Targeting the insulin-like growth factor-1 receptor to overcome bortezomib resistance in preclinical models of multiple myeloma.

Proteasome inhibition with bortezomib is a validated approach to the treatment of multiple myeloma, but drug resistance often emerges and limits its utility in the retreatment setting. To begin to identify some of the mechanisms involved, we developed bortezomib-resistant myeloma cell lines that, unlike previously reported models, showed no ß5 subunit mutations. Instead, up-regulation of the insulin-like growth factor (IGF)-1 axis was identified, with increased autocrine and paracrine secretion of IGF-1, leading to increased activation of the IGF-1 receptor (IGF-1R). Exogenous IGF-1 reduced cellular sensitivity to bortezomib, whereas pharmacologic or small hairpin RNA-mediated IGF-1R suppression enhanced bortezomib sensitivity in cell lines and patient samples. In vitro studies with OSI-906, a clinically relevant dual IGF-1R and insulin receptor inhibitor, showed it acted synergistically with bortezomib, and potently resensitized bortezomib-resistant cell lines and patient samples to bortezomib. Importantly, OSI-906 in combination with bortezomib also overcame bortezomib resistance in an in vivo model of myeloma. Taken together, these data support the hypothesis that signaling through the IGF-1/IGF-1R axis contributes to acquired bortezomib resistance, and provide a rationale for combining bortezomib with IGF-1R inhibitors like OSI-906 to overcome or possibly prevent the emergence of bortezomib-refractory disease in the clinic.

Pilot study to assess toxicity and pharmacokinetics of docetaxel in patients with metastatic breast cancer and impaired liver function secondary to hepatic metastases.

BACKGROUND: Limited clinical data are available regarding the safety of docetaxel in metastatic breast cancer patients with liver dysfunction. METHODS: Eligible patients had breast cancer with impaired liver function secondary to hepatic metastases and were candidates for docetaxel therapy. They were assigned to one of five groups on the basis of total bilirubin, alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase levels. All other causes of liver dysfunction were excluded, and bile duct obstruction was corrected, if possible, prior to study entry. Patients received docetaxel every three weeks. The chemotherapy dose was chosen on the basis of the patient's level of hepatic dysfunction and escalated as tolerated. The primary outcome of this study was safety. The secondary outcomes were pharmacokinetic data and efficacy in terms of time to disease progression. RESULTS: Twenty-three patients were enrolled. No unexpected toxicities occurred. Grade 3/4 fatigue (65%), neutropenia (30%), myalgias (26%), neutropenic fever (26%), vomiting (9%), and rash (9%) were the most common serious adverse events. The median time to progression was three months (range 1-18 months). Pharmacokinetic results indicated that patients with more severe hepatic dysfunction may have been underdosed based on our conservative dosing strategy. CONCLUSIONS: Docetaxel can be administered to patients with metastatic breast cancer and liver dysfunction after dose attenuation. However, because of a narrow therapeutic index in this clinical setting, therapy should be closely monitored with subsequent dose escalation when possible.

A phase I study of vorinostat in combination with idarubicin in relapsed or refractory leukaemia.

Histone deacetylase inhibitors (HDACi) affect chromatin remodelling and modulate the expression of aberrantly silenced genes. HDACi have single-agent clinical activity in haematological malignancies and have synergistic anti-leukaemia activity when combined with anthracyclines in vitro. We conducted a two-arm, parallel Phase I trial to investigate two schedules of escalating doses of vorinostat (Schedule A: thrice daily (TID) for 14 d; B: TID for 3 d) in combination with a fixed dose of idarubicin in patients with refractory leukaemia. Of the 41 patients enrolled, 90% had acute myeloid leukaemia, with a median of 3 prior therapies. Seven responses (17%) were documented (two complete response (5%), one complete response without platelet recovery (2.5%), and four marrow responses). The 3-d schedule of vorinostat was better tolerated than the 14-d schedule. The maximum tolerated dose for vorinostat was defined as 400 mg TID for 3 d. The most common grade 3 and 4 toxicities included mucositis, fatigue and diarrhoea. Correlative studies demonstrated histone acetylation in patients on therapy and modulation of CDKN1A and TOP2A (topoisomerase II) gene expression. Pharmacokinetic analysis confirmed a dose-related elevation in plasma vorinostat concentrations. The combination of vorinostat and idarubicin is generally tolerable and active in patients with advanced leukaemia and should be studied in the front-line setting.

