Antitumor activity of chemoendocrine therapy in premenopausal and postmenopausal models with human breast cancer xenografts.

We examined the efficacy of chemoendocrine therapy using capecitabine as a chemotherapeutic agent in premenopausal and postmenopausal models with estrogen receptor (ER)-positive human breast cancer xenografts. Tamoxifen and letrozole were used as endocrine therapeutic agents for premenopausal and postmenopausal models, respectively. The antitumor activity of capecitabine in combination was significantly superior to either monotherapy treatment in both premenopausal (p<0.01) and postmenopausal (p<0.05) models. No increase in toxicity in terms of body weight loss was observed during treatment in either of the xenograft models. In the premenopausal model, the level of thymidine phosphorylase (TP), a key enzyme generating 5-FU from capecitabine, was upregulated (p<0.05) in tumors by tamoxifen but not by letrozole treatment in the postmenopausal model. The combination of 5'-deoxy-5-fluorouridine (5'-DFUR; an intermediate of capecitabine) with 4-hydroxytamoxifen (4-OHT; an active form of tamoxifen) or letrozole was also evaluated in vitro by using estrogen-responsive element (ERE) reporter gene assays aimed to model premenopausal and postmenopausal breast cancer. Both combinations decreased the number of estrogen-responding cells in a concentration-dependent manner and further analysis by isobolograms revealed a synergistic effect of the combination of 5'-DFUR with 4-OHT, and at least an additive effect of the combination of 5'-DFUR with letrozole. These results suggest that chemoendocrine therapy using capecitabine may be a useful treatment modality for patients with hormone-receptor-positive breast cancer, regardless of the menopausal status and should be explored in clinical trials.

Erlotinib inhibits osteolytic bone invasion of human non-small-cell lung cancer cell line NCI-H292.

Previous preclinical and clinical findings have suggested a potential role of epidermal growth factor receptor (EGFR) in osteoclast differentiation and the pathogenesis of bone metastasis in cancer. In this study, we investigated the effect of erlotinib, an orally active EGFR tyrosine kinase inhibitor (TKI), on the bone invasion of human non-small-cell lung cancer (NSCLC) cell line NCI-H292. First, we established a novel osteolytic bone invasion model of NCI-H292 cells which was made by inoculating cancer cells into the tibia of scid mice. In this model, NCI-H292 cells markedly activated osteoclasts in tibia, which resulted in osteolytic bone destruction. Erlotinib treatment suppressed osteoclast activation to the basal level through suppressing receptor activator of NF-κB ligand (RANKL) expression in osteoblast/stromal cell at the bone metastatic sites, which leads to inhibition of osteolytic bone destruction caused by NCI-H292 cells. Erlotinib inhibited the proliferation of NCI-H292 cells in in vitro. Erlotinib suppressed the production of osteolytic factors, such as parathyroid hormone-related protein (PTHrP), IL-8, IL-11 and vascular endothelial growth factor (VEGF) in NCI-H292 cells. Furthermore, erlotinib also inhibited osteoblast/stromal cell proliferation in vitro and the development of osteoclasts induced by RANKL in vitro. In conclusion, erlotinib inhibits tumor-induced osteolytic invasion in bone metastasis by suppressing osteoclast activation through inhibiting tumor growth at the bone metastatic sites, osteolytic factor production in tumor cells, osteoblast/stromal cell proliferation and osteoclast differentiation from mouse bone marrow cells.

Schedule-dependent antitumor activity of the combination with erlotinib and docetaxel in human non-small cell lung cancer cells with EGFR mutation, KRAS mutation or both wild-type EGFR and KRAS.

Erlotinib is used as a standard treatment for recurrent advanced non-small cell lung cancer (NSCLC). Epidermal growth factor receptor (EGFR) mutations in NSCLC have been shown to be a predictive factor of erlotinib, although the relationship between K-ras oncogene (KRAS) mutations and erlotinib resistance is controversial. Recently, in vitro sequence-dependent interactions of erlotinib and docetaxel have been studied on as a novel therapeutic approach against NSCLC. The purpose of the present study was to determine the optimum novel regimen of erlotinib and docetaxel against NSCLC cells which have EGFR mutation (HCC827 cells), KRAS mutation (A549 cells) or both wild-type (NCI-H292 cells). First, we analyzed the effects of in vitro combination for cell proliferation-inhibition using a combination index. In all cell lines, docetaxel followed by erlotinib treatment showed nearly additive effects. On the other hand, erlotinib followed by docetaxel treatment showed remarkable antagonistic interactions. Second, we examined the effect of combinations on the in vitro apoptosis induction. Erlotinib followed by docetaxel treatment reduced apoptosis induction compared with docetaxel alone; in contrast, docetaxel followed by erlotinib treatment had no inhibitory effects on docetaxel-induced apoptosis in any of the cell lines. Finally, an in vivo tumor growth inhibition test was performed using xenograft models. Docetaxel followed by erlotinib administration resulted in significant tumor growth inhibition compared with erlotinib or docetaxel monotherapy in all models. In conclusion, we demonstrated that docetaxel followed by erlotinib therapy was a potentially optimum regimen against NSCLC regardless of the mutation status of EGFR and KRAS.

