RESUMO
The G1 restriction point is critical for regulating the cell cycle and is controlled by the Rb pathway (CDK4/6-cyclin D1-Rb-p16/ink4a). This pathway is important because of its inactivation in a majority of human tumors. Transition through the restriction point requires phosphorylation of retinoblastoma protein (Rb) by CDK4/6, which are highly validated cancer drug targets. We present the identification and characterization of a potent CDK4/6 inhibitor, LY2835219. LY2835219 inhibits CDK4 and CDK6 with low nanomolar potency, inhibits Rb phosphorylation resulting in a G1 arrest and inhibition of proliferation, and its activity is specific for Rb-proficient cells. In vivo target inhibition studies show LY2835219 is a potent inhibitor of Rb phosphorylation, induces a complete cell cycle arrest and suppresses expression of several Rb-E2F-regulated proteins 24 hours after a single dose. Oral administration of LY2835219 inhibits tumor growth in human tumor xenografts representing different histologies in tumor-bearing mice. LY2835219 is effective and well tolerated when administered up to 56 days in immunodeficient mice without significant loss of body weight or tumor outgrowth. In calu-6 xenografts, LY2835219 in combination with gemcitabine enhanced in vivo antitumor activity without a G1 cell cycle arrest, but was associated with a reduction of ribonucleotide reductase expression. These results suggest LY2835219 may be used alone or in combination with standard-of-care cytotoxic therapy. In summary, we have identified a potent, orally active small-molecule inhibitor of CDK4/6 that is active in xenograft tumors. LY2835219 is currently in clinical development.
Assuntos
Aminopiridinas/farmacologia , Antineoplásicos/farmacologia , Benzimidazóis/farmacologia , Quinase 4 Dependente de Ciclina/antagonistas & inibidores , Quinase 6 Dependente de Ciclina/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Aminopiridinas/uso terapêutico , Animais , Antineoplásicos/uso terapêutico , Benzimidazóis/uso terapêutico , Linhagem Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Desoxicitidina/uso terapêutico , Quimioterapia Combinada , Feminino , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Camundongos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologia , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/uso terapêutico , Proteína do Retinoblastoma/antagonistas & inibidores , Proteína do Retinoblastoma/metabolismo , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , GencitabinaRESUMO
Expression of eukaryotic translation initiation factor 4E (eIF4E) is commonly elevated in human and experimental cancers, promoting angiogenesis and tumor growth. Elevated eIF4E levels selectively increase translation of growth factors important in malignancy (e.g., VEGF, cyclin D1) and is thereby an attractive anticancer therapeutic target. Yet to date, no eIF4E-specific therapy has been developed. Herein we report development of eIF4E-specific antisense oligonucleotides (ASOs) designed to have the necessary tissue stability and nuclease resistance required for systemic anticancer therapy. In mammalian cultured cells, these ASOs specifically targeted the eIF4E mRNA for destruction, repressing expression of eIF4E-regulated proteins (e.g., VEGF, cyclin D1, survivin, c-myc, Bcl-2), inducing apoptosis, and preventing endothelial cells from forming vessel-like structures. Most importantly, intravenous ASO administration selectively and significantly reduced eIF4E expression in human tumor xenografts, significantly suppressing tumor growth. Because these ASOs also target murine eIF4E, we assessed the impact of eIF4E reduction in normal tissues. Despite reducing eIF4E levels by 80% in mouse liver, eIF4E-specific ASO administration did not affect body weight, organ weight, or liver transaminase levels, thereby providing the first in vivo evidence that cancers may be more susceptible to eIF4E inhibition than normal tissues. These data have prompted eIF4E-specific ASO clinical trials for the treatment of human cancers.
