Cytotoxic and genotoxic effects of 131 I and 60 Co in follicular thyroid cancer cell (WRO) with and without recombinant human thyroid-stimulating hormone treatment.

Normally, differentiated thyroid cancer (DTC) tends to be biologically indolent, highly curable and has an excellent prognosis. However, the treatment may fail when the cancer has lost radioiodine avidity. The present study was carried out in order to evaluate the cytotoxic and genotoxic effects of 131 I and 60 Co and radioiodine uptake in WRO cells, derived from DTC, harboring the BRAFV600E mutation. WRO cells showed a relatively slow cell cycle of 96.3 h with an unstable karyotype containing various double minutes. The genotoxicity assay (micronucleus test) showed a relative high radioresistance to 131 I (0.07-3.70 MBq/mL), independent of treatment with recombinant human thyroid-stimulating hormone (rhTSH). For the cytotoxicity assay, WRO cells were also relatively resistant to 60 Co (range: 0.2-8.3 Gy), but with a gradual decrease of viability as a function of time for higher doses (20 and 40 Gy, starting from the fifth to sixth day). For internal irradiation with 131 I, WRO cells showed a decline in viability at radioactive concentration higher than 1.85 MBq/mL; this was even more effective at 3.70 MBq/mL, but only when preceded by rhTSH, in coincidence with the highest level of 131 I uptake. These data show promising results, since the loss of the ability of thyroid cells to concentrate radioiodine is considered to be one of the main factors responsible for the failure of 131 I therapy in patients with DTC. The use of tumor-derived cell lines as a model for in vivo tumor requires, however, further investigations and deep evaluation of the corresponding in vivo effects. Environ. Mol. Mutagen. 58:451-461, 2017. © 2017 Wiley Periodicals, Inc.

Physical-chemical and biological characterization of different preparations of equine chorionic gonadotropin.

Ovarian stimulation with commercial preparations of equine chorionic gonadotropin (eCG) produces extremely variable responses in domestic animals, ranging from excessive stimulation to practically no stimulation, when applied on the basis of their declared unitage. This study was conducted to analyze four commercial preparations from different manufacturers via reversed-phase HPLC (RP-HPLC) in comparison with a reference preparation and an official International Standard from the World Health Organization. The peaks obtained by this qualitative and quantitative physical-chemical analysis were compared using an in vivo bioassay based on the ovarian weight gain of prepubertal female rats. The RP-HPLC data showed one or two peaks close to a main peak (tR= 27.9 min), which were related to the in vivo bioactivity. Commercial preparations that have this altered peak showed very little or no in vivo activity, as demonstrated by rat ovarian weight and in peripubertal gilts induced to ovulate. Overall, these findings indicate that RP-HPLC can be a rapid and reliable tool to reveal changes in the physicochemical profile of commercial eCG that is apparently related to decreased biological activity of this hormone.

Cytogenetic and dosimetric effects of (131)I in patients with differentiated thyroid carcinoma: comparison between stimulation with rhTSH and thyroid hormone withdrawal treatments.

A study directed to the cytogenetic and dosimetric aspects of radionuclides of medical interest is very valuable, both for an accurate evaluation of the dose received by the patients, and consequently of the genetic damage, and for the optimization of therapeutic strategies. Cytogenetic and dosimetric effects of (131)I in lymphocytes of thyroidectomized differentiated thyroid cancer (DTC) patients were evaluated through chromosome aberration (CA) technique: Euthyroid patients submitted to recombinant human thyroid-stimulating hormone (rhTSH) therapy (group A) were compared with hypothyroid patients left without levothyroxine treatment (group B). CA analysis was carried out prior to and 24 h, 1 week, 1 month and 1 year after radioiodine administration (4995-7030 MBq) in both groups. An activity-response curve of (131)I (0.074-0.740 MBq/mL) was elaborated, comparing dicentric chromosomes in vivo and in vitro in order to estimate the absorbed dose through Monte Carlo simulations. In general, radioiodine therapy induced a higher total CA rate in hypothyroid patients as compared to euthyroid patients. The frequencies of dicentrics obtained in DTC patients 24 h after treatment were equivalent to those induced in vitro (0.2903 ± 0.1005 MBq/mL in group A and 0.2391 ± 0.1019 MBq/mL in group B), corresponding to absorbed doses of 0.65 ± 0.23 Gy and 0.53 ± 0.23 Gy, respectively. The effect on lymphocytes of internal radiation induced by (131)I therapy is minimal when based on the frequencies of CA 1 year after the treatment, maintaining a higher quality of life for DTC patients receiving rhTSH-aided therapy.

Practical reversed-phase high-performance liquid chromatography method for laboratory-scale purification of recombinant human thyrotropin.

A small, semi-preparative C(4) RP-HPLC column was used to set up the conclusive laboratory-scale purification of Chinese hamster ovary-derived human thyrotropin (hTSH), after a preliminary concentration-purification of an extremely dilute and poorly ( approximately 0.6 microg hTSH/mL; mass fraction=0.35%) conditioned medium on a cation exchanger. Several fractions of this eluate were repeatedly injected on the semi-preparative column, obtaining, in a single run (<1h chromatographic time), a concentrated pool ( approximately 1.2 mg/mL) of highly purified hTSH that could be further concentrated to >3 mg/mL and then efficiently lyophilized. The overall recovery in the rapid RP-HPLC purification step, including concentration and lyophilization, was of the order of 80%. The final product, when tested via a precise, single-dose in vivo bioassay, confirmed that it did not suffer any loss of bioactivity. This same methodology can be easily adapted to the small-scale purification of other recombinant products, even when obtained from genetically modified organisms at extremely low concentrations and mass fractions.