Mainly because lapatinib is reported to become an equipotent inhibitor of the HER2 and EGFR kinases, we expected to locate that phos phorylation of EGFR, comparable to HER2, could be inhi bited in resistant cells. Having said that, examination of individual EGFR phosphotyrosine web-sites in lapatinib resistant cells unveiled a mixed pattern, as evidenced by variably persistent phosphorylation of tyrosines 992 and 1148, and marked inhibition of other phosphotyrosine web pages. These findings made it tempting to speculate that es cape from, or incomplete inhibition of EGFR tyrosine autophosphorylation websites in response to lapatinib, more than time, led to a switch in the regulation of cell survival from HER2 HER3 PI3K signaling in lapatinib naive HER2 breast cancer cells, to EGFR HER3 PI3K in cells that develop into resistance to lapatinib.
To check this hy pothesis, we molecularly knocked down EGFR selleck chemical 17-AAG in lapatinib resistant cells, which diminished HER3Y1197 phos phorylation and PI3K signaling, and led to improved apoptosis with a statistically substantial re duction in cell viability. As a result, the regulation of HER3 phosphorylation appears to switch from HER2 in treatment na ve cells, to EGFR in HER2 breast cancer cell lines that have grow to be resistant to lapatinib. Activation of a unfavorable suggestions loop in resistant tumor cells exclusively dephosphorylates AktS473 despite persistent PI3K pathway activation Inhibition of AktS473 phosphorylation in resistant cells appeared inconsistent using the persistent activation of your PI3K signaling pathway. In this context, PHLPPL is often a protein phosphatase that’s tran scriptionally regulated by mTORC1.
PHLPPL negatively feeds back on PI3K signaling by selectively dephosphorylating Akt on S473, not T308, ma king it tempting to speculate that PHLPPL could possibly be responsible for the pattern of Akt phosphorylation observed in lapatinib selleck chemical resistant cells. We located that ex pression of PHLPPL protein was increased in resistant cells compared with their parental cell counterparts. PHLPPL protein expression was de creased in parental cells treated with one uM lapatinib for 24 hours, constant with inhibition of PI3K mTOR signaling in lapatinib treated parental cells. In case the enhanced expression of PHLPPL in resistant cells had been relevant to persistent PI3K mTOR pathway ac tivation, then inhibition of PI3K signaling should block PHLPPL expression.
Certainly, PHLPPL expression was inhibited in resistant cells developing while in the presence of one uM lapatinib, following treatment with all the dual PI3K mTOR kinase inhibitor BEZ NVP 235. Also, molecular knockdown of EGFR, which blocked PI3K signaling, also inhibited PHLPPL protein expression. These findings suggest that AktS473 phosphorylation may not automatically signify a reputable pharmacodynamic readout to assess the effects of targeted therapies on PI3K signaling.