Internalization of the radioiodinated somatostatin analog [125I-Tyr3]octreotide by mouse and human pituitary tumor cells: increase by unlabeled octreotide
Recently, we developed a technique that allows the in vivo visualization
in man of somatostatin receptor-positive neuroendocrine tumors after i.v.
injection of [125I-Tyr3]octreotide or [111In-DTPA-D-Phe1]octreotide.
Radiotherapy of such tumors using somatostatin analogs coupled to alpha-
or beta-emitting radionuclides has been proposed as an application for
radiolabeled somatostatin analogs. To develop this concept further, it is
of importance to know whether the above-mentioned radiolabeled
somatostatin analogs are internalized by the tumor cells, and whether it
might be possible to manipulate the degree of internalization. In the
present study we investigated the internalization of a stable somatostatin
analog, [125I-Tyr3]octreotide, by mouse AtT20/D16V pituitary tumor cells
and primary cultures of human GH-secreting pituitary tumor cells.
Treatment of the cells with low pH was used to distinguish between
membrane-bound (acid-releasable) and internalize (acid-resistant)
radioligand. [125I-Tyr3]octreotide showed a time-dependent increasing
accumulation in AtT20 cells; after 4 h of incubation, values up to 6-8% of
the dose of radioligand added were obtained. Binding and internalization
of [125I-Tyr3]octreotide were temperature dependent and inhibited by
pertussis toxin. Inhibitors of lysosomal degradation did not increase the
amount of internalized radioligand. After 4 h of incubation, 88% of the
radioactivity present in the cells was still peptide bound, suggesting a
low intracellular breakdown of this radioligand. Six of seven human
GH-secreting adenoma cell cultures also internalized [125I-Tyr3]octreotide
(variation between 0.24-4.98% of the dose radioligand added). Displacement
of binding and internalization of [125I-Tyr3]octreotide by unlabeled
octreotide showed a bell-shaped curve in AtT20 cells. At low
concentrations (0.1 and 1 nM), binding and internalization were increased,
whereas at higher concentrations, saturation occurred. In contrast to
this, binding of [125I-Tyr3]octreotide to a broken cell preparation of
AtT20 cells was displaced in a dose-dependent manner by unlabeled
octreotide, with an IC50 of 0.1 nM. Similar observations were made in the
human GH-secreting adenoma cell cultures. In conclusion, a high amount of
[125I-Tyr3]octreotide is internalized in a specific-, time-, temperature-,
and pertussis toxin-sensitive GTP-binding protein-dependent manner by
mouse AtT20 and human GH-secreting pituitary tumor cells. In the presence
of a low concentration of unlabeled octreotide, a rapid increase in the
amount of [125I-Tyr3]octreotide internalized by AtT20 cells and by the
majority of the human GH-secreting adenoma cell cultures was
found.(ABSTRACT TRUNCATED AT 400 WORDS
