1 Transcriptomically-inferred PI 3 K activity and stemness show a 1 counterintuitive correlation with PIK 3 CA genotype in breast cancer 2 3

A PI3Kα-selective inhibitor has recently been approved for use in breast tumours harbouring mutations in PIK3CA, the gene encoding PI3Kα. Preclinical studies have suggested that the PI3K/AKT/mTORC1 signalling pathway influences stemness, a dedifferentiation-related cellular phenotype associated with aggressive cancer. No direct evidence for such a correlation has been demonstrated to date in human tumours. In two independent human breast cancer cohorts, encompassing nearly 3,000 tumour samples, transcriptional footprint-based analysis uncovered a positive linear association between transcriptionally-inferred PI3K signalling scores and stemness scores. Unexpectedly, stratification of tumours according to PIK3CA genotype revealed a “biphasic” relationship of mutant PIK3CA allele dosage with these scores. Relative to tumour samples without PIK3CA mutations, the presence of a single copy of a hotspot PIK3CA variant was associated with lower PI3K signalling and stemness scores, whereas tumours with multiple copies of PIK3CA hotspot mutations showed higher PI3K signalling and stemness scores. This observation was recapitulated in a human cell model of heterozygous and homozygous PIK3CAH1047R expression. Collectively, our analysis provides evidence for a signalling strength-dependent PI3K-stemness relationship in human breast cancer, which may aid future patient stratification for PI3K-targeted therapies.

[1]  K. Okkenhaug,et al.  PI3K inhibitors are finally coming of age , 2021, Nature Reviews Drug Discovery.

[2]  J. Saez-Rodriguez,et al.  Why do pathway methods work better than they should? , 2020, bioRxiv.

[3]  O. Elemento,et al.  The INPP4B Tumor Suppressor Modulates EGFR Trafficking and Promotes Triple Negative Breast Cancer. , 2020, Cancer discovery.

[4]  S. Miyano,et al.  Landscape and function of multiple mutations within individual oncogenes , 2020, Nature.

[5]  R. Irizarry.  ggplot2 , 2019, Introduction to Data Science.

[6]  Jared L. Johnson,et al.  Double PIK3CA mutations in cis increase oncogenicity and sensitivity to PI3Kα inhibitors , 2019, Science.

[7]  P. Vogt,et al.  PIK3CA Cooperates with KRAS to Promote MYC Activity and Tumorigenesis via the Bromodomain Protein BRD9 , 2019, Cancers.

[8]  Gennady Korotkevich,et al.  Fast gene set enrichment analysis , 2019, bioRxiv.

[9]  Ralitsa R. Madsen,et al.  NODAL/TGFβ signalling mediates the self-sustained stemness induced by PIK3CAH1047R homozygosity in pluripotent stem cells , 2019, Disease Models & Mechanisms.

[10]  H. Rugo,et al.  Alpelisib for PIK3CA‐Mutated, Hormone Receptor–Positive Advanced Breast Cancer , 2019, The New England journal of medicine.

[11]  Bin Liu,et al.  Dynamics of breast cancer relapse reveal late recurring ER-positive genomic subgroups , 2019, Nature.

[12]  A. Intlekofer,et al.  Metabolic signatures of cancer cells and stem cells , 2019, Nature Metabolism.

[13]  C. Sotiriou,et al.  pAKT pathway activation is associated with PIK3CA mutations and good prognosis in luminal breast cancer in contrast to p-mTOR pathway activation , 2019, npj Breast Cancer.

[14]  Ralitsa R. Madsen,et al.  Oncogenic PIK3CA promotes cellular stemness in an allele dose-dependent manner , 2018, Proceedings of the National Academy of Sciences.

[15]  Ralitsa R. Madsen,et al.  Cancer-Associated PIK3CA Mutations in Overgrowth Disorders , 2018, Trends in molecular medicine.

[16]  Ovidiu Lungu,et al.  Consolidation alters motor sequence-specific distributed representations , 2018, bioRxiv.

[17]  Allen W. Zhang,et al.  Cancer stemness, intratumoral heterogeneity, and immune response across cancers , 2018, Proceedings of the National Academy of Sciences.

[18]  Steven J. M. Jones,et al.  Oncogenic Signaling Pathways in The Cancer Genome Atlas. , 2018, Cell.

[19]  Joshua M. Stuart,et al.  Machine Learning Identifies Stemness Features Associated with Oncogenic Dedifferentiation. , 2018, Cell.

[20]  C. Finan,et al.  Linear regression and the normality assumption. , 2017, Journal of clinical epidemiology.

[21]  Icgc,et al.  Pan-cancer analysis of whole genomes , 2017, bioRxiv.

[22]  Lovelace J. Luquette,et al.  A Pan-Cancer Proteogenomic Atlas of PI3K/AKT/mTOR Pathway Alterations. , 2017, Cancer cell.

[23]  N. Blüthgen,et al.  Oncogenic β-catenin and PIK3CA instruct network states and cancer phenotypes in intestinal organoids , 2017, The Journal of cell biology.

[24]  Angela N. Brooks,et al.  A Next Generation Connectivity Map: L1000 Platform and the First 1,000,000 Profiles , 2017, Cell.

[25]  L. Fagnocchi,et al.  Multiple Roles of MYC in Integrating Regulatory Networks of Pluripotent Stem Cells , 2017, Front. Cell Dev. Biol..

