Yasuhito Fujisaka

Activation of ERBB2 Signaling Causes Resistance to the EGFR-Directed Therapeutic Antibody Cetuximab

Several cancers become resistant to cetuximab by activating a bypass signaling pathway and preventing cetuximab inhibition of ERK1/2-stimulated growth. Combating Resistance to an EGF Receptor Inhibitor Many promising anticancer drugs are effective only for a limited time, because the tumor cells develop resistance. Cetuximab, directed against the epidermal growth factor receptor (EGFR), is no exception, and patients with colorectal, head and neck, or non–small cell lung cancer eventually cease to respond to the drug. Yonesaka and colleagues have determined that cetuximab-resistant cancer cells—both in culture and in patients—can up-regulate signaling through the ERBB2 growth factor receptor in several ways, permanently turning on extracellular signal–regulated kinase 1/2 (ERK1/2)–mediated growth, differentiation, and survival. They further show that interference with the ERBB2 pathway restores the ability of cetuximab to control these cancers, pointing to a promising resistance-fighting approach. The authors generated clones of cetuximab-resistant non–small cell lung and colorectal cancer cell lines by exposing the cells to increasing concentration of the drug. In some of these resistant clones, the ERBB2 receptor oncogene was genetically amplified, resulting in activated ERK1/2 signaling. Down-regulation of ERBB2 with a small interfering RNA or antibody restored sensitivity. Other clones did not have amplified ERBB2 genes but did make excess heregulin, an activating ligand for the ERBB2 receptor. Heregulin depletion or ERBB2 inhibition restored cetuximab sensitivity. After replicating these studies in xenografts in mice, the authors also looked for evidence that these resistance-associated alterations pertain to human tumors. In several groups of patients with colorectal cancer, they saw decreased survival or decreased sensitivity to cetuximab in those who exhibited amplified ERBB2 gene or higher heregulin concentrations. The concordance of their cellular data with patient experience improves confidence that concomitant treatment of certain lung, head and neck, or colorectal cancers with cetuximab and an anti-ERBB2 drug may prevent or delay the development of drug resistance. These studies add to other successes for this approach, which has also been used for analysis of other molecular targeted therapies, including EGFR kinase inhibitors. Cetuximab, an antibody directed against the epidermal growth factor receptor, is an effective clinical therapy for patients with colorectal, head and neck, and non–small cell lung cancer, particularly for those with KRAS and BRAF wild-type cancers. Treatment in all patients is limited eventually by the development of acquired resistance, but little is known about the underlying mechanism. Here, we show that activation of ERBB2 signaling in cell lines, either through ERBB2 amplification or through heregulin up-regulation, leads to persistent extracellular signal–regulated kinase 1/2 signaling and consequently to cetuximab resistance. Inhibition of ERBB2 or disruption of ERBB2/ERBB3 heterodimerization restores cetuximab sensitivity in vitro and in vivo. A subset of colorectal cancer patients who exhibit either de novo or acquired resistance to cetuximab-based therapy has ERBB2 amplification or high levels of circulating heregulin. Collectively, these findings identify two distinct resistance mechanisms, both of which promote aberrant ERBB2 signaling, that mediate cetuximab resistance. Moreover, these results suggest that ERBB2 inhibitors, in combination with cetuximab, represent a rational therapeutic strategy that should be assessed in patients with cetuximab-resistant cancers.

A Phase I Dose-Escalation Study of ZD6474 in Japanese Patients with Solid, Malignant Tumors

Introduction: ZD6474 (vandetanib) is an orally available inhibitor of vascular endothelial growth factor receptor, epidermal growth factor receptor, and RET receptor tyrosine kinase activity. This study assessed the safety and tolerability of escalating doses of ZD6474 in Japanese patients with solid, malignant tumors. Methods: Adult patients with solid tumors refractory to standard therapy received a once-daily oral dose of ZD6474 (100–400 mg) in 28-day cycles, until disease progression or unacceptable toxicity was observed. Results: Eighteen patients were treated at doses of 100 mg (n = 3), 200 mg (n = 6), 300 mg (n = 6), and 400 mg (n = 3). Dose-limiting toxicities at the completion of cycle 2 were hypertension (n = 3), diarrhea (n = 1), headache (n = 1), toxic skin eruption (n = 1), and alanine aminotransferase increase (n = 1). A dose of 400 mg/day was considered to exceed the maximum tolerated dose (MTD). Toxicities were manageable with dose interruption and/or reduction. Objective tumor response was observed in four of nine patients with non-small cell lung cancer (NSCLC) at doses of either 200 or 300 mg. Terminal half-life was about 90–115 hours. Plasma trough concentrations achieved steady-state conditions after approximately 1 month of daily dosing. Conclusions: It was concluded that a dose of 400 mg/day was considered to exceed the MTD, and doses for phase II study were thought to be not more than 300 mg/day. The objective response observed in some NSCLC patients is encouraging for further studies in this tumor type.

