bims-pideca Biomed News
on Class IA PI3K signalling in development and cancer
Issue of 2021‒08‒08
fourteen papers selected by
Ralitsa Radostinova Madsen
University College London Cancer Institute

  1. Breast Cancer Res. 2021 Aug 03. 23(1): 81
      BACKGROUND: HER2-amplified breast cancer is a clinically defined subtype of breast cancer for which there are multiple viable targeted therapies. Resistance to these targeted therapies is a common problem, but the mechanisms by which resistance occurs remain incompletely defined. One mechanism that has been proposed is through mutation of genes in the PI3-kinase pathway. Intracellular signaling from the HER2 pathway can occur through PI3-kinase, and mutations of the encoding gene PIK3CA are known to be oncogenic. Mutations in PIK3CA co-occur with HER2-amplification in ~ 20% of cases within the HER2-amplified subtype.METHODS: We generated isogenic knockin mutants of each PIK3CA hotspot mutation in HER2-amplified breast cancer cells using adeno-associated virus-mediated gene targeting. Isogenic clones were analyzed using a combinatorial drug screen to determine differential responses to HER2-targeted therapy. Western blot analysis and immunofluorescence uncovered unique intracellular signaling dynamics in cells resistant to HER2-targeted therapy. Subsequent combinatorial drug screens were used to explore neuregulin-1-mediated resistance to HER2-targeted therapy. Finally, results from in vitro experiments were extrapolated to publicly available datasets.
    RESULTS: Treatment with HER2-targeted therapy reveals that mutations in the kinase domain (H1047R) but not the helical domain (E545K) increase resistance to lapatinib. Mechanistically, sustained AKT signaling drives lapatinib resistance in cells with the kinase domain mutation, as demonstrated by staining for the intracellular product of PI3-kinase, PIP3. This resistance can be overcome by co-treatment with an inhibitor to the downstream kinase AKT. Additionally, knockout of the PIP3 phosphatase, PTEN, phenocopies this result. We also show that neuregulin-1, a ligand for HER-family receptors, confers resistance to cells harboring either hotspot mutation and modulates response to combinatorial therapy. Finally, we show clinical evidence that the hotspot mutations have distinct expression profiles related to therapeutic resistance through analysis of TCGA and METABRIC data cohorts.
    CONCLUSION: Our results demonstrate unique intracellular signaling differences depending on which mutation in PIK3CA the cell harbors. Only mutations in the kinase domain fully activate the PI3-kinase signaling pathway and maintain downstream signaling in the presence of HER2 inhibition. Moreover, we show there is potentially clinical importance in understanding both the PIK3CA mutational status and levels of neuregulin-1 expression in patients with HER2-amplified breast cancer treated with targeted therapy and that these problems warrant further pre-clinical and clinical testing.
  2. Structure. 2021 Jul 30. pii: S0969-2126(21)00252-5. [Epub ahead of print]
      There is considerable interest in developing antibodies as modulators of signaling pathways. One of the most important signaling pathways in higher eukaryotes is the phosphoinositide 3-kinase (PI3K) pathway, which plays fundamental roles in growth, metabolism, and immunity. The class IB PI3K, PI3Kγ, is a heterodimeric complex composed of a catalytic p110γ subunit bound to a p101 or p84 regulatory subunit. PI3Kγ is a critical component in multiple immune signaling processes and is dependent on activation by Ras and G protein-coupled receptors (GPCRs) to mediate its cellular roles. Here we describe the rapid and efficient characterization of multiple PI3Kγ binding single-chain camelid nanobodies using hydrogen-deuterium exchange (HDX) mass spectrometry (MS) for structural and biochemical studies. We identify nanobodies that stimulated lipid kinase activity, block Ras activation, and specifically inhibited p101-mediated GPCR activation. Overall, our work reveals insight into PI3Kγ regulation and identifies sites that may be exploited for therapeutic development.