[26]  R. Touraine,et al.  Molecular diagnosis of PIK3CA-related overgrowth spectrum (PROS) in 162 patients and recommendations for genetic testing , 2017, Genetics in Medicine.

[27]  M. Garnett,et al.  Perturbation-response genes reveal signaling footprints in cancer gene expression , 2016, bioRxiv.

[28]  J. Shendure,et al.  PIK3CA-associated developmental disorders exhibit distinct classes of mutations with variable expression and tissue distribution , 2016, JCI insight.

[29]  M. Kulesz-Martin,et al.  Overexpression of PIK3CA in murine head and neck epithelium drives tumor invasion and metastasis through PDK1 and enhanced TGFβ signaling , 2015, Oncogene.

[30]  Gianluca Bontempi,et al.  TCGAbiolinks: an R/Bioconductor package for integrative analysis of TCGA data , 2015, Nucleic acids research.

[31]  J. Mesirov,et al.  The Molecular Signatures Database Hallmark Gene Set Collection , 2015 .

[32]  Ken Chen,et al.  Identification of Variant-Specific Functions of PIK3CA by Rapid Phenotyping of Rare Mutations. , 2015, Cancer research.

[33]  N. Socci,et al.  Identifying recurrent mutations in cancer reveals widespread lineage diversity and mutational specificity , 2015, Nature Biotechnology.

[34]  May Yin Lee,et al.  Reactivation of multipotency by oncogenic PIK3CA induces breast tumour heterogeneity , 2015, Nature.

[35]  Michael B. Stadler,et al.  PIK3CAH1047R induces multipotency and multi-lineage mammary tumours , 2015, Nature.

[36]  C. Perou,et al.  Mutant PIK3CA accelerates HER2-driven transgenic mammary tumors and induces resistance to combinations of anti-HER2 therapies , 2013, Proceedings of the National Academy of Sciences.

[37]  Justin Guinney,et al.  GSVA: gene set variation analysis for microarray and RNA-Seq data , 2013, BMC Bioinformatics.

[38]  Bonnie Berger,et al.  A gene expression profile of stem cell pluripotentiality and differentiation is conserved across diverse solid and hematopoietic cancers , 2012, Genome Biology.

[39]  Steven J. M. Jones,et al.  Comprehensive molecular portraits of human breast tumors , 2012, Nature.

[40]  Benjamin E. Gross,et al.  The cBio cancer genomics portal: an open platform for exploring multidimensional cancer genomics data. , 2012, Cancer discovery.

[41]  F. Markowetz,et al.  The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups , 2012, Nature.

[42]  R. Cardiff,et al.  Luminal expression of PIK3CA mutant H1047R in the mammary gland induces heterogeneous tumors. , 2011, Cancer research.

[43]  Stuart H. Orkin,et al.  A Myc Network Accounts for Similarities between Embryonic Stem and Cancer Cell Transcription Programs , 2010, Cell.

[44]  Adrian V. Lee,et al.  Proteomic and transcriptomic profiling reveals a link between the PI3K pathway and lower estrogen-receptor (ER) levels and activity in ER+ breast cancer , 2010, Breast Cancer Research.

[45]  Paul Ellis,et al.  PIK3CA mutations associated with gene signature of low mTORC1 signaling and better outcomes in estrogen receptor–positive breast cancer , 2010, Proceedings of the National Academy of Sciences.

[46]  M. Robinson,et al.  A scaling normalization method for differential expression analysis of RNA-seq data , 2010, Genome Biology.

[47]  Robert L Sutherland,et al.  PI3K pathway activation in breast cancer is associated with the basal‐like phenotype and cancer‐specific mortality , 2010, International journal of cancer.

[48]  S. O'toole,et al.  PI3K Pathway Activation in Breast Cancer Is Associated with the Basal-Like Phenotype and Cancer-Specific Mortality. , 2009 .

[49]  J. Watters,et al.  Can phenotypic pathway signatures improve the prediction of response to PI3K pathway inhibitors , 2009 .

[50]  L. Cantley,et al.  PI3K pathway alterations in cancer: variations on a theme , 2008, Oncogene.

[51]  R. Shamir,et al.  Regulatory networks define phenotypic classes of human stem cell lines , 2008, Nature.

[52]  Zhi Hu,et al.  An integrative genomic and proteomic analysis of PIK3CA, PTEN, and AKT mutations in breast cancer. , 2008, Cancer research.

[53]  A. Regev,et al.  An embryonic stem cell–like gene expression signature in poorly differentiated aggressive human tumors , 2008, Nature Genetics.

[54]  C. Creighton,et al.  A gene transcription signature of the Akt/mTOR pathway in clinical breast tumors , 2007, Oncogene.

[55]  L. Skoog,et al.  PIK3CA Mutations and PTEN Loss Correlate with Similar Prognostic Factors and Are Not Mutually Exclusive in Breast Cancer , 2007, Clinical Cancer Research.

[56]  Paul A Clemons,et al.  The Connectivity Map: Using Gene-Expression Signatures to Connect Small Molecules, Genes, and Disease , 2006, Science.

[57]  Steven J. M. Jones,et al.  Comprehensive molecular portraits of human breast tumours , 2013 .