Assuntos
Fator de Iniciação 4E em Eucariotos/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias/metabolismo , Neoplasias/terapia , Biossíntese de Proteínas/genética , Animais , Apoptose , Sequência de Bases , Células Cultivadas , Células Endoteliais/metabolismo , Fator de Iniciação 4E em Eucariotos/genética , Humanos , Camundongos , Neoplasias/irrigação sanguínea , Neoplasias/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Activation of protein kinase Cbeta (PKCbeta) has been repeatedly implicated in tumor-induced angiogenesis. The PKCbeta-selective inhibitor, Enzastaurin (LY317615.HCl), suppresses angiogenesis and was advanced for clinical development based upon this antiangiogenic activity. Activation of PKCbeta has now also been implicated in tumor cell proliferation, apoptosis, and tumor invasiveness. Herein, we show that Enzastaurin has a direct effect on human tumor cells, inducing apoptosis and suppressing the proliferation of cultured tumor cells. Enzastaurin treatment also suppresses the phosphorylation of GSK3betaser9, ribosomal protein S6(S240/244), and AKT(Thr308). Oral dosing with Enzastaurin to yield plasma concentrations similar to those achieved in clinical trials significantly suppresses the growth of human glioblastoma and colon carcinoma xenografts. As in cultured tumor cells, Enzastaurin treatment suppresses the phosphorylation of GSK3beta in these xenograft tumor tissues. Enzastaurin treatment also suppresses GSK3beta phosphorylation to a similar extent in peripheral blood mononuclear cells (PBMCs) from these treated mice. These data show that Enzastaurin has a direct antitumor effect and that Enzastaurin treatment suppresses GSK3beta phosphorylation in both tumor tissue and in PBMCs, suggesting that GSK3beta phosphorylation may serve as a reliable pharmacodynamic marker for Enzastaurin activity. With previously published reports, these data support the notion that Enzastaurin suppresses tumor growth through multiple mechanisms: direct suppression of tumor cell proliferation and the induction of tumor cell death coupled to the indirect effect of suppressing tumor-induced angiogenesis.
Assuntos
Apoptose/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Glioblastoma/tratamento farmacológico , Indóis/farmacologia , Proteína Quinase C/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Animais , Processos de Crescimento Celular/efeitos dos fármacos , Neoplasias do Colo/enzimologia , Neoplasias do Colo/patologia , Feminino , Glioblastoma/enzimologia , Glioblastoma/patologia , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Células HCT116 , Humanos , Masculino , Camundongos , Camundongos Nus , Fosforilação/efeitos dos fármacos , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/patologia , Proteína Quinase C beta , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Proteína S6 Ribossômica/antagonistas & inibidores , Proteína S6 Ribossômica/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The progression of human prostate cancer from the initial androgen-dependent phase to androgen independence involves diminished apoptosis and a release from the cell cycle block triggered by androgen ablation therapy. FOXO transcription factors play a central role in promoting expression of proapoptotic and cell cycle regulatory genes (e.g., FasL and p27KIP1). Reduced FOXO function might, therefore, play a role in androgen-independent progression of human prostate cancer. Herein, we show that FOXO function is compromised in androgen-independent prostate cancer cells (LNAI) versus androgen-dependent LNCaP cells. The FOXO3a protein, the most highly expressed FOXO family member in prostate cancer cells, is hyperphosphorylated in LNAI cells. FOXO3a expression is also markedly reduced in these androgen-independent LNAI cells when compared with parental LNCaP cells. Together, reduced FOXO3a expression coupled to FOXO3a hyperphosphorylation would suppress FOXO transcriptional activity. Accordingly, activity of the FOXO-responsive p27KIP1 promoter is reduced 60% in these LNAI cells when compared with LNCaP cells. Moreover, mutation of a conserved FOXO response element suppresses p27KIP1 promoter activity, substantiating a regulatory role for this FOXO response element in p27KIP1 promoter transactivation. Finally, we show that the activity of a distinct FOXO-responsive promoter, the 3X-IRS promoter, is also reduced in LNAI cells. Collectively, these data show that reduced FOXO3a expression coupled to increased FOXO3a phosphorylation coincide with reduced FOXO-responsive promoter activity in androgen-independent LNAI cells when compared with androgen-dependent LNCaP cells. To the extent that this model reflects human disease, these data suggest that FOXO function may be compromised with androgen-independent progression of human prostate cancer.
