The PI3K/AKT signaling pathway is one of the most frequently hyperactivated oncogenic axes in human biology, activated in approximately 30% of all solid tumors and implicated in cancer, metabolic disease, and immune dysfunction. Understanding how phosphatidylinositol 3-kinase (PI3K) converts extracellular growth signals into AKT-mediated pro-survival programs — and how mutations in PIK3CA and PTEN deregulate this circuit — is fundamental to modern cancer biology and to the rational design of targeted inhibitors now entering clinical practice.
PI3K: Classes, Isoforms, and Activation
Phosphoinositide 3-kinases are a family of lipid kinases grouped into three classes based on structure, substrate specificity, and regulation. Class I PI3Ks are the primary drivers of AKT signaling and are subdivided into two subclasses:
- Class IA: heterodimers of a catalytic p110 subunit (p110α encoded by PIK3CA, p110β by PIK3CB, or p110δ by PIK3CD) and a regulatory p85 subunit (p85α, p55α, p50α, p85β, or p55γ). The p110α and p110β isoforms are ubiquitously expressed; p110δ expression is enriched in leukocytes.
- Class IB: a single catalytic subunit, p110γ (encoded by PIK3CG), paired with a p101 or p84 regulatory subunit. p110γ is activated principally by G protein-coupled receptors (GPCRs) via Gβγ subunits and is highly expressed in myeloid cells.
Upon receptor tyrosine kinase (RTK) activation — by insulin, EGF, PDGF, or other growth factors — the p85 regulatory subunit binds phosphotyrosine motifs on receptor scaffolds, relieving its inhibitory constraint on p110 and recruiting the heterodimer to the plasma membrane. RAS GTPases provide a parallel activation input to Class IA p110 subunits. Once activated, Class I PI3K phosphorylates the 3′-hydroxyl of phosphatidylinositol-4,5-bisphosphate (PIP2) to generate phosphatidylinositol-3,4,5-trisphosphate (PIP3). PIP3 acts as a second messenger, recruiting pleckstrin homology (PH) domain-containing effectors — most critically AKT — to the inner leaflet of the plasma membrane.
Class II and Class III PI3Ks serve distinct intracellular roles. Class II PI3Ks (PI3K-C2α, -C2β, -C2γ) generate phosphatidylinositol-3-phosphate (PI3P) and PI3,4P2, involved in endosomal trafficking and autophagy. Class III PI3K (VPS34) regulates autophagosome nucleation and vesicle sorting. Neither class is directly linked to the AKT/mTOR oncogenic axis.
Of the Class I isoforms, PIK3CA (p110α) is by far the most commonly mutated in human cancer, making it a priority research target. Lewis Cantley's foundational work characterizing PI3K activity and its transforming potential laid the groundwork for the oncology field now built around this pathway Cantley, Science 2002.
AKT: Isoforms, Activation, and Downstream Targets
AKT (also called Protein Kinase B, PKB) is a serine/threonine kinase and the principal downstream effector of PIP3. Three highly homologous isoforms exist in mammals:
- AKT1 (PKBα) — ubiquitously expressed; the dominant isoform in most epithelial cells; promotes cell survival and growth.
- AKT2 (PKBβ) — enriched in insulin-responsive tissues (liver, muscle, adipose); plays a major role in glucose metabolism and insulin signaling.
- AKT3 (PKBγ) — preferentially expressed in brain and testes; contributes to neural development and brain tumor biology.
Full AKT activation requires two sequential phosphorylation events. PIP3 generated by PI3K recruits AKT to the plasma membrane via its PH domain, where 3-phosphoinositide-dependent protein kinase 1 (PDK1) phosphorylates AKT at Thr308 in the activation loop. This event alone yields partial kinase activity. Full activation requires phosphorylation at Ser473 in the hydrophobic motif by the mTOR complex 2 (mTORC2). Both phosphorylations are required for maximal AKT catalytic output and substrate engagement Manning & Toker, Cell 2017.