Phase I and Pharmacokinetic Study of HER2-targeted rhuMAb 2C4 (Pertuzumab, RO4368451) in Japanese Patients with Solid Tumors

Objective rhuMAb 2C4 (pertuzumab, RO4368451), a human epidermal growth factor receptor-2 (HER2) targeted antibody that binds to an epitope distinct from trastuzumab, blocks ligand-associated heterodimerization of HER2 with other HER receptor family members. This study evaluated the toxicity, pharmacokinetics and anti-tumor activities of pertuzumab in Japanese patients with solid tumors. Methods Patients with solid tumors refractory to standard therapy were administered pertuzumab 5, 10, 15, 20 and 25 mg/kg intravenously once every 3 weeks. Grade 3 toxicities were considered as dose limiting. The maximum tolerated dose (MTD) was a dose at which two out of six patients had Grade 3 toxicities. Results Eighteen patients, aged 38–66 (median 57) years, with solid tumors were enrolled and a total of 32 cycles of pertuzumab were administered. Toxicities were generally acceptable. Grade 3 elevation of gamma-glutamyl transpeptidase was observed in one patient at 25 mg/kg and was considered to be dose limiting. MTD was not reached up to a dose level of 25 mg/kg. The serum concentration of pertuzumab declined slowly (terminal half-life is approximately 3 weeks). The AUC proportionally increased over the dose range tested. There was limited evidence of activity (stable disease 2; progressive disease 13; and not evaluable 3); however, tumor shrinkage and tumor marker decrease were observed in an ovarian cancer and a non-small-cell lung cancer patient, respectively. Conclusions Pertuzumab is well tolerated up to 25 mg/kg. Although objective tumor response was not observed, it is worth evaluating as a flat dose and in combination with other cytotoxics and molecular-targeted agents.

Switching addictions between HER2 and FGFR2 in HER2-positive breast tumor cells: FGFR2 as a potential target for salvage after lapatinib failure.

Agents that target HER2 have improved the prognosis of patients with HER2-amplified breast cancers. However, patients who initially respond to such targeted therapy eventually develop resistance to the treatment. We have established a line of lapatinib-resistant breast cancer cells (UACC812/LR) by chronic exposure of HER2-amplified and lapatinib-sensitive UACC812 cells to the drug. The mechanism by which UACC812/LR acquired resistance to lapatinib was explored using comprehensive gene hybridization. The FGFR2 gene in UACC812/LR was highly amplified, accompanied by overexpression of FGFR2 and reduced expression of HER2, and a cell proliferation assay showed that the IC(50) of PD173074, a small-molecule inhibitor of FGFR tyrosine kinase, was 10,000 times lower in UACC812/LR than in the parent cells. PD173074 decreased the phosphorylation of FGFR2 and substantially induced apoptosis in UACC812/LR, but not in the parent cells. FGFR2 appeared to be a pivotal molecule for the survival of UACC812/LR as they became independent of the HER2 pathway, suggesting that a switch of addiction from the HER2 to the FGFR2 pathway enabled cancer cells to become resistant to HER2-targeted therapy. The present study is the first to implicate FGFR in the development of resistance to lapatinib in cancer, and suggests that FGFR-targeted therapy might become a promising salvage strategy after lapatinib failure in patients with HER2-positive breast cancer.

Randomised, phase III trial of epoetin-β to treat chemotherapy-induced anaemia according to the EU regulation

Background:Erythropoietin-stimulating agents (ESAs) effectively decrease the transfusion requirements of patients with chemotherapy-induced anaemia (CIA). Recent studies indicate that ESAs increase mortality and accelerate tumour progression. The studies also identify a 1.6-fold increased risk of venous thromboembolism. The ESA labelling was thus revised in Europe and the United States in 2008. This is the first randomised, phase III trial evaluating the efficacy and safety of epoetin-β (EPO), an ESA, dosed in accordance with the revised labelling, which specifies that ESAs should be administered to CIA patients with a haemoglobin level of ⩽10 g dl–1 and that a sustained haemoglobin level of >12 g dl–1 should be avoided.Methods:A total of 186 CIA patients (8.0 g dl–1⩽ haemoglobin ⩽10.0 g dl–1) with lung or gynaecological cancer were randomised to receive EPO 36 000 IU or placebo weekly for 12 weeks.Results:The proportion of patients receiving transfusions or with haemoglobin <8.0 g dl–1 between week 5 and the end of the treatment period as the primary end point was significantly lower in the EPO group (n=89) than in the placebo group (n=92; 10.0% vs 56.4%, P<0.001). The proportion receiving transfusions was significantly lower in the EPO group (4.5% vs 19.6%, P=0.002). Changes in quality of life were not different. No significant differences in adverse events – for example, the incidence of thromboembolic events was 1.1% for each group – or the 1-year overall survival were observed between groups.Conclusion:Weekly EPO administered according to the revised labelling approved by the European Medicines Agency is effective and well tolerated for CIA treatment. Further investigations are needed on the effect of ESAs on mortality.