Assuntos
Androgênios/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ligação a DNA/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Fatores de Transcrição/metabolismo , Ativação Transcricional , Proteínas Supressoras de Tumor/genética , Western Blotting , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p27 , Progressão da Doença , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead , Humanos , Masculino , Fosforilação , Plasmídeos/metabolismo , Regiões Promotoras Genéticas , Neoplasias da Próstata/genética , Elementos de Resposta , Transdução de Sinais , Transfecção , Proteínas Supressoras de Tumor/metabolismoRESUMO
PURPOSE: Angiogenin is a polypeptide involved in the formation and establishment of new blood vessels necessary for growth and metastasis of numerous malignant neoplasms, including prostatic adenocarcinoma. Antiangiogenin therapy inhibits the establishment, growth, and metastasis of prostatic adenocarcinoma in animal studies. In this study, we have investigated the expression of angiogenin in prostatic adenocarcinoma, high-grade prostatic intraepithelial neoplasia, and adjacent benign prostatic epithelium in a large cohort of prostatectomy specimens. METHODS: We have studied the expression of angiogenin by immunohistochemistry in prostatic adenocarcinoma, high-grade prostatic intraepithelial neoplasia, and adjacent benign prostatic tissue in 107 human total prostatectomy specimens. RESULTS: The percentage of cells staining positively for angiogenin in benign prostatic glandular epithelium (mean = 17%) was significantly less than for high-grade prostatic intraepithelial neoplasia (mean = 58%, P < 0.001) and prostatic adenocarcinoma (mean = 60%, P < 0.001). Compared with adjacent benign prostatic epithelium, the staining intensity was significantly greater in high-grade prostatic intraepithelial neoplasia (P < 0.001) and prostatic adenocarcinoma (P < 0.001). Furthermore, staining intensity has significantly stronger in prostatic adenocarcinoma versus high-grade prostatic intraepithelial neoplasia (P = 0.0023). However, there was no correlation of angiogenin expression with various clinical and pathologic variables examined, including age at surgery, Gleason scores, pathologic stage, tumor extent, angiolymphatic invasion, extraprostatic extension, seminal vesical invasion, lymph node metastasis, surgical margin status, presence of prostatic intraepithelial neoplasia, and perineural invasion. CONCLUSION: Angiogenin expression in prostatic tissue increases as prostatic epithelial cells evolve from a benign to an invasive phenotype. The increasing expression of prostatic adenocarcinoma in the progression from benign prostate to high-grade prostatic intraepithelial neoplasia and ultimately to prostatic adenocarcinoma are consistent with previous studies showing the influential role that angiogenin plays in the growth, invasion, and metastasis of prostatic adenocarcinoma and many other malignant tumors.
Assuntos
Neoplasias da Próstata/metabolismo , Ribonuclease Pancreático/metabolismo , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Adenocarcinoma/cirurgia , Adulto , Idoso , Ensaio de Imunoadsorção Enzimática , Humanos , Metástase Linfática/patologia , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/patologia , Prostatectomia , Hiperplasia Prostática/metabolismo , Hiperplasia Prostática/patologia , Hiperplasia Prostática/cirurgia , Neoplasia Prostática Intraepitelial/metabolismo , Neoplasia Prostática Intraepitelial/patologia , Neoplasia Prostática Intraepitelial/cirurgia , Neoplasias da Próstata/patologia , Neoplasias da Próstata/cirurgia , Resultado do TratamentoRESUMO
Trioxifene (LY133314) is a selective estrogen receptor modulator (SERM) with competitive binding activity against estradiol for estrogen receptor alpha (ERalpha) and antagonistic activity against ERalpha-mediated gene expression. The PAIII rat prostatic adenocarcinoma (PCa) is an androgen receptor-negative, ERalpha- and ERbeta-positive, spontaneously metastatic rodent tumor cell line. After s.c. implantation of 10(6) PAIII cells in the tail, s.c. administration of trioxifene (2.0, 4.0, 20.0, or 40.0 mg/kg-day) for 30 days produced significant (P < 0.05) inhibition of PAIII metastasis from the primary tumor in the tail to the gluteal and iliac lymph nodes (maximum nodal weight decreases, 86% and 88% from control values, respectively). PAIII metastasis to the lungs was significantly inhibited by trioxifene treatment. Numbers of pulmonary foci in PAIII-bearing rats were significantly (P < 0.05) reduced by trioxifene administration in a dose-related manner (maximal reduction, 98% from control values). Continual administration of the compound significantly (P < 0.05) extended survival of PAIII-bearing rats. Trioxifene inhibited the proliferation of PAIII cells at micromolar levels in vitro but did not slow growth of the primary tumor growth in the tail. Trioxifene administration also produced regression of male accessory sex organs. In PAIII-tumor-bearing animals, trioxifene administration produced a maximal regression of 76% for ventral prostate and 64% for seminal vesicle (P < 0.05 for both). SERMs may be preferable to estrogens given their efficacy in experimental PCa models and relative lack of side effects observed in clinical trials. Our data support the contention that trioxifene represents a SERM with potential antimetastatic efficacy for the treatment of androgen-independent, metastatic PCa.