Once fully activated, AKT phosphorylates a broad substrate network governing nearly every hallmark of cancer:
| Substrate | AKT Phosphorylation Site | Functional Consequence |
|---|---|---|
| GSK3β | Ser9 | Inhibition → increased glycogen synthesis; stabilization of β-catenin and Cyclin D1 |
| MDM2 | Ser166 | Nuclear translocation → p53 ubiquitination and degradation → evasion of apoptosis |
| FOXO transcription factors | Multiple Ser/Thr | Cytoplasmic sequestration via 14-3-3 binding → suppression of pro-apoptotic and cell-cycle-arrest gene programs |
| BAD | Ser136 | Dissociation from BCL-XL/BCL-2 → suppression of cytochrome c release → anti-apoptosis |
| TSC2 (Tuberin) | Multiple sites | Inhibition of TSC1/TSC2 complex → RHEB-GTP accumulation → mTORC1 activation → protein synthesis and cell growth |
| Caspase-9 | Ser196 | Inhibition of intrinsic apoptosis pathway → enhanced cell survival |
PTEN: The Key Brake on PI3K/AKT Signaling
Phosphatase and tensin homolog (PTEN) is the principal negative regulator of PI3K/AKT signaling and, after TP53, the most frequently inactivated tumor suppressor in human cancer. PTEN is a dual-specificity phosphatase that dephosphorylates both lipid and protein substrates, but its tumor-suppressive function is primarily exerted through its lipid phosphatase activity: PTEN converts PIP3 back to PIP2, directly opposing the action of PI3K and preventing AKT membrane recruitment and activation.
PTEN was first identified by Steck et al. in 1997 as a gene mapping to chromosome 10q23, deleted in multiple advanced cancers Steck et al., Nature Genetics 1997. The following year, Maehama and Dixon demonstrated that PTEN catalyzes the dephosphorylation of PIP3 at the 3-position, directly linking PTEN loss to constitutive PI3K/AKT activation Maehama & Dixon, JBC 1998.
PTEN loss of function occurs through multiple mechanisms in cancer: homozygous deletion (common in glioblastoma, prostate cancer), somatic point mutation, promoter methylation, post-translational degradation via NEDD4-1 ubiquitin ligase, and microRNA-mediated suppression. Germline PTEN mutations cause Cowden syndrome (OMIM #158350), a hamartoma tumor syndrome with elevated lifetime risks of breast, thyroid, endometrial, and colorectal cancers.
Beyond its cytoplasmic lipid phosphatase role, PTEN also localizes to the nucleus, where it maintains chromosomal integrity, regulates DNA damage repair via interaction with RAD51, and controls centromere stability. These nuclear functions are independent of lipid phosphatase activity and provide additional layers of tumor suppression.
PI3K/AKT Pathway Activation in Cancer and Therapeutic Inhibitors
Aberrant activation of the PI3K/AKT signaling pathway occurs through two primary routes in cancer: gain-of-function mutations in PIK3CA and loss-of-function alterations in PTEN.
PIK3CA mutations cluster at two hotspots in the p110α protein: the helical domain (E542K and E545K, encoded by exon 9) and the kinase domain (H1047R, encoded by exon 20). Both classes produce constitutively active PI3K that generates elevated PIP3 independently of upstream RTK input. PIK3CA mutations are among the most prevalent driver mutations across solid tumors: present in approximately 25–40% of HR-positive breast cancers, ~30% of endometrial carcinomas, and at lower but significant frequencies in colorectal cancer, head and neck squamous cell carcinoma, and bladder cancer Lawrence et al., Nature 2014.
PTEN loss is a hallmark of prostate cancer (~40% of tumors), glioblastoma multiforme (~40%), and endometrial carcinoma, and co-occurs with PIK3CA mutations in a subset of breast and colorectal tumors, compounding pathway activation.
PI3K inhibitors in clinical use:
- Alpelisib (Piqray, Novartis) — a selective PI3Kα (p110α) inhibitor, FDA-approved in May 2019 for postmenopausal women and men with PIK3CA-mutated, HR+/HER2− advanced breast cancer, in combination with fulvestrant.
