Vestibular Schwannoma Hypofractionated Stereotactic Radiation Therapy in Five Fractions.

AIM: To retrospectively analyse the long-term results of hypofractionated stereotactic radiation therapy (HSRT) applied in five fractions for vestibular schwannomas. MATERIALS AND METHODS: One hundred and thirty-four patients with vestibular schwannomas underwent medical treatment of HSRT. The median follow-up time interval was 54 months (range 6-121 months). All patients had a prescribed dose of 22 Gy in five fractions to D90. Restaging was carried out by thin-slice contrast-enhanced T1 magnetic resonance imaging. Progression was defined as 2 mm post-treatment tumour enlargement. Progression or death for any reason was counted as an event in progression-free survival rates. Acute toxicity was defined as adverse events occurring within 3 months of HSRT; long-term toxicity was defined as such events occurring after 3 months. RESULTS: In 74/128 patients who had >6 months of follow-up (54%), the HSRT resulted in a partial or a complete response. The mean time interval for response in 50% of these was 4 years, whereas in 49 patients (38%) vestibular schwannomas failed to show any response, resulting in stable disease. Five of 128 patients (4%) showed marked progressive vestibular schwannomas after treatment in the first 3 years; two of them received conventionally fractionated radiation therapy. Local control at 3, 5 and 7 years was 96%, 95% and 94%, respectively. Seven were lost to follow-up. The median planning target volume was 2.1 ml (range 0.78-8.66). The 3- and 5-year progression-free survival rates were 95% and 94%, respectively. Seven patients reported a marked deterioration in hearing ability. Post-radiation therapy magnetic resonance imaging showed variability in oedema collection, but no patient suffered from radio-necrosis. Grade 2 temporary facial nerve disorders were observed in 10 patients (8%) 3-6 months after HSRT. CONCLUSION: Delivering HSRT in five fractions for vestibular schwannoma appears safe and efficient, combining both efficiency and short treatment time while optimising neurological function preservation.

Combined effects of resveratrol and radiation in GH3 and TtT/GF pituitary adenoma cells.

OBJECTIVE: Resveratrol and radiation decrease viability in various tumor cells. This study aims to investigate combined effects of resveratrol and radiation on viability, induction of apoptosis and necrosis, and expression of apoptosis modulators in rodent GH3 and TtT/GF pituitary adenoma cells in vitro. METHODS: Cells were incubated with 10-100 µM resveratrol. Medium and medium with ethanol served as controls. After 2 h, cells were irradiated with 0-5 Gray (Gy) and further incubated for 48-72 h. Cell viability was quantified using a hemocytometer. Cell death was assessed with an enzyme-linked immunosorbent assay (ELISA) that detects free nucleosomes in cell lysates and free nucleosomes released to the culture medium. Expression of B-cell lymphoma-2 protein (BCL-2) and BCL-2 associated Xprotein (BAX) was measured using quantitative real time-polymerase chain reaction (qRT-PCR) to analyze changes in BAX/BCL-2 ratio. RESULTS: Resveratrol and irradiation with 4 Gy alone and in combination significantly decreased cell viability (p = 0.017 and less). In the ELISA, 10 µM resveratrol significantly induced apoptosis in TtT/GF cells at 0 Gy (p < 0.001), but not at 3 or 5 Gy. In the ELISA, 10 µM resveratrol significantly induced necrosis in GH3 cells at 0, 3 and 5 Gy (p < 0.001). While qRT-PCR did not demonstrate a significant effect of 10 µM resveratrol or radiation on expression of BAX or BCL-2, a significant increase in the BAX/BCL-2 ratio was found after irradiation with 5 Gy in GH3 cells (p = 0.0027). CONCLUSION: While moderate irradiation solely led to inhibited proliferation, resveratrol induced cell death in rodent pituitary adenoma cells.

A simple measure of reliability in clinical trials of surgical adjuvant radiotherapy.

AIMS: The 'number needed to treat' (NNT) is a measure of therapeutic effect and is defined as the number of patients needed to be treated, so that the treatment of one of them is successful. Since many patients or physicians find this way to describe therapeutic potency very intuitive, I calculated the NNTs for the surgical adjuvant radiotherapy of different malignant neoplasias. METHODS: This computation was applied to selected, prospective, randomized trials or meta-analyses on adjuvant or adjunctive radiotherapy of epithelial or glial malignancies. RESULTS: Among 17 NNTs calculated for trials reporting improvements in local relapse rates 2 are smaller than 5, 8 are in the range from 5 to 10 and 7 are larger than 10 (median follow-up 5 years; range 3-20). In contrast, only 2 NNTs from 10 trials reporting survival advantages are smaller than 10 (median follow-up 4 years; range 1-20). CONCLUSIONS: NNTs correctly reflect that adjuvant radiotherapy has a far more pronounced impact on the probability of local recurrence of a malignancy as opposed to patient's survival.

