EGFR mutation and ALK fusion-positive non-small cell lung cancer: a multicenter prospective cohort study in Nagano Prefecture, Japan

Introduction. We prospectively examined current clinical practices in patients with inoperable epidermal growth factor receptor (EGFR) mutation and anaplastic lymphoma kinase (ALK) fusion-positive (EGFR+ and ALK+, respectively) non-small cell lung cancer (NSCLC) in Nagano Prefecture, Japan.  Material and methods. The study population consisted of newly diagnosed patients with inoperable EGFR+ and ALK+ NSCLC in 14 hospitals in Nagano between May 2016 and March 2019. Both initial and subsequent treatment decisions were made at the discretion of the attending physician.  Results. A total of 281 patients with EGFR+ NSCLC (mean age, 74 years, 59.1% female) and 26 patients with ALK+ NSCLC (mean age, 66 years, 53.8% female) were included in the study. The study population consisted of 148/107/29/20/3 cases with performance status 0/1/2/3/4 and 6/2/31/194/75 cases with clinical stage I/II/III/IV/recurrence, respectively. First-line therapy with tyrosine kinase inhibitors was performed in 259 (92.2%) and 22 (84.6%) patients with EGFR+ and ALK+ NSCLC, respectively. The median overall survival rate was 41.2 months (95% CI 36.8–45.6 months) with EGFR+. It was not reached with ALK+ .  Conclusions. This observational analysis represents a valuable resource for evaluating the outcomes of treatment in patients with NSCLC

Enhanced anti-tumor effect of zoledronic acid combined with temozolomide against human malignant glioma cell expressing O6-methylguanine DNA methyltransferase.

Temozolomide (TMZ), a DNA methylating agent, is widely used in the adjuvant treatment of malignant gliomas. O6-methylguanine-DNA methyltranferase (MGMT), a DNA repair enzyme, is frequently discussed as the main factor that limits the efficacy of TMZ. Zoledronic acid (ZOL), which is clinically applied to treat cancer-induced bone diseases, appears to possess direct anti-tumor activity through apoptosis induction by inhibiting mevalonate pathway and prenylation of intracellular small G proteins. In this study, we evaluated whether ZOL can be effectively used as an adjuvant to TMZ in human malignant glioma cells that express MGMT. Malignant glioma cell lines, in which the expression of MGMT was detected, did not exhibit growth inhibition by TMZ even at a longer exposure. However, combination experiment of TMZ plus ZOL revealed that a supra-additive effect resulted in a significant decrease in cell growth. In combined TMZ/ZOL treatment, an increased apoptotic rate was apparent and significant activation of caspase-3 and cleavage of poly-(ADP-ribose) polymerase were observed compared with each single drug exposure. There were decreased amounts of Ras-GTP, MAPK and Akt phosphorylation and MGMT expression in the ZOL-treated cells. Subcutanous xenograft models showed significant decrease of tumor growth with combined TMZ/ZOL treatment. These results suggest that ZOL efficaciously inhibits activity of Ras in malignant glioma cells and potentiates TMZ-mediated cytotoxicity, inducing growth inhibition and apoptosis of malignant glioma cells that express MGMT and resistant to TMZ. Based on this work, combination of TMZ with ZOL might be a potential therapy in malignant gliomas that receive less therapeutic effects of TMZ due to cell resistance

<i>In vitro</i> growth-inhibitory effect of co-treatment of ZOL with TMZ on MGMT-expressing malignant glioma cells.

<p><b>A, Growth-inhibitory effect of ZOL in human malignant glioma T98G cell line.</b> One thousand cells per well were placed onto a 96-well tissue culture plate and incubated overnight. ZOL was added at concentrations of 0–80 µM and incubated for 0–5 days before an MTS assay was performed. T98G did not exhibit substantial growth inhibition by ZOL (40 µM) at 120 hours. <b>B, Growth-inhibitory effect of TMZ and ZOL combination in human malignant glioma cell lines. </b><i>Upper</i> One thousand cells per well were placed onto a 96-well tissue culture plate and incubated overnight. TMZ (100 µM) and/or ZOL (40 µM) was added and incubated for 0–5 days before an MTS assay was performed. This figure is representative of three independent experiments. Bars, SD. *, P<0.05. <i>Lower</i> In the same experiment as <i>Upper</i>, an MTS assay after 120 hours incubation was highlighted. T98G and LN-18 exhibited significant growth inhibition by 100 µM TMZ plus 40 µM ZOL despite modest inhibition by each drug. <b>C, Isobologram analysis of growth-inhibitory effect of TMZ and ZOL combination in human malignant glioma T98G cell line.</b> Cells were treated with TMZ and/or ZOL as a single drug or in combination. Combination effect was analyzed at the 50% growth-inhibition ratio endpoints. The isobologram was based upon the dose-response curves, as determined by MTS assay. The X- and Y-axis indicate the ZOL and TMZ concentrations (µM), respectively. The isobologram plotted at the 50% endpoint was curved downward, suggesting synergistic interaction between TMZ and ZOL in T98G.</p

A diagram of the proposed mechanism showing anti-tumor effect of TMZ in combination with ZOL against malignant glioma cells expressing MGMT.

<p>According to the results of the present study, ZOL inhibits the activity of Ras and the expression of MGMT in malignant glioma cells and potentiates TMZ-mediated cytotoxicity, inducing growth inhibition and apoptosis of malignant glioma cells that express MGMT and resistant to TMZ. The molecular mechanism leading to the pathway indicated by the dotted arrow remains to be determined.</p