- Idelalisib (Zydelig, Gilead) — a selective PI3Kδ inhibitor approved for relapsed chronic lymphocytic leukemia (CLL), follicular lymphoma, and small lymphocytic lymphoma. PI3Kδ is required for B-cell receptor signaling in B-cell malignancies.
- Copanlisib (Aliqopa, Bayer) — a pan-Class I PI3K inhibitor with predominant activity against PI3Kα and PI3Kδ, approved for relapsed follicular lymphoma.
AKT inhibitors:
- Capivasertib (Truqap, AstraZeneca) — a pan-AKT inhibitor (AKT1/2/3), FDA-approved in November 2023 in combination with fulvestrant for HR+/HER2− advanced breast cancer harboring PIK3CA, AKT1, or PTEN alterations.
- Ipatasertib (Roche) — a pan-AKT inhibitor in Phase III trials across breast, prostate, and other solid tumors.
Beyond oncology, PI3Kδ plays a critical role in T-cell and B-cell activation, making selective PI3Kδ inhibition a strategy in autoimmune disease. PI3Kγ, highly expressed in macrophages and neutrophils, regulates myeloid cell chemotaxis and inflammatory responses — and is being explored as a therapeutic target in inflammatory conditions and as an immunomodulatory strategy to reprogram tumor-associated macrophages.
Research Tools for PI3K/AKT Pathway Studies at BioHippo
Dissecting PI3K/AKT signaling at the bench requires reliable phosphorylation-state reagents and quantitative assay kits. BioHippo offers a curated portfolio of ELISA kits and primary antibodies for PI3K/AKT pathway components:
- Human Phospho-AKT (S473) ELISA Kit (ELK0792) — quantifies the active, mTORC2-phosphorylated form of AKT in cell lysates and tissue homogenates. A direct readout of pathway activation state in human samples.
- Mouse Phospho-AKT (S473) ELISA Kit (ELK0791) — validated for murine tumor models, enabling quantitative p-AKT measurement in syngeneic and xenograft studies.
- Human PTEN ELISA Kit (ELK1367) — quantifies total PTEN protein in serum, plasma, cell lysates, and tissue homogenates; supports studies of PTEN expression loss in cancer cell lines and clinical samples.
- Human PI3K ELISA Kit (ELK9879) — measures total PI3K protein levels in serum, plasma, and tissue homogenates; useful for expression-level comparisons across tumor types.
- Human PIK3CA ELISA Kit (EH1594) — isoform-specific quantification of p110α, the most frequently mutated PI3K subunit in cancer.
- Human mTOR ELISA Kit (ELK9237) — measures the downstream effector mTOR, enabling pathway profiling from PI3K through AKT to mTORC1.
Browse the full ELISA kit collection or search by target to find validated kits for additional PI3K/AKT pathway components including TSC2, GSK3β, FOXO, and S6K1.
Frequently Asked Questions
What is the PI3K/AKT signaling pathway?
The PI3K/AKT signaling pathway is an intracellular signal transduction cascade that transmits growth factor signals from cell-surface receptors to the nucleus, controlling cell growth, survival, proliferation, and metabolism. PI3K phosphorylates the plasma membrane lipid PIP2 to generate PIP3, which recruits and activates the serine/threonine kinase AKT. Activated AKT then phosphorylates a broad network of substrates that collectively promote cell survival, suppress apoptosis, stimulate protein synthesis, and drive cell-cycle progression. The pathway is one of the most frequently activated oncogenic signaling axes in human cancer, present in approximately 30% of solid tumors.
What does AKT do in the cell?