Modulation of prion protein structural integrity by geldanamycin.

The cellular prion protein PrPc is of crucial importance for the development of neurodegenerative diseases called transmissible spongiform encephalopathies. We investigated if the function of members of the HSP90 family is required for the integrity of the normal, nonpathogenic prion protein called PrPc. Eukaryotic cells were treated with the structurally unrelated HSP90-inhibitors geldanamycin (GA) or radicicol (RC). In either case the cellular prion protein was induced and exhibited faster migrating bands on western blot analysis, whereas geldampicin (GE), an analog of GA known not to bind to HSP90, had no effect. Ongoing protein and messenger RNA synthesis during treatment were found to be necessary for the appearance of these bands. Cotreatment with tunicamycin abrogated any effect of HSP90 inhibitors on the cellular prion protein. Finally, enzymatic deglycosylation with peptide:N-glycosidase F of the normal prion protein as well as the variant induced by benzoquinone ansamycins resulted in very similar band patterns. These experiments indicate that either altered glycosylation, or a change in conformation, or both are involved in the induction of faster migrating bands by HSP90 inhibitors. Thus the inhibition of the function of members of the HSP90 family of molecular chaperones results in profound changes in the physicochemical properties of PrPc.

Heat-shock protein 90: potential involvement in the pathogenesis of malignancy and pharmacological intervention.

Heat-shock protein 90 (Hsp90) is an essential, cytosolic protein. Its overexpression in a wide variety of malignant tumors makes it a candidate target for pharmacological intervention. The association with Hsp90 stabilizes key regulatory proteins like Fak, Bcr-Abl, ErbB2, mutant p53 and Raf-1. The disruption of these heterocomplexes by Hsp90 inhibitors causes the rapid degradation of Hsp90-client proteins by the proteasome. Benzoquinone ansamycins were the first group of compounds for which interference with Hsp90 function was shown to be the major mechanism of action. They are in the early phase of clinical development. Radicicol and its derivatives are functional analogues of benzoquinone ansamycins without structural similarity. Flavonoids and stresgenin B share the ability to suppress heat-shock protein synthesis. Recently, it became apparent that coumarin antibiotics, cisplatin and paclitaxel also bind to Hsp90. The clinical value of the newly characterized agents with activity towards Hsp90 remains to be determined.

The benzoquinone ansamycin geldanamycin stimulates proteolytic degradation of focal adhesion kinase.

FAK is a nonreceptor tyrosine kinase involved in adhesion-mediated signal transduction whose level of expression is related to the invasiveness of malignant tumors. In seeking strategies to downregulate FAK, we treated various cell lines in vitro with the benzoquinone ansamycin geldanamycin (GA) which was previously described as a tyrosine kinase inhibitor, but recently has been shown to exert its effects by interfering with the chaperone function of members of the hsp90 family of heat-shock proteins. We evaluated the effects of benzoquinone ansamycins on FAK steady-state protein level and FAK half-life in breast and prostate carcinoma, Ewing's sarcoma, and 3T3 fibroblasts. Our data demonstrate that GA stimulates the proteolytic degradation of FAK in all cell lines examined and markedly reduces the half-life of newly synthesized FAK protein without significantly altering the level of FAK mRNA. These data demonstrate FAK to be another tyrosine kinase sensitive to the destabilizing effects of benzoquinone ansamycins and further show that small molecule-mediated pharmacologic modulation of FAK protein level is a feasible approach to the interdiction of FAK function.

The amino-terminal domain of heat shock protein 90 (hsp90) that binds geldanamycin is an ATP/ADP switch domain that regulates hsp90 conformation.

Many functions of the chaperone, heat shock protein 90 (hsp90), are inhibited by the drug geldanamycin that specifically binds hsp90. We have studied an amino-terminal domain of hsp90 whose crystal structure has recently been solved and determined to contain a geldanamycin-binding site. We demonstrate that, in solution, drug binding is exclusive to this domain. This domain also binds ATP linked to Sepharose through the gamma-phosphate. Binding is specific for ATP and ADP and is inhibited by geldanamycin. Mutation of four glycine residues within two proposed ATP binding motifs diminishes both geldanamycin binding and the ATP-dependent conversion of hsp90 to a conformation capable of binding the co-chaperone p23. Since p23 binding requires regions outside the 1-221 domain of hsp90, these results indicate a common site for nucleotides and geldanamycin that regulates the conformation of other hsp90 domains.