sirolimus and alovudine

Mammalian target of rapamycin (mTOR) is a new promising oncological target. However, most clinical studies reported only modest antitumor activity during mTOR-targeted monotherapies, including studies in osteosarcomas, emphasizing a need for improvement. We hypothesized that the combination with rationally selected other therapeutic agents may improve response. In this study, we examined the efficacy of the mTOR inhibitor temsirolimus combined with cisplatin or bevacizumab on the growth of human osteosarcoma xenografts (OS-33 and OS-1) in vivo, incorporating functional imaging techniques and microscopic analyses to unravel mechanisms of response. In both OS-33 and OS-1 models, the activity of temsirolimus was significantly enhanced by the addition of cisplatin (TC) or bevacizumab (TB). Extensive immunohistochemical analysis demonstrated apparent effects on tumor architecture, vasculature, apoptosis and the mTOR-pathway with combined treatments. 3'-Deoxy-3'-(18) F-fluorothymidine ((18) F-FLT) positron emission tomography (PET) scans showed a remarkable decrease in (18) F-FLT signal in TC- and TB-treated OS-1 tumors, which was already noticeable after 1 week of treatment. No baseline uptake was observed in the OS-33 model. Both immunohistochemistry and (18) F-FLT-PET demonstrated that responses as determined by caliper measurements underestimated the actual tumor response. Although (18) F-FLT-PET could be used for accurate and early response monitoring for temsirolimus-based therapies in the OS-1 model, we could not evaluate OS-33 tumors with this molecular imaging technique. Further research on the value of the use of (18) F-FLT-PET in this setting in osteosarcomas is warranted. Overall, these findings urge the further exploration of TC and TB treatment for osteosarcoma (and other cancer) patients.

Topics: Angiogenesis Inhibitors; Animals; Antibodies, Monoclonal, Humanized; Antineoplastic Combined Chemotherapy Protocols; Bevacizumab; Bone Neoplasms; Cell Line, Tumor; Cisplatin; Dideoxynucleosides; Female; Fluorodeoxyglucose F18; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Osteosarcoma; Positron-Emission Tomography; Protein Kinase Inhibitors; Sirolimus; Tomography, X-Ray Computed; TOR Serine-Threonine Kinases; Xenograft Model Antitumor Assays

The prognosis of relapsed or refractory aggressive lymphoma is poor. The huge variety of currently evolving targeted treatment approaches would benefit from tools for early prediction of response or resistance. We used various ALK-positive anaplastic large cell lymphoma (ALCL) cell lines to evaluate two inhibitors, the HSP90 inhibitor NVP-AUY922, and the mTOR inhibitor everolimus, both of which have shown to interfere with ALK-dependent oncogenic signal transduction. Their therapeutic effect was determined in vitro by MTT assay, [(18)F]fluorodeoxyglucose (FDG)- and [(18)F]fluorothymidine (FLT)-uptake, and by biochemical analysis of ALK-induced signaling. Micro-FDG- and FLT-positron emission tomography (PET) imaging studies in immunodeficient mice bearing ALCL xenotransplants were carried out with the cell lines SUDHL-1 and Karpas299 to assess early treatment response to NVP-AUY922 or everolimus in vivo. SUDHL-1 cells showed sensitivity to both inhibitors in vitro. Importantly, we detected a significant reduction of FLT-uptake in SUDHL-1 bearing animals using both inhibitors compared with baseline as early as 5 days after initiation of targeted therapy. Immunostaining showed a decrease in Ki-67 and an increase in cleaved caspase-3 staining. In contrast, FDG-uptake did not significantly decrease at early time points. Karpas299 xenotransplants, which are resistant to NVP-AUY922 and sensitive to everolimus treatment, showed an increase of mean FLT-uptake on day 2 after administration of NVP-AUY299, but a significant reduction in FLT-uptake upon everolimus treatment. In conclusion, we show that FLT-PET but not FDG-PET is able to predict response to treatment with specific inhibitors very early in the course of treatment and thus enables early prediction of treatment efficacy.

Topics: Animals; Blotting, Western; Cell Line; Cell Line, Tumor; Cell Survival; Dideoxynucleosides; Enzyme Inhibitors; Everolimus; Female; Flow Cytometry; Fluorine Radioisotopes; Fluorodeoxyglucose F18; HSP90 Heat-Shock Proteins; Humans; Isoxazoles; Lymphoma, Large-Cell, Anaplastic; Mice; Mice, SCID; Positron-Emission Tomography; Protein-Tyrosine Kinases; Resorcinols; Sirolimus; TOR Serine-Threonine Kinases; Treatment Outcome; Tumor Burden; Xenograft Model Antitumor Assays