AKT is a master serine/threonine kinase that integrates PI3K-derived PIP3 signals into a diverse program of cellular responses. Its principal functions include: (1) suppressing apoptosis by phosphorylating and inactivating BAD, Caspase-9, and FOXO transcription factors; (2) promoting cell survival by phosphorylating MDM2 (Ser166), which targets the tumor suppressor p53 for degradation; (3) stimulating protein synthesis and cell growth by activating mTORC1 via TSC2 phosphorylation; (4) enhancing glucose uptake and glycogen synthesis by inhibiting GSK3β (Ser9); and (5) driving cell-cycle entry by stabilizing Cyclin D1 and inhibiting p27. The three AKT isoforms — AKT1, AKT2, and AKT3 — share these core functions but have distinct tissue distributions and can have non-overlapping, sometimes opposing, roles in specific cancer contexts.
What is the role of PTEN in PI3K/AKT signaling?
PTEN (phosphatase and tensin homolog) acts as the primary brake on PI3K/AKT signaling by catalyzing the dephosphorylation of PIP3 back to PIP2, directly opposing PI3K. By reducing cellular PIP3 levels, PTEN prevents AKT membrane recruitment and activation, keeping the pathway in check. Loss of PTEN function — through deletion, mutation, promoter methylation, or protein degradation — results in constitutively elevated PIP3 and persistent AKT activation even in the absence of upstream growth factor signals. This is why PTEN is one of the most frequently mutated tumor suppressor genes in human cancer: its loss effectively phenocopies constitutively active PI3K.
How is the PI3K/AKT pathway activated in cancer?
The PI3K/AKT pathway becomes constitutively active in cancer through several mechanisms: (1) PIK3CA gain-of-function mutations at hotspots E542K/E545K (helical domain) and H1047R (kinase domain), which account for the majority of oncogenic PIK3CA alterations; (2) loss-of-function alterations in PTEN (deletion, mutation, methylation), removing the primary negative regulator; (3) amplification or activating mutations of upstream RTKs (e.g., EGFR, HER2/ERBB2, FGFR); (4) activating AKT1 mutations (e.g., E17K); and (5) overactivation of RAS, which directly activates Class IA PI3K catalytic subunits. Any one of these alterations can decouple the pathway from normal growth factor-dependent control.
What are PI3K inhibitors and which are FDA-approved?
PI3K inhibitors are small-molecule kinase inhibitors that competitively block the ATP-binding site of PI3K catalytic subunits, preventing PIP3 generation and downstream AKT activation. Isoform-selective inhibitors have improved the therapeutic window compared to pan-PI3K agents. FDA-approved PI3K inhibitors include: alpelisib (Piqray), a selective PI3Kα inhibitor approved in 2019 for PIK3CA-mutated HR+/HER2− breast cancer; idelalisib (Zydelig), a selective PI3Kδ inhibitor approved for B-cell malignancies including CLL and follicular lymphoma; and copanlisib (Aliqopa), a pan-PI3K inhibitor with predominant PI3Kα/δ activity approved for follicular lymphoma. Resistance to PI3K inhibitors remains a major clinical challenge, driven by feedback reactivation of RTKs, RAS, and mTOR.
What is the difference between PI3K, AKT, and mTOR?
PI3K, AKT, and mTOR are three distinct kinases that function as sequential nodes in the same signaling cascade. PI3K is a lipid kinase activated at the plasma membrane by RTKs and RAS; it generates the second messenger PIP3. AKT is a serine/threonine protein kinase recruited and activated by PIP3; it is the primary effector that phosphorylates a broad substrate network. mTOR (mechanistic target of rapamycin) is a serine/threonine kinase that exists in two complexes: mTORC1 (rapamycin-sensitive; promotes protein synthesis via S6K1 and 4E-BP1) and mTORC2 (rapamycin-insensitive; phosphorylates AKT at Ser473 to complete its activation). The three enzymes are therefore ordered: PI3K → PIP3 → AKT → mTORC1. mTORC2 sits upstream of AKT as a co-activating kinase. This interconnection means that inhibiting any one node can trigger compensatory feedback — for example, mTORC1 inhibition with rapamycin relieves negative feedback on RTKs, paradoxically increasing PI3K and AKT activity. For a detailed treatment of mTOR signaling, see our article on the mTOR pathway and cell growth regulation.