Effect of erythropoietin on human tumor growth in xenograft models.

Recombinant human erythropoietin (rhEPO) has been used in the EU and the United States for the treatment of anemia in cancer patients after myelosuppressive chemotherapy or radiotherapy. However, several conflicting results have been reported concerning the detrimental effect of rhEPO on survival benefit in cancer patients. In experimental studies, contradictory results were also reported in in vitro tumor cell proliferation studies and in vivo tumor growth studies using tumor cells expressing EPO-receptor (EPO-R). Therefore, we tried to clarify the effect of epoetin ß, a product of rhEPO, on tumor growth in xenograft models using five EPO-R-positive human cancer cell lines, namely the MCF7 breast, 786-O renal, SCH gastric, A549 lung and SK-OV-3 ovarian cancer cell lines. Epoetin ß was administered once a week for 3 weeks at doses of 1,000, 3,000 and 10,000 IU/kg in accordance with the clinical administration schedule and dosages. As a result, no enhancement of tumor growth from the administration of epoetin ß was observed in any of the xenograft models throughout the experiment duration. The effect of epoetin ß on the antitumor activity of bevacizumab, an anti-angiogenic agent, was additionally examined using A549 and MCF7 xenograft models, since rhEPO reportedly stimulates tumor neovascularization. Epoetin ß showed no significant effect on the antitumor activity of bevacizumab in either xenograft model. These findings suggest that epoetin ß is not involved in in vivo tumor growth promotion.

Antitumor activity of erlotinib in combination with gemcitabine in in vitro and in vivo models of KRAS-mutated pancreatic cancers.

Erlotinib treatment in combination with gemcitabine is a standard therapy for patients with locally advanced pancreatic cancer in many countries, including the US and the EU. Since mutations of the K-ras oncogene (KRAS) occur in approximately 90% of pancreatic cancers, we examined the antitumor activity of erlotinib in combination with gemcitabine in KRAS-mutated pancreatic cancer cell lines, HPAC and Capan-1, which have the KRAS mutation G12D and G12V, respectively. We analyzed the mode of inhibition of in vitro tumor cell proliferation by means of a combination index and found that a combination treatment of erlotinib plus gemcitabine had an additive effect in the two cell lines. We then examined the effect of erlotinib and gemcitabine on the phosphorylation of epidermal growth factor receptor (EGFR). Erlotinib strongly suppressed, while gemcitabine augmented the phosphorylation of EGFR, which was completely blocked by erlotinib in the two cell lines. An in vivo tumor growth inhibition test was then performed using the HPAC tumor xenograft model. The combination therapy of erlotinib and gemcitabine resulted in a significant inhibition of tumor growth compared with erlotinib or gemcitabine monotherapy. To the best of our knowledge, this is the first study to show the combination effect of erlotinib and gemcitabine in vivo using a xenograft model of a KRAS-mutated pancreatic cancer cell line.

Enhancement of capecitabine efficacy by oxaliplatin in human colorectal and gastric cancer xenografts.

We have evaluated the antitumor activity of XELOX (a combination therapy of capecitabine (Xeloda) and oxaliplatin) in human colorectal and gastric cancer xenograft models. In human colorectal cancer model CXF280, antitumor activity of the combination at two-thirds of the maximum tolerated dose (MTD) was superior to that of each monotherapy at MTD. Furthermore, in human colorectal cancer model COL-05-JCK and human gastric cancer xenograft model GXF 97, the combination also showed at least additive antitumor activity. In addition, toxicity was not augmented with the combination therapy in these three models. As demonstrated using ELISA or immunohistochemistry, oxaliplatin in xenograft model tumors up-regulated the level of thymidine phosphorylase (dThdPase), a key enzyme for the metabolism of capecitabine to 5-fluorouracil. These results suggest that oxaliplatin might potentiate the antitumor activity of capecitabine by up-regulating the tumor level of dThdPase. Based on these results, clinical trials of XELOX against colorectal and gastric cancers are warranted.