RESUMO
Novel, well-tolerated drugs are needed for the management of canine mammary cancer. Many of these cancers are promoted in their growth by estrogen. Alpha-fetoprotein (AFP) is a ubiquitous mammalian protein that has anti-estrogenic properties. AFPep (the anti-estrogenic site of AFP) has been developed into a readily synthesizable drug. AFPep has been shown to have anti-mammary cancer activity in several models of this disease, both in cell culture and in rodents. The purpose of the study reported herein was to determine the tolerability of AFPep in normal and tumor-bearing dogs. AFPep was given to dogs via both parenteral and oral routes in a single application and in repeated daily doses. Full clinical chemistry and hematology values were determined before and after drug administration. Blood levels of the drug were achieved in dogs that had been previously found to be oncostatic in rodents. No changes in clinical chemistry, hematology, and clinical behaviors were found in dogs following drug administration. The data support the further development of AFPep for clinical use against canine mammary cancer.
RESUMO
PURPOSE: The purpose of this study is to assess the efficacy and safety profile of AFPep, a 9-amino acid cyclic peptide prior to its entry into pre-clinical toxicology analyses en route to clinical trials. METHODS: AFPep was assessed for anti-estrogenic activity in a mouse uterine growth assay and for breast cancer therapeutic efficacy in a human tumor xenograft model in mice. AFPep was assessed for tolerability in a variety of in vivo models, notably including assessment for effects on rat liver and human hepatocellular carcinoma cell lines and xenografts. RESULTS: AFPep arrests the growth of human MCF-7 breast cancer xenografts, inhibits the estrogen-induced growth of mouse uteri, and does not affect liver growth nor stimulate growth of human hepatocellular carcinoma cell lines when growing in vitro or as xenografts in vivo. AFPep is well tolerated in mice, rats, dogs, and primates. CONCLUSIONS: AFPep is effective for the treatment of ER-positive breast cancer and exhibits a therapeutic index that is substantially wider than that for drugs currently in clinical use. The data emphasize the importance of pursuing pre-clinical toxicology studies with the intent to enter clinical trials.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Antagonistas de Estrogênios/uso terapêutico , Fragmentos de Peptídeos/uso terapêutico , Peptídeos Cíclicos/uso terapêutico , alfa-Fetoproteínas/uso terapêutico , Animais , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Cães , Antagonistas de Estrogênios/farmacologia , Feminino , Células Hep G2 , Humanos , Células MCF-7 , Macaca mulatta , Camundongos , Camundongos SCID , Fragmentos de Peptídeos/farmacologia , Peptídeos Cíclicos/farmacologia , Ratos , Ratos Sprague-Dawley , Resultado do Tratamento , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/fisiologia , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , alfa-Fetoproteínas/farmacologiaRESUMO
Epidemiologic studies associate elevated maternal serum levels of α-fetoprotein (AFP) with reduced breast cancer risk for parous women. Laboratory studies demonstrate direct anti-breast cancer activity of AFP. Here, we review the development of a small cyclic peptide that is an active site analog of AFP, referred to as AFPep, which is composed exclusively of amino acids, is orally active, has no discernable toxicity, and is effective for the treatment and prevention of breast cancer in animal models.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/uso terapêutico , Neoplasias da Mama/prevenção & controle , Domínio Catalítico , Descoberta de Drogas , alfa-Fetoproteínas/química , Animais , Antineoplásicos/química , Neoplasias da Mama/epidemiologia , Domínio Catalítico/efeitos dos fármacos , Quimioprevenção/métodos , Feminino , Humanos , Paridade/fisiologia , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/uso terapêutico , GravidezRESUMO
In earlier work, we synthesized a cyclic 9-amino acid peptide (AFPep, cyclo[EKTOVNOGN]) and showed it to be useful for prevention and therapy of breast cancer. In an effort to explore the structure-function relationships of AFPep, we have designed analogs that bear a short 'tail' (one or two amino acids) attached to the cyclic peptide distal to its pharmacophore. Analogs that bore a tail of either one or two amino acids, either of which had a hydrophilic moiety in the side chain (e.g., cyclo[EKTOVNOGN]FS) exhibited greatly diminished biological activity (inhibition of estrogen-stimulated uterine growth) relative to AFPep. Analogs that bore a tail of either one or two amino acids which had hydrophobic (aliphatic or aromatic) side chains (e.g., cyclo[EKTOVNOGN]FI) retained (or had enhanced) growth inhibition activity. Combining in the same biological assay a hydrophilic-tailed analog with either AFPep or a hydrophobic-tailed analog resulted in decreased activity relative to that for AFPep or for the hydrophobic-tailed analog alone, suggesting that hydrophilic-tailed analogs are binding to a biologically active receptor. An analog with a disrupted pharmacophore (cyclo[EKTOVGOGN]) exhibited little or no growth inhibition activity. An analog with a hydrophilic tail and a disrupted pharmacophore (cyclo[EKTOVGOGN]FS) exhibited no growth inhibition activity of its own and did not affect the activity of a hydrophobic-tailed analog, but enhanced the growth inhibition activity of AFPep. These results are discussed in the context of a two-receptor model for binding of AFPep and ring-and-tail analogs. We suggest that tails on cyclic peptides may comprise a useful method to enhance diversity of peptide design and specificity of ligand-receptor interactions.