Phase I/II study of G3139 (Bcl-2 antisense oligonucleotide) in combination with doxorubicin and docetaxel in breast cancer.

PURPOSE: Preclinical data showed enhancement of breast cancer cell death when G3139 was combined with anthracyclines and taxanes. We evaluated the efficacy and safety of a Bcl-2 antisense oligonucleotide, G3139, in combination with doxorubicin (A) and docetaxel (T) in patients with locally advanced breast cancer (LABC). EXPERIMENTAL DESIGN: Following a brief phase I to determine the phase II dose, patients with locally advanced breast cancer received G3139 administered by continuous i.v. infusion for 5 to 7 days with bolus A (50 mg/m2) and T (75 mg/m2) administered on either day 3 or 6 of therapy with G3139. Cycles were repeated every 21 days x 6 in the neoadjuvant setting. Serial plasma samples were obtained for pharmacokinetic analysis. Tissue samples were obtained before and after therapy for pharmacodynamic analysis of Bcl-2 expression. RESULTS: Thirty patients (median age, 49 years; range, 24-71 years) received 160 cycles. During the phase I portion of the trial, the dose of G3139 was escalated from 3 to 7 mg/kg/d (i.v. for 5 days) in combination with AT. During the phase II portion of the trial, several doses and schedules of G3139 were evaluated. There were no pathologic complete responses. Pharmacodynamic studies showed limited Bcl-2 down-regulation in the primary tumors. CONCLUSIONS: G3139 in combination with doxorubicin and docetaxel is well tolerated. No pathologic complete response was seen and pharmacodynamic studies showed very little down-regulation of Bcl-2 in primary tumors, perhaps related to issues with insufficient drug delivery to the intact tumor.

The novel triterpenoid C-28 methyl ester of 2-cyano-3, 12-dioxoolen-1, 9-dien-28-oic acid inhibits metastatic murine breast tumor growth through inactivation of STAT3 signaling.

We and others have reported that C-28 methyl ester of 2-cyano-3, 12-dioxoolen-1, 9-dien-28-oic acid (CDDO-Me) effectively inhibits the growth of multiple cancer cell types. Our previous studies indicated that prolonged CDDO-Me treatment inactivated extracellular signal-regulated kinase signaling in acute myelogenous leukemia cells. Whether treatment with CDDO-Me has an earlier effect on other proteins that are important for either signal transduction or oncogenesis is unknown. Constitutively activated signal transducer and activator of transcription 3 (STAT3) is frequently found in human breast cancer samples. Constitutively activated STAT3 was shown to up-regulate c-Myc in several types of cancer and has a feedback effect on Src and Akt. To examine the effects of CDDO-Me on STAT3 signaling in breast cancer, we used the murine 4T1 breast tumor model, which is largely resistant to chemotherapy. In vitro, after treatment of 4T1 cells with 500 nmol/L CDDO-Me for 2 h, we found (a) inactivation of STAT3, (b) inactivation of Src and Akt, (c) 4-fold reduction of c-Myc mRNA levels, (d) accumulation of cells in G(2)-M cell cycle phase, (e) abrogation of invasive growth of 4T1 cells, and (f) lack of apoptosis induction. In in vivo studies, CDDO-Me completely eliminated 4T1 breast cancer growth and lung metastases induced by 4T1 cells in mice when treatment started 1 day after tumor implantation and significantly inhibited tumor growth when started after 5 days. In vivo studies also indicated that splenic mature dendritic cells were restored after CDDO-Me treatment. In summary, these data suggest that CDDO-Me may have therapeutic potential in breast cancer therapy, in part, through inactivation of STAT3.

Safety and pharmacokinetic effects of TNP-470, an angiogenesis inhibitor, combined with paclitaxel in patients with solid tumors: evidence for activity in non-small-cell lung cancer.

