Development of molecular agents for IGF receptor targeting.

The type 1 insulin-like growth factor receptor (IGF1R) is a promising anticancer treatment target, being frequently overexpressed by tumours, and mediating proliferation, motility and apoptosis protection. Design of specific kinase inhibitors is problematic because of homology between the IGF1R and insulin receptor. This obstacle can be circumvented using sequence-specific molecular agents including antisense, triplex and ribozymes. Recent studies indicate that profound sequence-specific IGF1R gene silencing can be induced by small interfering RNAs that mediate RNA interference in mammalian cells. IGF1R downregulation blocks tumour growth and metastasis, and enhances sensitivity to cytotoxic drugs and irradiation. In murine melanoma cells, radiosensitisation is associated with impaired activation of Atm, which is required for initiation of cell cycle checkpoints and DNA repair pathways after double-strand DNA breaks. Furthermore, tumour cells killed in vivo following IGF1R downregulation can provoke an immune response, protecting against tumour rechallenge. After years of studying the role of the IGF system in tumour biology, novel agents for IGF1R targeting will soon be available for clinical testing. This review summarises the development of molecular agents, and considers factors that will influence clinical activity, including the requirement of established tumours for IGF signalling, and the efficacy and toxicity of IGF1R inhibitors.

Downregulation of the type 1 insulin-like growth factor receptor in mouse melanoma cells is associated with enhanced radiosensitivity and impaired activation of Atm kinase.

The type 1 insulin-like growth factor receptor (IGF1R) is required for growth, tumorigenicity and protection from apoptosis. IGF1R overexpression is associated with radioresistance in breast cancer. We used antisense (AS) RNA to downregulate IGF1R expression in mouse melanoma cells. Cells expressing AS-IGF1R transcripts were more radiosensitive in vitro and in vivo than controls. Also they showed reduced radiation-induced p53 accumulation and p53 serine 18 phosphorylation, and radioresistant DNA synthesis. These changes were reminiscent of the cellular phenotype of the human genetic disorder ataxia-telangiectasia (A-T), caused by mutations in the ATM gene. Cellular Atm protein levels were lower in AS-IGF1R-transfected cells than in control cells, although there was no difference in Atm expression at the transcriptional level. AS-IGF1R cells had detectable basal Atm kinase activity, but failed to induce kinase activity after irradiation. This suggests that IGF1R signalling can modulate the function of Atm, and supports the concept of targeted IGF1R downregulation as a potential treatment for malignant melanoma and other radioresistant tumours.

Phase I study of the novel cyclic AMP (cAMP) analogue 8-chloro-cAMP in patients with cancer: toxicity, hormonal, and immunological effects.

The cyclic AMP (cAMP)-dependent protein kinase regulatory subunit RI is overexpressed in cancer cells. 8-Chloro-cAMP (8-Cl-cAMP) is an RII site-specific analogue that down-regulates RI and inhibits the growth of a wide range of cancer cells in vitro and in vivo. We performed a Phase I trial of 8-Cl-cAMP in 32 patients with malignancies that were refractory to standard treatments. 8-Cl-cAMP was initially given in a 1-month cycle by constant infusion at 0.005 mg/kg/h for 21 days, followed by 1 week of rest. The dose was escalated to 0.045 mg/kg/h, but hypercalcemia became the dose-limiting toxicity. The length of drug administration was, therefore, reduced to 5 days per week for the first 3 weeks of the cycle, but it was not possible to increase the drug dose without producing hypercalcemia. Hence, the length of drug administration was reduced to 3 days per week for the first 3 weeks of the cycle. The maximum tolerated dose for this regimen was 0.15 mg/kg/h, and the dose-limiting toxicities were reversible hypercalcemia and hepatotoxicity. Stable disease for > or =4 months was observed in two patients treated at > or =0.045 mg/kg. cAMP-dependent protein kinase is involved in hormone- and cytokine-mediated signaling, and so representative hormone, cytokine, and peripheral lymphocyte subsets were measured. The drug had a parathyroid hormone-like effect on calcium homeostasis and significantly increased circulating luteinizing hormone and 17-hydoxyprogesterone levels (P < 0.02 and P < 0.0006, respectively). We conclude that 8-Cl-cAMP is well tolerated without attendant myelotoxicity, and in this study, it was associated with biological effects. In Phase II studies, a dose of 0.11 mg/kg/h for 3 days per week would be appropriate.

