The humanised monoclonal antibody pertuzumab (Omnitarg, rhuMAb 2C4), directed against the extracellular domain of HER2/erbB2, is the first in a new class of targeted therapeutic agents known as HER dimerisation inhibitors (HDIs). In a phase I study it has shown clinical activity in ovarian cancer (Agus et al., Proc ASCO, 771, 2003) but the determinants of sensitivity remain undefined. This study sought to identify indicators of sensitivity to pertuzumab in a panel of 13 ovarian cancer cell lines. They were treated with TGFα (activator of EGF receptor / HER1) or NRG1β (activator of HER3 and HER4) (1nM) with or without pertuzumab (100 nM) for 72h and cell proliferation assessed by the SRB assay. The PE01, 41M, PE06, PE04, CAOV3 and OVCAR3 cell lines were the most responsive to NRG1β while the OVCAR4, 59M, PEA2 and SKOV-3 cell lines showed no growth change. In general, the growth responses to NRG1β and TGFα were similar (r = 0.83 ; Pearson). The magnitude of NRG-driven growth stimulation was significantly associated with the fold-increase in ERK-2 activation (p=0.019; Pearson) but not Akt stimulation (p =0.99; Pearson). Addition of 100nM pertuzumab to cells being stimulated by 1nM NRG1β produced varying degrees of growth inhibition in all cell lines that were stimulated by NRG1β. In certain cell lines e.g. PE06 and PE04, the NRG1β stimulation was completely reversed, while in most cell lines it typically produced a 40-60% reversal. Although addition of 100nM pertuzumab to cells being stimulated by 1nM TGFα also produced complete growth inhibition in the PE04, PE06 and PE01 lines, in contrast to its general inhibitory effect in NRG1β-stimulated cells, it was ineffective in the remaining cell lines (possibly as a result of EGFR homodimerisation, thereby bypassing HER2). Investigation of signaling using phospho-specific antibodies in 7 cell lines (3 sensitive and 4 resistant to pertuzumab) indicated that in the 3 pertuzumab-responsive cell lines, NRG1β increased phosphorylation at Tyr877 of HER2 which was reversed by pertuzumab. In the 4 resistant cell lines, NRG1β alone did not increase Tyr877 phosphorylation, whilst addition of pertuzumab increased the signal. NRG1β also stimulated phosphorylation of HER2 Tyr1023 and Tyr1248 in some sensitive cell lines which was not reversed by pertuzumab. In cell lines growth stimulated by NRG1β, addition of pertuzumab reduced activation of both ERK-2 and AKT. These data indicate that a subset of ovarian cancer cell lines in which NRG1β induces HER2 phosphorylation at Tyr877 are sensitive to pertuzumab. Whether this marker holds potential for identifying patients benefiting from pertuzumab treatment remains to be tested in the clinic.
[Proc Amer Assoc Cancer Res, Volume 46, 2005]