    Keywords:  HDX-MS; PI3K; PI3Kγ; PIK3CG; hydrogen exchange; nanobodies; p110; p110γ; phosphoinositide 3-kinases
  3. Front Pediatr. 2021 ;9 697706
      Activated phosphoinositide 3-kinase delta syndrome (APDS), caused by mutations in PI3Kδ catalytic p110δ (PIK3CD) or regulatory p85α (PIK3R1) subunits, is a primary immunodeficiency affecting both humoral and cellular immunity, which shares some phenotypic similarities with hyper-IgM syndromes and common variable immunodeficiency (CVID). Since its first description in 2013, over 200 patients have been reported worldwide. Unsurprisingly, many of the newly diagnosed patients were recruited later in life from previously long-standing unclassified immunodeficiencies and the early course of the disease is, therefore, often less well-described. In this study, we report clinical and laboratory features of eight patients followed for APDS, with particular focus on early warning signs, longitudinal development of their symptoms, individual variations, and response to therapy. The main clinical features shared by our patients included recurrent bacterial and viral respiratory tract infections, gastrointestinal disease, non-malignant lymphoproliferation, autoimmune thyroiditis, and susceptibility to EBV. All patients tolerated vaccination with both attenuated live and subunit vaccines with no adverse effects, although some failed to mount adequate antibody response. Laboratory findings were characterized by dysgammaglobulinaemia, elevated serum IgM, block in B-cell maturation with high transitional B cells, and low naïve T cells with CD8 T-cell activation. All patients benefited from immunoglobulin replacement therapy, whereas immunosuppression with mTOR pathway inhibitors was only partially successful. Therapy with specific PI3K inhibitor leniolisib was beneficial in all patients in the clinical trial. These vignettes, summary data, and particular tell-tale signs should serve to facilitate early recognition, referral, and initiation of outcome-improving therapy.
    Keywords:  APDS; PI3K; activated phosphoinositide 3-kinase delta syndrome; immunodeficiency; immunoglobulins; infection; lymphoproliferation
  4. J Clin Invest. 2021 Aug 03. pii: 146415. [Epub ahead of print]
      Decreased skeletal muscle strength and mitochondrial dysfunction are characteristic of diabetes. Action of insulin and IGF-1 through insulin receptor (IR) and IGF-1 receptor (IGF1R) maintain muscle mass via suppression of FoxOs, but whether FoxO activation coordinates atrophy in concert with mitochondrial dysfunction is unknown. We show that mitochondrial respiration and complex-I activity were decreased in streptozotocin (STZ) diabetic muscle, but these defects were reversed following muscle-specific FoxO1/3/4 triple knockout in STZ-FoxO TKO. In the absence of systemic glucose or lipid abnormalities, muscle-specific IR knockout (M-IR-/-) or combined IR/IGF1R knockout (MIGIRKO) impaired mitochondrial respiration, decreased ATP production, and increased ROS. These mitochondrial abnormalities were not present in muscle-specific IR/IGF1R and FoxO1/3/4 quintuple knockout mice (M-QKO). Acute tamoxifen-inducible deletion of IR/IGF1R also decreased muscle pyruvate respiration, complex-I activity, and supercomplex assembly. Although autophagy was increased when IR/IGF1R were deleted in muscle, mitophagy was not increased. Mechanistically, RNA-seq revealed that complex-I core subunits were decreased in STZ-diabetic and MIGIRKO muscle, and these changes were not present with FoxO knockout in STZ-FoxO TKO and M-QKO. Thus, insulin-deficient diabetes or loss of insulin/IGF-1 action in muscle decreases complex-I driven mitochondrial respiration and supercomplex assembly, in part by FoxO-mediated repression of Complex-I subunit expression.
    Keywords:  Endocrinology; Insulin; Mitochondria; Muscle
  5. Nat Rev Cancer. 2021 Aug 02.
      Translational control of mRNAs during gene expression allows cells to promptly and dynamically adapt to a variety of stimuli, including in neoplasia in response to aberrant oncogenic signalling (for example, PI3K-AKT-mTOR, RAS-MAPK and MYC) and microenvironmental stress such as low oxygen and nutrient supply. Such translational rewiring allows rapid, specific changes in the cell proteome that shape specific cancer phenotypes to promote cancer onset, progression and resistance to anticancer therapies. In this Review, we illustrate the plasticity of mRNA translation. We first highlight the diverse mechanisms by which it is regulated, including by translation factors (for example, eukaryotic initiation factor 4F (eIF4F) and eIF2), RNA-binding proteins, tRNAs and ribosomal RNAs that are modulated in response to aberrant intracellular pathways or microenvironmental stress. We then describe how translational control can influence tumour behaviour by impacting on the phenotypic plasticity of cancer cells as well as on components of the tumour microenvironment. Finally, we highlight the role of mRNA translation in the cellular response to anticancer therapies and its promise as a key therapeutic target.
  6. J Vasc Surg Venous Lymphat Disord. 2021 Aug 03. pii: S2213-333X(21)00391-7. [Epub ahead of print]
      BACKGROUND: PIK3CA-related overgrowth syndromes (PROS) include a variety of clinical presentations that are associated with hypertrophy of different parts of the body.AIM: Perform a systematic literature review to assess the current treatment options and their efficacy and safety in PROS.
    METHODS: A literature search was performed in EMBASE, MEDLINE (Ovid), Web of Science Core Collection, Cochrane Central Register of Controlled Trials, and Google Scholar to retrieve publications on the treatment for hypertrophy in PROS and randomized controlled trials, cohort studies or case series including ≥10 patients reporting were included in the review. Titles, abstracts and full texts were assessed by two reviewers independently. The Risk of Bias (RoB) was assessed using the Newcastle Ottawa Scale.