Cyclophosphamide, methotrexate and infusional 5-fluorouracil (infusional CMF) in metastatic breast cancer.

Bolus 5-fluorouracil (5-FU) is a phase-specific drug with a short plasma half-life that is used in combination with bolus cyclophosphamide and methotrexate in the treatment of breast cancer. The efficacy of 5-FU can be improved by continuous intravenous infusion using portable infusion pumps (infusional 5-FU). Infusional 5-FU, 200 mg m(-2) day(-1), in combination with standard doses of bolus cyclophosphamide and methotrexate, was evaluated in a phase I/II dose-finding study. The cyclophosphamide and methotrexate were administered in 28-day cycles as follows: cohort 1, cyclophosphamide 600 mg m(-2), days 1 and 8, and methotrexate 40 mg m(-2), day 1; cohort 2, cyclophosphamide 400 mg m(-2), days 1 and 8, and methotrexate 40 mg m(-2), day 1; cohort 3, cyclophosphamide 480 mg (m-2), days 1 and 8, and methotrexate 40 mg m(-2), day 1; cohort 4, cyclophosphamide 480 mg m(-2), days 1 and 8, and methotrexate 40 mg m(-2), days 1 and 8. Median overall survival was 10 months (range 3-21 months). Objective tumour responses were seen in 9 of 25 patients (36%, 95% CI 18-58%), including 3 of 13 patients (23%) previously treated for metastatic disease. Cohorts 1 and 4 proved to be too toxic, with five of six patients in cohort 1 and three of four in cohort 4 developing grade III/IV neutropenia. The dose intensity of cyclophosphamide achieved was as follows: cohort 1, 82%; cohort 2, 86%; cohort 3, 97%; cohort 4, 90%. Infusional 5-FU can be administered safely and is effective in combination with cyclophosphamide 480 mg m(-2), days 1 and 8, and methotrexate 40 mg m(-2), day 1, in the treatment of metastatic breast cancer.

A novel cyclic adenosine monophosphate analog induces hypercalcemia via production of 1,25-dihydroxyvitamin D in patients with solid tumors.

The treatment of cancer patients with conventional chemotherapy is sometimes associated with severe systemic toxicity and only a minimal survival benefit. Because of this, new less toxic and more efficacious treatments have been sought. 8-Chloro-cAMP (8-Cl-cAMP) is one of a new generation of anticancer drugs that act at the level of signal transduction. In preclinical models, 8-Cl-cAMP modulates protein kinase A (PKA) leading to growth inhibition and increased differentiation of cancer cells. 8-Cl-cAMP was given to 16 patients with advanced cancer as an infusion via an indwelling subclavian venous catheter. We showed that 8-Cl-cAMP had a parathyroid hormone-like effect leading to increased synthesis of renal 1,25-dihydroxyvitamin D [up to 14 times the baseline value, median 3.6 times; P = 0.00001 (Student's paired t test)]. This produced the dose-limiting toxicity of reversible hypercalcemia that could not be controlled by the administration of either pamidronate or dexamethasone. The treatment was otherwise well tolerated, and other cAMP-dependent pathways (cortisol and TSH) were not affected, emphasizing the marked differences between organs in their sensitivity to this cAMP analog. Our results have shown that 8-Cl-cAMP is biologically active, and it is feasible that if the hypercalcemia can be controlled, then this drug may have a role as a single agent, or as a short infusion between cycles of chemotherapy.

A phase II study of mitomycin C and oral etoposide for advanced adenocarcinoma of the upper gastrointestinal tract.