PURPOSE: Preclinical studies suggested that the antiangiogenic agent TNP-470 was synergistic with cytotoxic therapy. TNP-470 was administered with paclitaxel to adults with solid tumors to define the safety and optimal dose of the combination regimen and to assess pharmacokinetic interactions. PATIENTS AND METHODS: Thirty-two patients were enrolled chronologically onto one of two treatment arms. Arm A involved a fixed TNP-470 dose with escalating doses of paclitaxel, and Arm B involved a fixed paclitaxel dose with escalating doses of TNP-470. Paclitaxel and TNP-470 pharmacokinetics were evaluated along with toxicity. RESULTS: The combination of TNP-470 administered at 60 mg/m(2) three times per week and paclitaxel 225 mg/m(2) administered over 3 hours every 3 weeks was defined as both the maximum-tolerated dose and the optimal dose. Myelosuppression was similar to that expected with paclitaxel alone. Mild to moderate neurocognitive impairment was observed; however, the majority of changes were subclinical and reversible as determined by prestudy and poststudy neuropsychiatric test results. A clinically insignificant decrease of paclitaxel clearance was observed for the combination. Median survival for all patients was 14.1 months. Partial responses were reported in eight (25%) of 32 patients and in six (38%) of 16 patients with NSCLC, 60% of whom had received prior chemotherapy. CONCLUSION: The combination of TNP-470 and paclitaxel, each at full single-agent dose, seems well tolerated, with minimal pharmacokinetic interaction between the two agents. Further studies of TNP-470 with chemotherapy regimens are warranted in NSCLC and other solid tumors.

Development of biologic markers of response and assessment of antiangiogenic activity in a clinical trial of human recombinant endostatin.

PURPOSE: Angiogenesis is a target for the treatment of cancer and other diseases, and its complex biology suggests that establishing the appropriate dose and schedule for antiangiogenic treatment will require extensive study. We present the initial results of a dose-finding clinical trial of recombinant human endostatin (rh-Endo) that examined potential surrogates for response to antiangiogenic therapy. PATIENTS AND METHODS: Twenty-five patients were treated with escalating doses of rh-Endo. Positron emission tomography (PET) was used to assess tumor blood flow (with [15O]H2O) and metabolism (with [18F]fluorodeoxyglucose) before the start of therapy and then every 4 weeks. To directly assess the effects of rh-Endo on endothelial cells within the tumors, biopsy specimens of tumor tissue were obtained before therapy and again at 8 weeks and evaluated for endothelial cell and tumor cell apoptosis. RESULTS: Tumor blood flow and metabolism as measured by PET scans generally decreased with increasing doses of rh-Endo; however, the effects were complex and in some analyses nonlinear. Tumor biopsy analysis revealed a significant increase in tumor cell apoptosis (P =.027) and endothelial cell apoptosis (P =.027) after 8 weeks of therapy. However, there was no statistically significant relationship between rh-Endo dose and induction of tumor cell or endothelial cell apoptosis. CONCLUSION: These initial data suggest that rh-Endo has measurable effects on tumor blood flow and metabolism and induces endothelial and tumor cell apoptosis even in the absence of demonstrable anticancer effects. Further study and validation of these biomarkers in the context of antiangiogenic therapy will be required.

Phase I and pharmacological study of oral 9-aminocamptothecin colloidal dispersion (NSC 603071) in patients with advanced solid tumors.

PURPOSE: 9-Aminocamptothecin colloidal dispersion (9-ACCD; NSC 603071) is a specific inhibitor of topoisomerase I that can be given p.o. This Phase I trial was conducted to determine the toxicity profile, maximal tolerated dose, and pharmacokinetics profile, including bioavailability, of p.o. 9-ACCD in patients with advanced solid tumors. EXPERIMENTAL DESIGN: After receiving one i.v. dose of 9-ACCD, patients were treated with 9-ACCD p.o., starting with a 2-week schedule, to establish the safety. Once safety was established, patients were treated continuously for 4 weeks followed by a rest period of 2 weeks at dosages of 0.2, 0.3, 0.45, 0.56, 0.7, and 0.63 mg/m(2)/day. Serial blood samples were collected for the pharmacokinetics study on day 1 after the i.v. dose and day 2 after p.o. administration. Lactone and total 9-aminocamptothecin were analyzed by high-pressure liquid chromatographic assay. RESULTS: Thirty-two patients were treated on the study. The dose-limiting toxicity was myelosuppression at the dosage of 0.7 mg/m(2)/day. Other toxic effects included nausea, vomiting, fatigue, and transient elevation of the total bilirubin level. The maximal tolerated dose was 0.63 mg/m(2)/day. There was no objective response. The mean terminal half-life of p.o. total 9-ACCD was 1.2 +/- 1.2 h, and the volume of distribution was 17.7 +/- 20.6 l/m(2). The mean bioavailability of total 9-ACCD was 68.1 +/- 36.4%. CONCLUSIONS: Despite good tolerance of p.o. administration, the lack of clinical activity and variable absorption of 9-ACCD suggested that further development might not be warranted.