A Phase 1 Clinical Study of Temsirolimus (CCI-779) in Japanese Patients with Advanced Solid Tumors

OBJECTIVE Temsirolimus (CCI-779) is a novel inhibitor of the mammalian target of rapamycin. This Phase 1 study was aimed at investigating the maximum-tolerated dose, toxicity, pharmacokinetics and antitumor activity in Japanese patients with advanced solid tumors. METHODS Temsirolimus was given as a 30 min intravenous infusion once a week. Patients with solid tumors not amenable to standard forms of treatment were eligible. Dose escalation of temsirolimus was planned from 15, 45, 80 to 165 mg/m(2). The pharmacokinetics of temsirolimus and sirolimus in whole blood were examined for cycles 1, 2, 4 and 5 of treatment. RESULTS Ten patients (median age 60.5 years; range 41-69 years) with advanced solid tumors were enrolled. Their primary cancers were renal cell carcinoma (five patients), lung cancer (three patients) and colorectal cancer (two patients). The major toxicities were hypophosphatemia diarrhea, hyperglycemia, stomatitis, pyrexia, elevated aspartate aminotransferase, rash, reduced neutrophil count, elevated alanine aminotransferase, anorexia, hypertriglyceridemia and somnolence. Two of three patients who received temsirolimus 45 mg/m(2) developed dose-limiting toxicities of Grade 3 stomatitis (one patient) and Grade 3 diarrhea (two patients). The maximum-tolerated dose was 15 mg/m(2). The peak blood concentrations of temsirolimus and sirolimus, a major active metabolite, increased in a dose-dependent manner. The area under the concentration-versus-time curve of sirolimus, but not temsirolimus, increased in a dose-dependent manner. CONCLUSIONS The recommended dose for Phase 2 clinical studies of temsirolimus in Japanese patients with advanced solid tumors is 15 mg/m(2) intravenously once a week.

Phase I pharmacokinetic and pharmacogenomic study of E7070 administered once every 21 days

E7070 is a novel sulfonamide anticancer agent that disrupts the G1/S phase of the cell cycle. The objectives of this phase I study of E7070 were to estismate the maximal tolerated dose (MTD), to determine the recommended dose for phase II, and to clarify the pharmacokinetic profile of E7070 and its relation to polymorphisms of CYP2C9 (*2, *3) and CYP2C19 (*2, *3) in Japanese patients. Patients received 1–2‐h i.v. infusions of E7070 (400, 600, 700, 800 or 900 mg/m2) on day 1 of a 21‐day cycle. Twenty‐one patients received between one and eight cycles of E7070. The dose‐limiting toxicities (DLT) comprised leukopenia, neutropenia, thrombocytopenia, elevation of aspartate aminotransferase, colitis, and ileus. The mean area under the plasma concentration–time curve (AUC) for successive dose levels increased in a non‐dose‐proportional manner. Two patients were heterozygous for the CYP2C9 mutation. For CYP2C19, eight patients were wild type and the remainder had heterozygous (n = 8) or homozygous mutations (n = 5). Regarding the CYP2C19 genotype, the AUC of patients with mutant alleles were higher than those of patients with wild type at a dose of 600 mg/m2 or more. The severity of toxic effects, such as myelosuppression, seemed to depend on the AUC. No partial responses were observed. One patient treated at a dose of 700 mg/m2 experienced a maximum tumor volume reduction of 22.5%. The MTD was estimated to be 900 mg/m2. A dose of 800 mg/m2 is recommended for further phase II studies. The pharmacokinetic/pharmacodynamic properties of E7070 seemed to be influenced by CYP2C19 genotype. The observed safety profile and preliminary evidence of antitumor activity warrant further investigation of this drug in monotherapy or in combination chemotherapy. (Cancer Sci 2005; 96: 721 –728)

CYP2C19 genotype-based phase I studies of a c-Met inhibitor tivantinib in combination with erlotinib, in advanced/metastatic non-small cell lung cancer

