RBL2/p130 is a direct AKT target and is required to induce apoptosis upon AKT inhibition in lung cancer and mesothelioma cell lines

Our plain-English summary of RBL2/p130 is a direct AKT target and is required to induce apoptosis upon AKT inhibition in lung cancer and mesothelioma cell lines, by F. Pentimalli, I.M. Forte, L. Esposito, et al (Oncogene, 2018).

The study

• Authors: F. Pentimalli, I.M. Forte, L. Esposito, P. Indovina, C.A. Iannuzzi, L. Alfano, C. Costa, D. Barone, G. Rocco, A. Giordano
• Journal: Oncogene
• Published: 2018;37(27):3657-3671
• Cited by: 29 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

The retinoblastoma (RB) protein family includes RB1/p105, RBL1/p107, and RBL2/p130, which are key factors in cell-cycle regulation and stand at the crossroads of multiple pathways dictating cell fate decisions. The role of RB proteins in apoptosis is controversial because they can inhibit or promote apoptosis depending on the context, on the apoptotic stimuli and on their intrinsic status, impacting on the response to antitumoral treatments. Here we identified RBL2/p130 as a direct substrate of the AKT kinase, a key antiapoptotic factor hyperactive in multiple cancer types. We showed that RBL2/p130 and AKT1 physically interact and AKT phosphorylates RBL2/p130 Ser941, located in the pocket domain, but not when this residue is mutated into Ala. We found that pharmacological inhibition of AKT, through the highly selective AKT inhibitor VIII (AKTiVIII), impairs RBL2/p130 Ser941 phosphorylation and increases RBL2/p130 stability, mRNA expression and nuclear levels in both lung cancer and mesothelioma cell lines, mirroring the more extensively studied effects on the p27 cell-cycle inhibitor. Consistently, AKT inhibition reduced cell viability, induced cell accumulation in G0/G1, and triggered apoptosis, which proved to be largely dependent on RBL2/p130 itself, as shown upon RBL2/p130 silencing. AKT inhibition induced RBL2/p130-dependent apoptosis also in HEK-293 cells, in which re-expression of a short hairpin-resistant RBL2/p130 was able to rescue AKTiVIII-induced apoptosis upon RBL2/p130 silencing. Our data also showed that the combination of AKT and cyclin-dependent kinases (CDK) inhibitors, which converge on the re-activation of RBL2/p130 antitumoral potential, could be a promising anticancer strategy.

In plain English

This laboratory study used human cells grown in dishes; it involved no patients and did not study asbestos. The authors studied two proteins: RBL2/p130, which helps restrain cell division, and AKT, an enzyme that helps cells survive. They tested lung cancer, pleural mesothelioma and human embryonic kidney (HEK-293) cell lines. They reported that AKT binds RBL2/p130 and adds a phosphate group at one site, Ser941. A drug that blocks AKT (AKT inhibitor VIII) lowered cell viability in all cancer lines tested. In the A549 lung cancer and MSTO-211H mesothelioma lines, it raised RBL2/p130 levels, made more cells gather in an early phase of the cell cycle (G0/G1) and triggered cell death (apoptosis). When RBL2/p130 was switched off, apoptosis after the drug fell markedly. In HEK-293 cells, restoring RBL2/p130 brought it back. With roscovitine, combination index values were always below 1, indicating synergy (a stronger joint effect); the authors also reported synergy with cisplatin. The authors concluded that RBL2/p130 is a direct AKT target with an active role in cell death. They noted that results may depend on cell type and that some findings were preliminary.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) study using cultured human cell lines; no patients or animals
• Who was studied: Human cell lines: non-small cell lung cancer (A549, NCI-H358); pleural mesothelioma (MMB, NCI-H2452, IST-MES 2, REN, MSTO-211H, NCI-H2052); MET-5A cells derived from normal mesothelium; human embryonic kidney HEK-293 cells
• Exposure: Asbestos was not studied. Cells were treated with the AKT inhibitor VIII (AKTiVIII), either alone or combined with roscovitine (a CDK inhibitor) or with cisplatin. Doses included AKTiVIII at each cell line's IC50 value and 25 µM in HEK-293 cells.
• Main results:
  • Purified active AKT1 added a phosphate group to the RBL2/p130 pocket region. It did not do so when the Ser941 site was changed to alanine.
  • AKTiVIII reduced long-term colony formation in A549 (lung cancer) and MSTO-211H (mesothelioma) cells.
  • In A549 and MSTO-211H cells, AKT inhibition increased total RBL2/p130 protein. It also increased the activity of the RBL2 and CDKN1B (p27) genes.
  • After 24 hours of AKTiVIII, more A549 and MSTO-211H cells accumulated in the G0/G1 phase of the cell cycle.
  • AKTiVIII induced apoptosis in A549 and MSTO-211H cells. A caspase blocker inhibited this apoptosis.
  • In cells where RBL2/p130 had been silenced, apoptosis after AKTiVIII was markedly lower. The authors showed one representative experiment out of two.
  • In silenced HEK-293 cells, re-introducing RBL2/p130 restored apoptosis after AKT inhibition.
  • In A549 and MSTO-211H cells, combining AKTiVIII with roscovitine always gave combination index values below 1, which indicates synergy. The authors also reported synergy for AKTiVIII combined with cisplatin.
  • The drug combinations did not significantly affect the viability of MET-5A cells, which were derived from normal mesothelium.
• The authors' conclusion: The authors concluded that RBL2/p130 is a new direct target of AKT. They also concluded that, beyond restraining cell division, RBL2/p130 has an active role in promoting cell death in various cancer cell types.
• Limitations the authors state:
  • The mechanisms and biological effects may depend on the cell type and need to be studied separately in other cell types.
  • The TP73 findings were described as preliminary, and the role of p73 still needs to be characterized.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Apoptosis, Lung Cancer, Mesothelioma.

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