Murine pharmacokinetics and metabolism of oleandrin, a cytotoxic component of Nerium oleander.

Pharmacokinetic studies of [3H]oleandrin, a cardiac glycoside component of Anvirzel, were conducted in mice after either an i.v. dose (40 micrograms/kg) or a p.o. dose (80 micrograms/kg). Oleandrin was rapidly absorbed after oral dosing (Cmax at 20 min) although the elimination half-life was longer (2.3 +/- 0.5 h) than that after i.v. dosing (0.4 +/- 0.1 h). The AUC0-infinity values obtained after i.v. and p.o. dosing were 24.6 +/- 11.1 and 14.4 +/- 4.3 (ng.h/ml), respectively, resulting in an oral bioavailability of approximately 30%. After i.v. administration, oleandrin concentration in liver was approximately twice that measured in heart or kidney tissue. Oleandrigenin, the aglycone of oleandrin, was also found in these tissues. At 5 min, > 60% of the total radioactivity in liver was due to oleandrin while 28% of the given dose was present as oleandrigenin. Twenty-four hours following injection, 8% of total radioactivity was excreted in urine and contained both oleandrigenin (4.4% of the injected dose) and oleandrin (1.9%). Sixty-six percent of injected radioactivity was found in feces and consisted of oleandrin and oleandrigenin in equal amounts. Uptake of oleandrin in brain after i.p. injection of oleandrin (3 mg/kg) or oleander extract (700 mg/kg) was examined. Measured by LC/MS/MS, oleandrin content in brain was higher following injection of extract than it was with an equivalent dose of oleandrin. The data suggest that components within oleander extract may enhance transport of oleandrin across the blood brain barrier.

Plasma and cellular pharmacology of 8-chloro-adenosine in mice and rats.

PURPOSE: The nucleoside 8-chloro-adenosine (8-Cl-Ado) is currently being developed for treatment of multiple myeloma and leukemias. Although accumulation of the phosphorylated drug product is known to occur within cell lines, its metabolic fate in plasma or circulating cells in animals is unclear. The purpose of the present study was to determine the pharmacology of 8-Cl-Ado in rodents through examination of plasma and cellular levels of parent drug and metabolites. In addition, we sought to determine whether an inhibitor of adenosine deaminase, 2'-deoxycoformycin (dCF), could enhance intracellular formation of 8-Cl-ATP by preventing degradation of 8-Cl-Ado to 8-Cl-inosine (8-Cl-Ino). METHODS: A validated HPLC assay permitted simultaneous determination of 8-Cl-Ado, 8-Cl-adenine (8-Cl-Ade), dCF, and 8-Cl-Ino. Radiolabeled cellular nucleotides were obtained from peripheral blood mononuclear cells (PBMC) of both mice and rats using a perchloric acid extraction procedure and were separated by HPLC. RESULTS: Stability of 8-Cl-Ado in the presence or absence of dCF was examined in fresh plasma from mice, rats and humans. Conversion of 8-Cl-Ado to 8-Cl-Ino was only marginally affected by coincubation with dCF. In CD(2)F(1) mice given 8-Cl-Ado i.p. at 100 mg/kg, there was rapid appearance in plasma of both 8-Cl-Ade and 8-Cl-Ino. The identities of the metabolites were confirmed by mass spectrometry. The plasma [(3)H]8-Cl-Ado concentration 1 h after drug injection was 1.3 micro M in mice while the intracellular levels of [(3)H]8-Cl-AMP and [(3)H]8-Cl-ATP were 1 m M and 350 micro M, respectively. Mice that had received dCF (2 mg/ml) 30 min prior to [(3)H]8-Cl-Ado had 27% less intracellular [(3)H]8-Cl-ATP in PBMC compared to mice without dCF pretreatment. The pharmacokinetics of 8-Cl-Ado were examined in greater detail in Sprague-Dawley rats. Animals were given [(3)H]8-Cl-Ado (42.5 mg/kg, i.v.) by itself or 30 min following injection of dCF (4 mg/kg). Mononuclear cells in mice accumulated 350 or 1200 micro M [(3)H]8-Cl-ATP 1 h after injection of either 50 or 100 mg [(3)H]8-Cl-Ado, respectively. The major metabolite in these cells was the monophosphate, which was four- to sevenfold higher in concentration than the triphosphate metabolite. In rats, [(3)H]8-Cl-AMP concentrations in PBMC were similar to those of the triphosphate metabolite which achieved a peak of 90 micro M 2 h after a bolus injection of 8-Cl-Ado (40 mg/kg). Cellular clearance of 8-Cl-ATP appeared to be slow: 24 h after injection of 8-Cl-Ado the cellular concentration of 8-Cl-ATP was still 40 micro M. CONCLUSIONS: The use of dCF did not significantly alter 8-Cl-ATP levels in PBMC and is not considered to be a useful therapeutic strategy. Even though a portion of 8-Cl-Ado is metabolically inactivated in plasma, high levels of cytotoxic 8-Cl-ATP accumulated intracellularly in these animals and were retained for a considerable length of time. Further development of 8-Cl-Ado is recommended.