    RESULTS: 16 articles for the treatment of hypertrophy in PROS patients were included, 13 (81.3%) from clinical retrospective studies and 3 (13.7%) from prospective cohort studies. The ROB grade was low for 2, medium for 12 and high for 2 studies. 13 articles reported surgical treatment, while 3 reported pharmacological treatment using PIK3/mTOR pathway inhibitors in PROS patients. In 3 studies, PROS was defined by a mutation in the PIK3CA gene, while the other studies relied on a clinical definition of PROS. Surgical therapy was beneficial for a specific subgroup of PROS (macrodactyly), but little was reported concerning surgery and potential benefits in other PROS entities. Reported side effects in surgical therapy were mostly prolonged wound healing or scarring. PIK3/mTOR pathway inhibition was beneficial in patients with PROS reducing hypertrophy as well as systemic symptoms. Adverse effects reported included infection, changes in blood count, liver enzymes and metabolic measures.
    CONCLUSION: Surgery is a locally limited treatment option in specific types of PROS. A promising treatment option in PROS is the pharmacological PIK3CA inhibition. However, the level of evidence on treatment of overgrowth in PROS patients is limited.
    Keywords:  Phosphatidylinositol 3-Kinase; Phosphoinositide-3 Kinase Inhibitors; TOR Serine-Threonine Kinases
  7. Nat Commun. 2021 08 03. 12(1): 4670
      Spontaneous canine cancers are valuable but relatively understudied and underutilized models. To enhance their usage, we reanalyze whole exome and genome sequencing data published for 684 cases of >7 common tumor types and >35 breeds, with rigorous quality control and breed validation. Our results indicate that canine tumor alteration landscape is tumor type-dependent, but likely breed-independent. Each tumor type harbors major pathway alterations also found in its human counterpart (e.g., PI3K in mammary tumor and p53 in osteosarcoma). Mammary tumor and glioma have lower tumor mutational burden (TMB) (median < 0.5 mutations per Mb), whereas oral melanoma, osteosarcoma and hemangiosarcoma have higher TMB (median ≥ 1 mutations per Mb). Across tumor types and breeds, TMB is associated with mutation of TP53 but not PIK3CA, the most mutated genes. Golden Retrievers harbor a TMB-associated and osteosarcoma-enriched mutation signature. Here, we provide a snapshot of canine mutations across major tumor types and breeds.
  8. Genes (Basel). 2021 Jul 19. pii: 1094. [Epub ahead of print]12(7):
      Ras proteins are essential mediators of a multitude of cellular processes, and its deregulation is frequently associated with cancer appearance, progression, and metastasis. Ras-driven cancers are usually aggressive and difficult to treat. Although the recent Food and Drug Administration (FDA) approval of the first Ras G12C inhibitor is an important milestone, only a small percentage of patients will benefit from it. A better understanding of the context in which Ras operates in different tumor types and the outcomes mediated by each effector pathway may help to identify additional strategies and targets to treat Ras-driven tumors. Evidence emerging in recent years suggests that both oncogenic Ras signaling in tumor cells and non-oncogenic Ras signaling in stromal cells play an essential role in cancer. PI3K is one of the main Ras effectors, regulating important cellular processes such as cell viability or resistance to therapy or angiogenesis upon oncogenic Ras activation. In this review, we will summarize recent advances in the understanding of Ras-dependent activation of PI3K both in physiological conditions and cancer, with a focus on how this signaling pathway contributes to the formation of a tumor stroma that promotes tumor cell proliferation, migration, and spread.
    Keywords:  PI3-Kinase; Ras oncogenes
  9. Elife. 2021 Aug 04. pii: e71610. [Epub ahead of print]10
      Changes in MAPK signaling allow lung cancer cells to transition between lineages that respond differently to treatment.
    Keywords:  ERK signaling; cancer biology; human; lineage switching; lineage transformation; lung cancer
  10. Cell Rep Med. 2021 Jul 20. 2(7): 100350
      Inhibition of the extracellular signal-regulated kinases ERK1 and ERK2 (ERK1/2) offers a promising therapeutic strategy in cancers harboring activated RAS/RAF/MEK/ERK signaling pathways. Here, we describe an orally bioavailable and selective ERK1/2 inhibitor, ASN007, currently in clinical development for the treatment of cancer. In preclinical studies, ASN007 shows strong antiproliferative activity in tumors harboring mutations in BRAF and RAS (KRAS, NRAS, and HRAS). ASN007 demonstrates activity in a BRAFV600E mutant melanoma tumor model that is resistant to BRAF and MEK inhibitors. The PI3K inhibitor copanlisib enhances the antiproliferative activity of ASN007 both in vitro and in vivo due to dual inhibition of RAS/MAPK and PI3K survival pathways. Our data provide a rationale for evaluating ASN007 in RAS/RAF-driven tumors as well as a mechanistic basis for combining ASN007 with PI3K inhibitors.
