Method of treating cancer with anti-neurotrophin agents
A technology of neurotrophin and protein, which is applied in the field of oncology and neurotrophin, and can solve unsatisfactory problems
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Embodiment 1
[0054] Example 1: Neutralization of trk receptors by anti-NGF
[0055]Antibodies were injected into PC-3 and / or TSU-pr1 xenografts previously shown to respond to CEP-751 (Dionne et al., Clin. Canc. Res., 1998, 4, 187- 1898). The following experiments were performed to confirm the neutralizing ability of anti-neurotrophin antibodies.
[0056] Anti-NGF antibody reduces ligand-stimulated trk autophosphorylation in NIH3T3-trkA cells. NGF (10 ng / ml) and various concentrations of antibodies were pre-incubated in 6 ml tissue culture medium. The NGF / antibody mixture was added to NIH3T3-trkA cells. Trk protein was immunoprecipitated from lysates using the pan-Trk antibody CEP-21, and samples were probed on immunoblots with anti-phosphotyrosine antibody 4G10. Optical density scan values (integer OD units) are as follows (as displayed by NGF (10ng / ml) / anti-NGF (μg / ml)): - / - channel, 0.3; + / - channel, 6.5; 0.001 mg / ml, 5.0; 0.01 mg / ml, 4.5; 0.1 mg / ml, 2.5; 1.0 mg / ml, 3.1; 10.0 mg / m...
Embodiment 2
[0057] Example 2: Neutralization of trk receptors by anti-NT-3
[0058] Anti-NT-3 antibody reduces ligand-stimulated autophosphorylation of trk in NIH3T3-trkC cells. NT-3 (10 ng / ml) and various concentrations of antibodies were pre-incubated in 6 ml tissue culture medium. The NT-3 / antibody mixture was added to NIH3T3-trkC cells. Trk protein was immunoprecipitated from lysates using the pan-Trk antibody CEP-21, and samples were probed on immunoblots with anti-phosphotyrosine antibody 4G10. Densitometric scan values (integer OD units) are as follows (shown as NT-3 (10 ng / ml) / anti-NT-3 (μg / ml)): - / - lane, 0.1; + / - lane, 4.0; IgG, 7.5 ; 0.001 mg / ml, 6.2; 0.01 mg / ml, 4.0; 0.1 mg / ml, 3.6; 1.0 mg / ml, 7.8; 10.0 mg / ml, 1.2. Thus, in cells treated with 10 ng / ml NT-3 for 5 minutes, anti-NT-3 (Pepro Tech 500-P82) at a concentration of 10 μg / ml reduced trk phosphorylation by approximately 70% relative to no anti-NGF treatment.
[0059] The following examples show the results of exper...
Embodiment 3
[0060] Example 3: Inhibition of PC-3 Prostate Cancer Xenograft Growth by Neurotrophin Antibodies
[0061] The following neurotrophin antibodies were used: anti-NGF (Pepro Tech 500-P85), anti-BDNF (Pepro Tech 500-P84), anti-NT-3 (Pepro Tech 500-P82), anti-NT-4 / 5 (Pepro Tech 500- P83). Anti-NGF and anti-NT3 antibodies blocked trkA and trkC autophosphorylation after neurotrophin treatment of cultured cells. Individual antibodies have been shown to block the activity of their cognate neurotrophins in bioassays in which ChAT activity was measured in rat forebrain basal cell cultures.
[0062] PC-3 human prostate tumor cells (5×10 6 cells / mouse) were injected subcutaneously into the flank of 8-10 week old female athymic nude mice (nu / nu; Charles River, Raleigh, NC). On the day of tumor implantation, mice weighed 22-25 g. In xenograft tumors grow to 100-500mm 3 Afterwards, the mice were randomly divided into experimental groups. Some experimental groups were administered steril...
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