Phase I study of the combination of sorafenib and temsirolimus in patients with metastatic melanoma.

PURPOSE: This phase I clinical trial was conducted to determine the safety, efficacy, and molecular effects of sorafenib with temsirolimus in patients with advanced melanoma. PATIENTS AND METHODS: Patients with stage IV or unresectable or recurrent stage III melanoma and Eastern Cooperative Oncology Group performance status of 0 to 1 were eligible. Sorafenib was given orally once or twice daily and temsirolimus was given i.v. weekly, both starting on day 1, with a 4-week cycle. Responses were assessed every 2 cycles per Response Evaluation Criteria in Solid Tumors criteria. Consenting patients with accessible tumors underwent optional tumor biopsies before treatment and after the second infusion of temsirolimus. Tumor biopsies were analyzed for activating mutations in BRAF and NRAS, and for expression of P-extracellular signal-regulated kinase (P-ERK) and P-S6 proteins. RESULTS: A total of 25 patients were accrued to the study. The maximum tolerated doses were sorafenib 400 mg every morning and 200 mg every evening and temsirolimus 25 mg i.v. weekly. Dose-limiting toxicities included thrombocytopenia, hand-foot syndrome, serum transaminase elevation, and hypertriglyceridemia. There were no complete or partial responses with the combination; 10 patients achieved stabilization of disease as their best response. The median progression-free survival was 2.1 months. Matching pretreatment and day 15 tumor biopsies showed marked inhibition of P-S6 with treatment in 3 of 4 evaluable patients, but minimal inhibition of P-ERK. CONCLUSIONS: Combination therapy with sorafenib and temsirolimus resulted in significant toxicity at higher dose levels, failed to achieve any clinical responses in genetically unselected patient population, and did not inhibit P-ERK.

Phase I trial of hepatic arterial infusion of nanoparticle albumin-bound paclitaxel: toxicity, pharmacokinetics, and activity.

Because liver involvement in patients with metastatic cancer has limited options and poor outcomes, we conducted a phase I study to determine the safety, activity, and pharmacokinetic characteristics of hepatic arterial infusion of nanoparticle albumin-bound paclitaxel (HAI nab-paclitaxel). Cohorts of three patients having predominant hepatic metastases received HAI nab-paclitaxel at three dose levels (180, 220, and 260 mg/m(2), respectively) infused for more than 1 hour every 3 weeks (3 + 3 design). Some patients participated in comparative pharmacokinetic studies (i.v. vs. HAI), receiving their first course i.v., to determine peak concentrations and effect of first-pass hepatic extraction compared with subsequent courses administered by HAI. The highest dose level was expanded to determine the safety and activity of HAI nab-paclitaxel. Thirty-eight patients were treated. There were no dose-limiting toxicities at doses up to 260 mg/m(2). Common adverse events included alopecia, fatigue, myelosuppresion, nausea, and vomiting. Three patients had stable disease for 4 or more months and 2 patients (1 of 12 with breast cancer and 1 of 1 with cervical cancer) achieved a partial response lasting for 5 and 15 months, respectively. Peak concentrations were lower (∼50%) with greater hepatic extraction of drug (∼42%) following HAI than i.v. infusion based on area under the curve comparison of drug exposure. HAI nab-paclitaxel showed partial hepatic extraction. At doses 260 mg/m(2) or less given for 1 hour every 3 weeks, the treatment was well-tolerated and showed activity in advanced cancer patients with predominant liver metastases.