Assuntos
Adenocarcinoma/tratamento farmacológico , Neoplasias da Próstata/tratamento farmacológico , Pirrolidinas/farmacologia , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Peso Corporal/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Modelos Animais de Doenças , Receptor alfa de Estrogênio , Receptor beta de Estrogênio , Genitália Masculina/efeitos dos fármacos , Luciferases/antagonistas & inibidores , Luciferases/metabolismo , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Metástase Linfática , Masculino , Tamanho do Órgão/efeitos dos fármacos , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Pirrolidinas/metabolismo , Ratos , Receptores Androgênicos/biossíntese , Receptores de Estrogênio/biossíntese , Receptores de Estrogênio/metabolismo , Moduladores Seletivos de Receptor Estrogênico/metabolismo , Testículo/anatomia & histologia , Testículo/efeitos dos fármacosRESUMO
BACKGROUND: PSA mediates growth factor responses that stimulate proliferation of prostatic and other cellular types. Androgen-sensitive TE85 human osteosarcoma cells were used to study PSA as a potential mediator of prostatic cancer growth and osseous metastasis. MATERIALS AND METHODS: TE85 cells were probed for PSA mRNA and protein levels under testosterone (T)-replete and--depleted conditions. TE85 proliferative responses to PSA were evaluated in the absence and presence of LY312340, a monocyclic beta-lactam inhibitor of PSA enzymatic activity. RESULTS: A 3.1-fold increase in PSA mRNA was observed following T stimulation. Low levels of immunoreactive PSA (iPSA) were detected in media of androgen-stimulated TE85 cells while iPSA was not found in control media. Conversely, iPSA was detected in TE85 cell pellets from control but not in androgen-stimulated cell cultures. Exogenously added enzymatically active PSA stimulated TE85 proliferation in a bi-phasic manner. LY312340 inhibited PSA-induced increases in TE85 cell numbers but had no effect on basal or T- stimulated cellular proliferation. CONCLUSION: While the PSA levels produced by TE-85 cells in response to androgen stimulation are too low to be biologically active, PSA produced by metastatic PCa cells may mediate paracrine stimulation of osteogenic PCa metastasis. Inhibitors of PSA enzymatic activity could be useful therapeutic agents.
Assuntos
Benzoatos/farmacologia , Neoplasias Ósseas/patologia , Lactamas/farmacologia , Neoplasias Hormônio-Dependentes/patologia , Osteossarcoma/patologia , Antígeno Prostático Específico/antagonistas & inibidores , Adolescente , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Divisão Celular/efeitos dos fármacos , Divisão Celular/fisiologia , Feminino , Humanos , Neoplasias Hormônio-Dependentes/genética , Neoplasias Hormônio-Dependentes/metabolismo , Osteossarcoma/genética , Osteossarcoma/metabolismo , Antígeno Prostático Específico/biossíntese , Antígeno Prostático Específico/genética , Antígeno Prostático Específico/fisiologia , Testosterona/deficiência , Testosterona/farmacologia , Testosterona/fisiologia , Células Tumorais CultivadasRESUMO
BACKGROUND: Androgen-deprivation remains the standard of care for metastatic prostate cancer, yet its total impact on the course of disease is incompletely established. METHODS: We have examined the long-term effects of castration upon the progression of established cancer in the TRAMP transgenic mouse model of prostate cancer. Mice castrated at 15-weeks of age, as well as intact littermates, were followed until spontaneous death from cancer. RESULTS: Statistical analyses of age-at-death versus primary tumor mass revealed that mice segregate into two categories of response to androgen-deprivation. In Category One, the act of castration paradoxically triggers the growth of microscopic androgen-independent lesions, as evidenced by a statistical synchronization of primary tumor growth. Delaying castration until 20-weeks of age delays the synchronized growth of these tumors, as well as the deaths of the host mice. In Category Two, castration eliminates or substantially delays primary tumor growth, but fails to eliminate metastases. so that castration is found to impart no long-term survival advantage. CONCLUSIONS: We propose a two-step model for the alteration of androgen signaling in prostate cancer capable of explaining the above paradoxical results, which is based upon the dualistic role of androgens as both survival and differentiation factors. This model makes specific predictions for clinical intervention that are discussed in light of published studies.