Background:A previous clinical study in non-small cell lung cancer (NSCLC) patients in Western countries suggested the potential for combination of a first-in-class non-ATP-competitive c-Met inhibitor tivantinib with an epidermal growth factor receptor-tyrosine kinase inhibitor erlotinib. Polymorphisms of CYP2C19, the key metabolic enzyme for tivantinib, should be addressed to translate the previous Western study to Asian population, because higher incidence of poor metabolisers (PMs) is reported in Asian population.Methods:Japanese patients with advanced/metastatic NSCLC received tivantinib in combination with erlotinib to evaluate safety and pharmacokinetics. Doses of tivantinib were escalated separately for extensive metabolisers (EMs) and PMs.Results:Tivantinib, when combined with erlotinib, was well tolerated up to 360 mg BID for EMs and 240 mg BID for PMs, respectively. Among 25 patients (16 EMs and 9 PMs), the adverse events (AEs) related to tivantinib and/or erlotinib (>20%, any grade) were rash, diarrhoea, dry skin and nausea. Grade ⩾3 AEs were leukopenia, anaemia and neutropenia. No dose-limiting toxicity was observed. Pharmacokinetics profile of tivantinib was not clearly different between the combination and monotherapy. Three partial response and three long-term stable disease (⩾24 weeks) were reported.Conclusion:Two doses of tivantinib in combination with erlotinib were recommended based on CYP2C19 genotype: 360 mg BID for EMs and 240 mg BID for PMs.

Meta-analysis of epoetin beta and darbepoetin alfa treatment for chemotherapy-induced anemia and mortality: Individual patient data from Japanese randomized, placebo-controlled trials

Erythropoiesis‐stimulating agents (ESA) reduce the need for transfusions and improve the quality of life in patients receiving chemotherapy, but several clinical trials have suggested that ESA might have a negative impact on survival. To evaluate the efficacy and safety of ESA, epoetin beta and darbepoetin alfa, including their impact on overall survival and thromboembolic events, we conducted an individual data‐based meta‐analysis of three randomized, placebo‐controlled trials studying Japanese patients with chemotherapy‐induced anemia. All trials were conducted in compliance with Good Clinical Practice. A total of 511 patients with solid tumor or lymphoma (epoetin beta or darbepoetin alfa, n = 273; placebo, n = 238) were included. The ESA significantly reduced the risk of transfusion (relative risk, 0.47; 95% confidence interval, 0.29–0.76). No significant effect of the ESA on overall survival was observed (unadjusted hazard ratio, 1.00; 95% confidence interval, 0.75–1.34). A prespecified subgroup analysis showed no strong interaction between the baseline hemoglobin concentration and the effect of ESA on overall survival. Among the ESA‐treated patients, the highest hemoglobin achieved during the treatment period in each patient had no impact on mortality. No increase in thromboembolic events was observed in the ESA‐treated patients (0.7% vs 1.7% placebo). The ESA reduced the risk of transfusion without a negative impact on the survival of patients with chemotherapy‐induced anemia.

Phase I trial of OTS11101, an anti-angiogenic vaccine targeting vascular endothelial growth factor receptor 1 in solid tumor

OTS11101 is a novel peptide vaccine that acts as an angiogenesis inhibitor by inducing cytotoxic T lymphocyte (CTL) cells that specifically target vascular endothelial cells expressing vascular endothelial growth factor (VEGF) receptor 1. We conducted a phase I study to evaluate the safety, tolerability, maximum tolerated dose, and pharmacodynamic biomarker status of this vaccine. Nine patients with advanced solid tumors received 1.0, 2.0, or 3.0 mg of OTS11101 subcutaneously, once a week in a 28‐day cycle. Three patients experienced grade 1 injection site reactions, which were the most frequent adverse events. Grade 2 proteinuria and hypertension each occurred in one patient. As other toxicities were generally mild, the maximum tolerated dose was not reached. Furthermore, we explored the induction of specific activated CTLs, and biomarkers related to angiogenesis. A pharmacodynamics study revealed that induction of specific CTLs was observed for a dose of 2.0 and 3.0 mg. The serum concentrations of soluble VEGF receptor 1 and 2 after vaccination increased significantly compared with baseline. A microarray was performed to give a comprehensive analysis of gene expression, suggesting that OTS11101 vaccination resulted in T cell activation in a clinical setting. In conclusion, OTS11101 was well tolerated in patients up to 3.0 mg once weekly and our biomarker analysis suggested that this anti‐angiogenesis vaccine is biologically active. (Cancer Sci 2013; 104: 98–104)