pher2 y877  (Danaher Inc)

Journal: Breast Cancer Research : BCR

Article Title: Direct inhibition of PI3K in combination with dual HER2 inhibitors is required for optimal antitumor activity in HER2+ breast cancer cells

doi: 10.1186/bcr3601

Figure Lengend Snippet: PIK3CA mutation uncouples phosphoinositide 3-kinase signaling from HER2 inhibition by lapatinib. (A) BT474 and SKBR3 cells infected with wild-type, E545K or H1047R constructs were treated with lapatinib at the indicated doses, and lysates were analyzed by immunoblotting with the indicated antibodies. (B) Lysates from PIK3CA wild-type or mutant expressing cells treated with a range of lapatinib doses (0.0016 to 5 μM) were analyzed by ELISA for pHER2, pAkt and pS6. Half-maximal concentration (IC 50 ) values were calculated, and the mean log IC 50 ± SEM values for three replicate dose–inhibitor curves are shown. * P < 0.05, ** P < 0.01, *** P < 0.001 and **** P < 0.0001. (C) HER2+ cell lines with wild-type PIK3CA (BT474 or SKBR3) or with a PIK3CA mutation (MDA361, HCC1954, SUM190 or UACC893) were treated with varying lapatinib doses and analyzed as described in (B) . Mean log IC 50 values from three replicates ± SEM are shown. Mean IC 50 data are shown in Table .

Article Snippet: Antibodies from the following sources were used for analysis: pHER2 Y1248 (R&D Systems, Minneapolis, MN, USA); Y877 pHER2 (Epitomics, Burlingame, CA, USA); Y1221/2 pHER2, Y11197 and Y1289 pHER3, S473 pAkt, Akt, S240/44 pS6, pErk1/2, Erk, and p110α PI3K (Cell Signaling Technology, Danvers, MA, USA); p85 N-terminal Src homology 2 (SH2) domain (EMD Millipore); HA (Covance, Princeton, NJ, USA); actin (Sigma-Aldrich, St Louis, MO, USA); HER2 (Thermo Fisher, Pittsburgh, PA, USA); and glyceraldehyde 3-phosphate dehydrogenase (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Mutagenesis, Inhibition, Infection, Construct, Western Blot, Expressing, Enzyme-linked Immunosorbent Assay, Concentration Assay

Journal: PLoS ONE

Article Title: Trastuzumab effects depend on HER2 phosphorylation in HER2-negative breast cancer cell lines

doi: 10.1371/journal.pone.0234991

Figure Lengend Snippet: Y877 is localized in the activation loop of HER2 tyrosine kinase domain. Recep-L: Receptor-L-domain. Furin-like: Furin-like cysteine rich region domain. GF-Recep: Growth factor receptor domain IV. T and TM: Transmembrane domain. TK domain: Tyrosine-kinase domain. C-loop: Catalytic loop. A-loop: Activation loop. Arrows: beta sheets. Cylinders: alpha helix.

Article Snippet: HER2 status was obtained by using the Herceptest TM kit–Dako and pHER2 Y877 was using by IHC with a specific anti-pHER2 Y877 antibody.

Techniques: Activation Assay

Journal: PLoS ONE

Article Title: Trastuzumab effects depend on HER2 phosphorylation in HER2-negative breast cancer cell lines

doi: 10.1371/journal.pone.0234991

Figure Lengend Snippet: pHER2 Y877 status was evaluated using the 2013 ASCO/CAP scoring guidelines after staining by IHC with anti-pHER2 Y877 antibody. Score 2+ were considered as positive. HER2 status was evaluated by IHC (HercepTestTM kit) according to the 2013 ASCO/CAP scoring. Molecular subtypes were identified using the ER and PR status evaluates by IHC and the global HER2 status (IHC + FISH status). A) pHER2 Y877 prevalence in the cohort. B) pHER2 Y877 distribution according to HER2 status (defined by IHC). C) pHER2 Y877 distribution according to molecular subtypes.

Article Snippet: HER2 status was obtained by using the Herceptest TM kit–Dako and pHER2 Y877 was using by IHC with a specific anti-pHER2 Y877 antibody.

Techniques: Staining

Journal: PLoS ONE

Article Title: Trastuzumab effects depend on HER2 phosphorylation in HER2-negative breast cancer cell lines

doi: 10.1371/journal.pone.0234991

Figure Lengend Snippet: pHER2 Y877 distribution according to estimation from Public Health Agency of Canada (PHAC) and U.S Breast Cancer Statistics estimation.

Article Snippet: HER2 status was obtained by using the Herceptest TM kit–Dako and pHER2 Y877 was using by IHC with a specific anti-pHER2 Y877 antibody.

Techniques:

Journal: PLoS ONE

Article Title: Trastuzumab effects depend on HER2 phosphorylation in HER2-negative breast cancer cell lines

doi: 10.1371/journal.pone.0234991

Figure Lengend Snippet: Breast cancer cell lines selected depending on their HER2 and pHER2 Y877 status. HER2 status was obtained by using the Herceptest TM kit–Dako and pHER2 Y877 was using by IHC with a specific anti-pHER2 Y877 antibody. ER (Estrogen Receptor) and PR (Progesterone Receptor) were confirmed by literature. HER2 (Human Epidermal Receptor 2), ER (Estrogen Receptor), PR (Progesteron Receptor), TNBC (Triple Negatif Breast Cancer).

Article Snippet: HER2 status was obtained by using the Herceptest TM kit–Dako and pHER2 Y877 was using by IHC with a specific anti-pHER2 Y877 antibody.

Techniques:

Journal: PLoS ONE

Article Title: Trastuzumab effects depend on HER2 phosphorylation in HER2-negative breast cancer cell lines

doi: 10.1371/journal.pone.0234991

Figure Lengend Snippet: A. Overexpression of HER2 with non-overphosphorylation of HER2 (HER2+; pHER2-). B. Non-overexpression of HER2 with overphosphorylation of HER2 (HER2-; pHER2+). In this study we show that BT-474 and SKBR3, which are HER2+/pHER2 Y877 + have a better response to trastuzumab than MDA-MB-453, which is also HER2+ but not phosphorylated at Y877 (HER2+/pHER2 Y877 −). This is concordant with studies reporting that HER2 phosphorylation leads to a better response to trastuzumab in HER2-positive BC tumors [ – ]. As shown in a study by Giuliani et al ., among HER2+ BC patients treated with trastuzumab, 89% with pHER2Y 1248 + showed a positive response while only 49% of pHERY 1248 − presented a positive response . These results suggest that the combination of HER2+/pHER Y877 + could indeed predict a better response to trastuzumab. However, our study is the first to examine HER2 phosphorylation at position Y877 in HER2-negative BC cell lines with regard to trastuzumab treatment. We demonstrated here that the decrease in proliferation in HER2-negative BC cell lines is Y877-phosphorylation-specific, as the TNBC cell line MDA-MB-468, which is HER2−/pHER2 Y877 +, displays sensitivity to trastuzumab. Studies have reported that Y877 phosphorylation is a marker of HER2 activation. This again is in agreement with our results showing that HER2 over-activation by over-phosphorylation at Y877 could be an additional biomarker in BC diagnosis.

Article Snippet: HER2 status was obtained by using the Herceptest TM kit–Dako and pHER2 Y877 was using by IHC with a specific anti-pHER2 Y877 antibody.

Techniques: Over Expression, Marker, Activation Assay, Biomarker Assay