors.lastname: Chen
authors.affiliation: Department of Bioinformatics and Computational Biology, University of Texas M.D. Anderson Cancer Center, Houston, Texas.
authors.forename: Tenghui
authors.initials: T
authors.lastname: Chen
authors.affiliation: Department of Bioinformatics and Computational Biology, University of Texas M.D. Anderson Cancer Center, Houston, Texas.
authors.forename: Zechen
authors.initials: Z
authors.lastname: Chong
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, Texas. The Human Genome Sequencing Center, Baylor College of Medicine, One Baylor Plaza, Houston, Texas.
authors.forename: Vivek
authors.initials: V
authors.lastname: Appadurai
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, Texas.
authors.forename: Armel
authors.initials: A
authors.lastname: Dogruluk
authors.affiliation: Department of Systems Biology, University of Texas M.D. Anderson Cancer Center, Houston, Texas.
authors.forename: Agna Karina
authors.initials: AK
authors.lastname: Eterovic
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, Texas. The Human Genome Sequencing Center, Baylor College of Medicine, One Baylor Plaza, Houston, Texas.
authors.forename: Penelope E
authors.initials: PE
authors.lastname: Bonnen
authors.affiliation: Dan L. Duncan Cancer Center, Baylor College of Medicine, One Baylor Plaza, Houston, Texas. Department of Medicine, Baylor College of Medicine, One Baylor Plaza, Houston, Texas.
authors.forename: Chad J
authors.initials: CJ
authors.lastname: Creighton
authors.affiliation: Department of Bioinformatics and Computational Biology, University of Texas M.D. Anderson Cancer Center, Houston, Texas.
authors.forename: Ken
authors.initials: K
authors.lastname: Chen
authors.affiliation: Department of Systems Biology, University of Texas M.D. Anderson Cancer Center, Houston, Texas.
authors.forename: Gordon B
authors.initials: GB
authors.lastname: Mills
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Houston, Texas. Dan L. Duncan Cancer Center, Baylor College of Medicine, One Baylor Plaza, Houston, Texas. kls1@bcm.edu.
authors.forename: Kenneth L
authors.initials: KL
authors.lastname: Scott
mesh_terms: D000818:Animals; D001943:Breast Neoplasms / Q000235:genetics*; D045744:Cell Line, Tumor; D058534:Class I Phosphatidylinositol 3-Kinases; D004252:DNA Mutational Analysis / Q000379:methods*; D005260:Female; D064593:Heterografts; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D015151:Immunoblotting; D051379:Mice; D008819:Mice, Nude; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics*; D016133:Polymerase Chain Reaction; D014161:Transduction, Genetic
chemical_list: D019869:Phosphatidylinositol 3-Kinases; D058534:Class I Phosphatidylinositol 3-Kinases; C484760:PIK3CA protein, human