Assuntos
Androgênios/farmacologia , Castração/veterinária , Neoplasias da Próstata/patologia , Animais , Modelos Animais de Doenças , Progressão da Doença , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias da Próstata/veterinária , Transdução de Sinais , Análise de SobrevidaRESUMO
OBJECTIVES: To study the safety, pharmacokinetics, biologic activity, and preliminary efficacy of the bispecific 5-alpha-reductase inhibitor (LY320236) in prostate cancer. METHODS: Fifty-one patients with recurrent or metastatic prostate cancer were sequentially (nonrandomly) assigned in cohorts to receive one of five single daily oral doses of LY320236 (10, 50, 150, 250, and 500 mg). Serial evaluations included serum testosterone, dihydrotestosterone, androstenediol glucuronide, estradiol, and pharmacokinetics on days 1, 29, and 57. Toxicity assessments, x-rays/scans, and blood tests, including serum prostate-specific antigen (PSA) determination, were done at regular intervals. RESULTS: Overall, treatment was well tolerated, with 3 of 51 patients developing reversible grade 3-4 toxicity (one diarrhea, two elevated liver enzymes). Peak blood levels (2 to 3 hours after drug administration) were greater for doses of 150 mg or greater compared with less than 150-mg doses with slow accumulation. Serum levels of testosterone, dihydrotestosterone, and androstenediol glucuronide did not change significantly during treatment; however, a statistically significant increase occurred in serum estradiol levels in both the castration and noncastration groups. One of 26 in the noncastration group and 4 (27%) of 15 in the castration group with baseline PSA levels of 5 ng/mL or greater had a 50% or greater PSA decline for 4 weeks or longer. CONCLUSIONS: LY320236 treatment is associated with modest reversible toxicity. An elevation of estradiol levels was seen in both castration and noncastration groups, although PSA declines were primarily seen in the castration group. The absence of cardiovascular toxicity suggests that this agent may be a promising alternative to exogenous estrogens in patients with prostate cancer who demonstrate evidence of disease progression after initial androgen deprivation treatment.
Assuntos
Inibidores de 5-alfa Redutase , Adenocarcinoma/tratamento farmacológico , Antagonistas de Androgênios/uso terapêutico , Benzoquinonas/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Estradiol/sangue , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias da Próstata/tratamento farmacológico , Adenocarcinoma/sangue , Adenocarcinoma/enzimologia , Adenocarcinoma/cirurgia , Antagonistas de Androgênios/administração & dosagem , Antagonistas de Androgênios/efeitos adversos , Antagonistas de Androgênios/farmacologia , Androstenodióis/sangue , Benzoquinonas/administração & dosagem , Benzoquinonas/efeitos adversos , Benzoquinonas/farmacologia , Biomarcadores Tumorais/sangue , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Di-Hidrotestosterona/sangue , Intervalo Livre de Doença , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/efeitos adversos , Inibidores Enzimáticos/farmacologia , Humanos , Isoenzimas/antagonistas & inibidores , Tábuas de Vida , Masculino , Proteínas de Neoplasias/sangue , Orquiectomia , Antígeno Prostático Específico/sangue , Neoplasias da Próstata/sangue , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/cirurgia , Segurança , Especificidade por Substrato , Testosterona/sangue , Resultado do TratamentoRESUMO
Phospholipase A2 (PLA2) enzymes release arachidonic acid from cellular phospholipids in a variety of mammalian tissues, including prostate. Group IIa secretory PLA2 (sPLA2) can generate arachidonate from cellular phospholipids. We examined the group IIa sPLA2 expression in benign prostatic tissues, prostatic intraepithelial neoplasia (PIN), and adenocarcinoma to determine whether sPLA2 expression is altered in the carcinogenesis of human prostatic cancer. Thirty-three of 74 total cases (45%) of benign prostatic tissue showed positive immunohistochemical staining for group IIA sPLA2, whereas 63 of 69 total cases (91%) of high-grade PINs and 70 of 78 total cases (90%) of adenocarcinomas gave positive results. Four of 10 cases of low-grade PIN showed positive immunoreactivity for sPLA2. The number of cells staining for sPLA2 was significantly less in benign epithelium (4%) and low-grade PIN (4%) compared to high-grade PIN (40%) or adenocarcinoma (38%) (P < 0.001). There was no significant difference between high-grade PIN and adenocarcinoma in the number of cells staining positively for sPLA2. The intensity of sPLA2 immunoreactivity was also different among benign prostatic tissue, low-grade PIN, high-grade PIN, and prostatic adenocarcinoma specimens. The malignant cells demonstrated more intense immunohistochemical staining (moderate to strong staining in 81% and 69% cases for high-grade PIN and adenocarcinoma, respectively) than benign glands (moderate staining in 11% of cases). No strong staining was observed in benign glands or low-grade PIN. Our data are consistent with the contention that group IIA sPLA2 expression is elevated in neoplastic prostatic tissue and support the hypothesis that dysregulation of sPLA2 may play a role in prostatic carcinogenesis.