RESUMO
OBJECTIVE: To develop a reproducible orthotopic model of papillary thyroid carcinoma for the BRAF(V600E) mutation (GenBank NM004333) and an RET/PTC rearrangement (GenBank M31213) that recapitulates the clinical picture in humans. DESIGN: In vitro and in vivo study. SETTING: Department of Head and Neck Surgery, M. D. Anderson Cancer Center. SUBJECTS: Eight- to 12-week-old athymic female nude mice. INTERVENTIONS: Either BRAF-mutated or RET/PTC1-rearranged papillary thyroid carcinoma cells were injected into the thyroid glands of athymic female nude mice. The mice were euthanized when the tumor burden exceeded 1.0 cm or when they exhibited significant morbidity. MAIN OUTCOME MEASURES: Tumorigenicity, extent of tumor invasion and metastasis, cell invasion and migration, and median survival. RESULTS: All the BRAF-mutated cell lines and 1 selected RET/PTC1-rearranged cell line were 100% tumorigenic in mice. These mouse tumor models exhibited a wide range of biological potential, including laryngeal invasion, lymph node metastasis, and pulmonary metastasis, thus reflecting the clinical spectrum of papillary carcinoma. CONCLUSIONS: An orthotopic model of papillary thyroid carcinoma was successfully established in nude mice using BRAF-mutated and RET/PTC1-rearranged cell lines. These models mimic the human disease and will thus be useful for evaluating the clinical potential of novel targeted therapies.
Assuntos
Carcinoma Papilar , Modelos Animais de Doenças , Neoplasias da Glândula Tireoide , Animais , Carcinoma Papilar/genética , Carcinoma Papilar/metabolismo , Carcinoma Papilar/patologia , Ensaios de Migração Celular , Feminino , Imuno-Histoquímica , Técnicas In Vitro , Camundongos , Camundongos Nus , Dados de Sequência Molecular , Invasividade Neoplásica , Proteínas de Fusão Oncogênica/genética , Mutação Puntual , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologiaRESUMO
Headpin (SERPINB13) is a novel member of the serine proteinase inhibitor (Serpin) gene family that was originally cloned from a keratinocyte cDNA library. Western blot analysis using a headpin-specific antiserum recognized a protein with the predicted M(r) of 44kDa in lysates derived from a transformed keratinocyte cell line known to express headpin mRNA. Similarity of the reactive-site loop (RSL) domain of headpin, notably at the P1-P1(') residues, with other serpins that inhibit cysteine and serine proteinases suggests that headpin may inhibit similar proteinases. This study demonstrates that recombinant headpin indeed inhibits cathepsins K and L, but not chymotrypsin, elastase, trypsin, subtilisin A, urokinase-type plasminogen activator, plasmin, or thrombin. The second-order rate constants (k(a)) for the inhibitory reactions of rHeadpin with cathepsins K and L were 5.1+/-0.6x10(4) and 4.1+/-0.8x10(4)M(-1)s(-1), respectively. Headpin formed SDS-stable complexes with cathepsins K and L, a characteristic property of inhibitory serpins. Interactions of the RSL domain of headpin with cathepsins K and L were indicated by cleavage of headpin near the predicted P1-P1(') residues by these proteinases. These results demonstrate that the serpin headpin possesses specificity for inhibiting lysosomal cysteine proteinases.