Targeting the mammalian target of rapamycin (mTOR) pathway is a potential means of overcoming cisplatin resistance in ovarian cancer patients. Because mTOR inhibition affects cell proliferation, we aimed to study whether 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) PET could be useful for monitoring early response to treatment with mTOR inhibitors in an animal model of cisplatin-resistant ovarian tumor.. BALB/c nude mice bearing subcutaneous human SKOV3 ovarian cancer xenografts were treated with either the mTOR inhibitor everolimus (5 mg/kg) or vehicle, and (18)F-FLT PET was performed at baseline, day 2, and day 7 of treatment. (18)F-FLT uptake was evaluated by calculation of mean standardized uptake value (SUVmean) corrected for partial-volume effect. Ex vivo immunohistochemistry studies were performed on separate cohorts of mice treated as above and sacrificed at the same time points as for the PET studies. The ex vivo analysis included bromodeoxyuridine incorporation as a marker of cell proliferation, and phosphorylation of ribosomal protein S6 as a downstream marker of mTOR activation.. During the treatment period, no significant change in tumor (18)F-FLT uptake was observed in the vehicle group, whereas in everolimus-treated mice, (18)F-FLT SUVmean decreased by 33% (P = 0.003) at day 2 and 66% (P < 0.001) at day 7, compared with baseline. Notably, the reduction of (18)F-FLT uptake observed at day 2 in the everolimus group preceded changes in tumor volume, and a significant difference in (18)F-FLT uptake was observed between vehicle and drug-treated tumors at both day 2 (P = 0.0008) and day 7 (P = 0.01). In ex vivo studies, everolimus treatment resulted in a 98% reduction in phosphorylated ribosomal protein S6 immunostaining at day 2 (P = 0.02) and 91% reduction at day 7 (P = 0.003), compared with the vehicle group. Bromodeoxyuridine incorporation was reduced by 65% at day 2 (not significant) and by 41% at day 7 (P = 0.02) in drug versus vehicle groups.. Reduction in (18)F-FLT uptake correlates well with the level of mTOR inhibition by everolimus in the SKOV3 ovarian tumor model. These data suggest that early treatment monitoring by (18)F-FLT PET may be of use in future preclinical or clinical trials evaluating treatment of cisplatin-resistant ovarian tumors by mTOR inhibitors.

Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Cisplatin; Dideoxynucleosides; Disease Models, Animal; Drug Resistance, Neoplasm; Everolimus; Female; Humans; Mice; Mice, Nude; Ovarian Neoplasms; Positron-Emission Tomography; Radiopharmaceuticals; Reproducibility of Results; Sensitivity and Specificity; Sirolimus; TOR Serine-Threonine Kinases; Treatment Outcome

To be a reliable predictor of response, tracer uptake should reflect changes in the amount of active tumor cells. However, uptake of (18)F-FDG, the most commonly used PET tracer, is disturbed by the inflammatory cells that appear early after cytotoxic therapy. The first aim of this study was to investigate whether 3'-(18)F-fluoro-3'-deoxy-l-thymidine ((18)F-FLT), a marker of cellular proliferation, is a better tracer for response assessment early after cytotoxic therapy. A second objective of this study was to investigate whether (18)F-FDG and (18)F-FLT responses were comparable early after mammalian target of rapamycin (mTOR) inhibition, as an example of proliferation-targeting therapies.. Severe combined immunodeficient mice were subcutaneously inoculated with Granta-519 cells, a human cell line derived from a leukemic mantle cell lymphoma. Half the mice were treated with cyclophosphamide and the other half with mTOR inhibition. (18)F-FDG and (18)F-FLT uptake was evaluated by small-animal PET on day 0 (D0; before treatment), D+1, D+2, D+4, D+7, D+9, D+11, and D+14. At each time point, 2 mice of each treatment condition were sacrificed, and tumors were excised for histopathology.. After cyclophosphamide, (18)F-FDG and (18)F-FLT uptake decreased, with a maximum reduction of -29% for (18)F-FDG and -25% for (18)F-FLT uptake at D+2, compared with baseline. Although (18)F-FDG uptake increased from D+4 on, with a maximum on D+7, (18)F-FLT uptake remained virtually stable. Histology showed an increase in apoptotic or necrotic tumor fraction, followed by an influx of inflammatory cells. In mTOR-inhibited mice, (18)F-FDG uptake dropped until D+2 after therapy (-43%) but increased at D+4 (-27%) to form a plateau on D+7 and D+9 (-14% and -16%, respectively). Concurrently, (18)F-FLT uptake decreased to -31% on D+2, followed by an increase with a peak value of +12% on D+7, after which (18)F-FLT uptake decreased again. Cyclin D1 expression dropped from D+1 until D+4 and returned to baseline at D+7.. Because (18)F-FLT uptake is not significantly influenced by the temporary rise in inflammatory cells early after cyclophosphamide, it more accurately reflects tumor response. However, a formerly unknown temporary rise in (18)F-FLT uptake a few days after the administration of mTOR inhibition was defined, which makes it clear that drug-specific responses have to be considered when using PET for early treatment monitoring.

Topics: Animals; Antineoplastic Agents, Alkylating; Cell Line, Tumor; Cyclophosphamide; Dideoxynucleosides; Fluorodeoxyglucose F18; Humans; Lymphoma; Metabolic Clearance Rate; Mice; Mice, SCID; Protein Kinase Inhibitors; Protein Kinases; Radionuclide Imaging; Radiopharmaceuticals; Sirolimus; Tissue Distribution; TOR Serine-Threonine Kinases; Treatment Outcome