Immunohistochemistry of tumor tissue for Ki-67 expression and TUNEL, relation of [¹⁸F]FLT and [¹⁸F]FDG uptake to Ki-67 expression, and measurement of tumor volume for the assessment of treatment response.

<p>(A) Frozen tissue was stained for Ki-67 and TUNEL (magnification 10×). Columns, average number of TUNEL positive cells (green cells) were counted in three randomly selected field (area 0.625mm²) in two tumor samples for each cell line. The Ki-67 labeling index as assessed by the percentage of nuclei stained with MIB-1 per total number of nuclei was compared to uptake ratios of [¹⁸F]FLT and [¹⁸F]FDG (B). Effects of daily Erlotinib treatment on the tumor size of the xenografts for the assessment of tumor response (C).</p

[¹⁸F]FLT PET indicates response to therapy after 2 days of erlotinib treatment.

<p>In (A) a representative [¹⁸F]FLT PET image of a mouse bearing the sensitive PC9, HCC827 and the resistant H1975 xenografts before beginning of treatment, 48h and 96h after daily erlotinib treatment (Tarceva, 50mg/kg). (B) Quantitative analysis of changes in [¹⁸F]FLT and [¹⁸F]FDG uptake ratios after 48h and 96h daily erlotinib treatment vs. vehicle only as control (PC9: n = 8; vehicle, n = 2; HCC827: n = 7; vehicle, n = 2; H1975: n = 8; vehicle, n = 2).</p

Erlotinib treatment induces down-regulation of EGFR/EGFR-coupled signaling pathways and cell cycle arrest with subsequent induction of apoptosis in EGFR sensitive tumor cells.

<p>The erlotinib sensitive cell lines HCC827 and PC9 and the erlotinib-resistant cell line H1975 were treated with the indicated doses of erlotinib for 24 hours. Whole-cell lysates were subjected to immunoblotting with the indicated antibodies (A). PC9, HCC827 and H1975 cells were treated with erlotinib (0.5 µM) for 24h, 48h and 72h; nuclei were prepared, stained with propidium iodide and analyzed by flow cytometry. Results are shown for the G1 and S phases of the cell cycle (A). Apoptotic effects of erlotinib on EGFR-sensitive cell lines in comparison to the T790M mutant H1975 (B). Annexin V FACS was performed 12h, 24h, 36h, 48h, 72h and 96h after 0.5 µM erlotinib treatment. Images show Annexin V-positive cells after 48h in the different cell lines.</p