    Keywords:  ASN007; ERK; KRAS; PI3K; RAF/RAS-driven cancers; biomarker; combinational therapy; kinase inhibitor; lymphoma; solid tumors
  11. J Cell Sci. 2021 Aug 01. pii: jcs254920. [Epub ahead of print]134(15):
      Elongation of vascular endothelial cells (ECs) is an important process in angiogenesis; however, the molecular mechanisms remain unknown. The actin-crosslinking protein TAGLN (transgelin, also known as SM22 or SM22α) is abundantly expressed in smooth muscle cells (SMCs) and is widely used as a canonical marker for this cell type. In the course of studies using mouse embryonic stem cells (ESCs) carrying an Tagln promoter-driven fluorescence marker, we noticed activation of the Tagln promoter during EC elongation. Tagln promoter activation co-occurred with EC elongation in response to vascular endothelial growth factor A (VEGF-A). Inhibition of phosphoinositide 3-kinase (PI3K)-Akt signaling and mTORC1 also induced EC elongation and Tagln promoter activation. Human umbilical vein endothelial cells (HUVECs) elongated, activated the TAGLN promoter and increased TAGLN transcripts in an angiogenesis model. Genetic disruption of TAGLN augmented angiogenic behaviors of HUVECs, as did the disruption of TAGLN2 and TAGLN3 genes. Tagln expression was found in ECs in mouse embryos. Our results identify TAGLN as a putative regulator of angiogenesis whose expression is activated in elongating ECs. This finding provides insight into the cytoskeletal regulation of EC elongation and an improved understanding of the molecular mechanisms underlying the regulation of angiogenesis.
    Keywords:  Angiogenesis; Cell elongation; Endothelial cells; TAGLN
  12. Cancers (Basel). 2021 Jul 30. pii: 3850. [Epub ahead of print]13(15):
      Insulin-like growth factor-1 receptor (IGF-1R), an important factor in promoting cancer cell growth and survival, is commonly upregulated in cancer cells. However, amplification of the IGF1R gene is extremely rare in tumors. Here, we have provided insights into the mechanisms underlying the regulation of IGF-1R protein expression. We found that PKM2 serves as a non-metabolic protein that binds to and increases IGF-1R protein expression by promoting the interaction between IGF-1R and heat-shock protein 90 (HSP90). PKM2 depletion decreases HSP90 binding to IGF-1R precursor, thereby reducing IGF-1R precursor stability and the basal level of mature IGF-1R. Consequently, PKM2 knockdown inhibits the activation of AKT, the key downstream effector of IGF-1R signaling, and increases apoptotic cancer cell death during hypoxia. Notably, we clinically verified the PKM2-regulated expression of IGF-1R through immunohistochemical staining in a tissue microarray of 112 lung cancer patients, demonstrating a significant positive correlation (r = 0.5208, p < 0.0001) between PKM2 and IGF-1R expression. Together, the results of a previous report demonstrated that AKT mediates PKM2 phosphorylation at serine-202; these results suggest that IGF-1R signaling and PKM2 mutually regulate each other to facilitate cell growth and survival, particularly under hypoxic conditions, in solid tumors with dysregulated IGF-1R expression.
    Keywords:  HSP90; IGF-1R; PKM2; hypoxia; survival
  13. Nat Commun. 2021 08 03. 12(1): 4688
      Internalization and intracellular trafficking of G protein-coupled receptors (GPCRs) play pivotal roles in cell responsiveness. Dysregulation in receptor trafficking can lead to aberrant signaling and cell behavior. Here, using an endosomal BRET-based assay in a high-throughput screen with the prototypical GPCR angiotensin II type 1 receptor (AT1R), we sought to identify receptor trafficking inhibitors from a library of ~115,000 small molecules. We identified a novel dual Ras and ARF6 inhibitor, which we named Rasarfin, that blocks agonist-mediated internalization of AT1R and other GPCRs. Rasarfin also potently inhibits agonist-induced ERK1/2 signaling by GPCRs, and MAPK and Akt signaling by EGFR, as well as prevents cancer cell proliferation. In silico modeling and in vitro studies reveal a unique binding modality of Rasarfin within the SOS-binding domain of Ras. Our findings unveil a class of dual small G protein inhibitors for receptor trafficking and signaling, useful for the inhibition of oncogenic cellular responses.