BACKGROUND: Mitomycin C and etoposide have both demonstrated activity against gastric carcinoma. Etoposide is a topoisomerase II inhibitor with evidence for phase-specific and schedule-dependent activity. PATIENTS AND METHOD: Twenty-eight consecutive patients with advanced upper gastrointestinal adenocarcinoma were treated with intravenous (i.v.) bolus mitomycin C 6 mg/m2 on day 1 every 21 days to a maximum of four courses. Oral etoposide capsules 50 mg b.i.d. (or 35 mg b.i.d. liquid) were administered days 1 to 10 extending to 14 days in subsequent courses if absolute neutrophil count > 1.5 x 10(9)/l on day 14 of first course, for up to six courses. RESULTS: Twenty-six patients were assessed for response of whom 12 had measurable disease and 14 evaluable disease. Four patients had a documented response (one complete remission, three partial remissions) with an objective response rate of 15% (95% confidence interval (95% CI) 4%-35%). Eight patients had stable disease and 14 progressive disease. The median survival was six months. The schedule was well tolerated with no treatment-related deaths. Nine patients experienced leucopenia (seven grade II and two grade III). Nausea and vomiting (eight grade II, one grade III), fatigue (eight grade II, two grade III) and anaemia (seven grade II, two grade III) were the predominant toxicities. CONCLUSION: This out-patient schedule is well tolerated and shows modest activity in the treatment of advanced upper gastrointestinal adenocarcinoma. Further studies using protracted schedules of etoposide both orally and as infusional treatment should be developed.

A phase II study evaluating the effect of tamoxifen on DNA repair in melanoma patients treated with dacarbazine.

The addition of tamoxifen to dacarbazine containing chemotherapy regimens used in the treatment of melanoma, has been shown to increase response rates, but the mechanism of any interaction is uncertain. The object of this study was to determine whether the addition of tamoxifen to dacarbazine, would modify DNA repair in-vivo and cause an increase in O6-meG adducts in peripheral blood leucocytes. This would provide some insight into the nature of the interaction between these two drugs. Twenty three patients with metastatic malignant melanoma received dacarbazine (DTIC) 1 g/m2 every three weeks for a maximum of six cycles. Tamoxifen 20 mg daily, was started after the first cycle of chemotherapy and then taken continuously during the treatment. Adduct levels after the second cycle of treatment were significantly higher than those after the first cycle (p = 0.0001). A similar rise however, was also produced when a cohort of patients were given dacarbazine without tamoxifen during the second cycle of treatment. This study did not show an additional increase of O6-meG adducts when tamoxifen was administered and therefore this mechanism does not support a postulated interaction between tamoxifen and dacarbazine. This is in agreement with the recent randomised study which did not show any significant increase in response rate with the addition of tamoxifen.

Bryostatin 1, a novel antineoplastic agent and protein kinase C activator, induces human myalgia and muscle metabolic defects: a 31P magnetic resonance spectroscopic study.

Bryostatin 1, a novel antineoplastic agent and protein kinase C (PKC) activator, has been found to induce myalgia (muscle pain) 48 h after administration in clinical trials. This is the dose-limiting toxicity and has restricted the duration of therapy in phase I trials. To investigate the mechanisms and try to increase toleration of the drug, we studied calf muscle metabolism of 14 patients at rest and during exercise and subsequent recovery using 31P magnetic resonance spectroscopy (MRS) before and 4 h, 48-72 h and 1-2 weeks following bryostatin therapy. In resting muscle there was a significant (P < 0.001) increase in the phosphodiester/adenosine 5'-triphosphate (PDE/ATP) ratio 48 h post bryostatin and in patients with myalgia compared with pre-bryostatin control studies. Following exercise, patients with myalgia showed significantly slower phosphocreatine (PCr) and ADP recovery half-time (P < or = 0.05) suggesting impaired mitochondrial (oxidative) energy production, possibly due to a direct effect on the mitochondria or secondary to reduced blood flow. The apparent proton efflux rate following exercise was significantly reduced 4 h after bryostatin (P < or = 0.05), suggesting reduced blood flow. The rate of post-exercise reoxygenation was studied in four patients by near-infrared spectroscopy 4 h post bryostatin. In three of these the rate was reduced, consistent with reduced muscle blood flow. Bryostatin 1 appeared to cause a long-lasting impairment of oxidative metabolism and proton washout from muscle, consistent with a vasoconstrictive action. Thus these studies provide evidence for two mechanisms of the dose-limiting toxicity for bryostatin. Prospective studies on the use of vasodilators to improve the tolerance of the drug should be carried out.

Unilateral absence of the hand in second cousins.

This paper reports two second cousins with absence of the left hand.