Assuntos
Aminoácidos/química , Antineoplásicos/síntese química , Peptídeos Cíclicos/química , Sequência de Aminoácidos , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Sítios de Ligação , Estradiol/administração & dosagem , Estradiol/farmacologia , Feminino , Interações Hidrofóbicas e Hidrofílicas , Camundongos , Modelos Moleculares , Peptídeos Cíclicos/síntese química , Peptídeos Cíclicos/farmacologia , Conformação Proteica , Receptores de Superfície Celular/química , Técnicas de Síntese em Fase Sólida , Relação Estrutura-Atividade , Especificidade por Substrato , Útero/efeitos dos fármacos , Útero/crescimento & desenvolvimento , Ensaios Antitumorais Modelo de Xenoenxerto , alfa-Fetoproteínas/químicaRESUMO
Parity in women is associated with reduced lifetime risk of breast cancer, and hormones of pregnancy [estrogen (E), progesterone (P), human chorionic gonadotropin (hCG)] are implicated. Parity also reduces mammary cancer risk in carcinogen-exposed rats, and administering pregnancy hormones to these animals is similarly effective. Because pregnancy hormones are also able to stimulate cancer growth, we proposed to resolve this dichotomy by determining whether administered pregnancy hormones elicit the cancer-inhibiting agent alpha-fetoprotein (AFP) from the liver, which would implicate AFP as a proximal effector of hormonal anticancer activity. Accordingly, we treated groups of nitrosomethylurea-exposed rats with saline, E(3), E(2) + P, E(3) + P, hCG, or allowed them to experience pregnancy, and then monitored mammary cancer incidence and serum levels of AFP over time. Each hormone treatment reduced mammary cancer incidence and elevated serum AFP levels. To challenge human tissues, human HepG2 liver cells in culture were treated with the same hormonal agents. Each hormone regimen increased the levels of AFP in the culture medium. Medium containing AFP elicited by hCG inhibited the E(2)-stimulated proliferation of cultured human MCF7 breast cancer cells, whereas hCG alone did not inhibit their growth. Furthermore, antibodies to AFP neutralized the growth-inhibiting effect of AFP-containing HepG2 medium. We conclude that in the treatment of carcinogen-exposed rats with the hormones of pregnancy, and by inference in women who have experienced pregnancy, that AFP is a proximal agent that inhibits mammary gland cancer.
Assuntos
Neoplasias da Mama/etiologia , Gonadotropina Coriônica/metabolismo , Estradiol/metabolismo , Gravidez/fisiologia , Progesterona/metabolismo , alfa-Fetoproteínas/metabolismo , Animais , Neoplasias da Mama/epidemiologia , Gonadotropina Coriônica/farmacologia , Estradiol/farmacologia , Feminino , Humanos , Neoplasias Mamárias Experimentais/epidemiologia , Neoplasias Mamárias Experimentais/etiologia , Paridade/fisiologia , Progesterona/farmacologia , Ratos , Ratos Sprague-Dawley , Fatores de RiscoRESUMO
Pregnancy lowers the risk of breast cancer, largely attributable to alpha-fetoprotein (AFP). A small AFP-derived peptide (AFPep) which mimics the active site of AFP has been developed and may be useful for decreasing the risk of breast cancer for women. AFPep has been shown previously to stop the growth of estrogen-dependent human breast cancer xenografts in mice and prevent carcinogen-induced breast cancer in a rat model. Since AFPep disrupts an estrogen-responsive pathway, it is essential to assess its effects on the female reproductive cycle and fertility. Ten cycling female Sprague-Dawley rats (age 81 days) were given 100 microg AFPep in saline s.c. daily for 20 days. A second group of ten rats was given 50 microg tamoxifen s.c. daily and a third group received saline only. Vaginal smears were obtained twice per day and stained to assess estrous cycle phase. After completion of estrous cycle assessment (five cycles, 21 days), rats were maintained on drug and allowed to mate. Effects on birth of offspring and maternal body weights were assessed. AFPep had no significant effect on the incidence or duration of any estrous cycle phase, and no effect on reproductive potential or maternal body mass. Tamoxifen significantly increased the length of diestrus, locking the cycle in this phase for most animals. Only half of the tamoxifen-treated rats mated, and none became pregnant. Tamoxifen significantly slowed the rate of body mass increase. In rats, AFPep has no toxicity and no effect on female reproduction. This molecule may be developed into an attractive modality for prevention of breast cancer in women.
Assuntos
Anticarcinógenos/toxicidade , Antineoplásicos Hormonais/toxicidade , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/prevenção & controle , Moduladores de Receptor Estrogênico/toxicidade , Estro/efeitos dos fármacos , Fertilidade/efeitos dos fármacos , Fragmentos de Peptídeos/toxicidade , Tamoxifeno/toxicidade , alfa-Fetoproteínas/toxicidade , Animais , Animais Recém-Nascidos , Anticarcinógenos/administração & dosagem , Antineoplásicos Hormonais/administração & dosagem , Peso ao Nascer/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Moduladores de Receptor Estrogênico/administração & dosagem , Feminino , Injeções Subcutâneas , Tamanho da Ninhada de Vivíparos/efeitos dos fármacos , Fragmentos de Peptídeos/administração & dosagem , Gravidez , Ratos , Ratos Sprague-Dawley , Tamoxifeno/administração & dosagem , Fatores de Tempo , alfa-Fetoproteínas/administração & dosagemRESUMO
Cyclo[EKTOVNOGN] (AFPep), a cyclic 9-amino acid peptide derived from the active site of alpha-fetoprotein, has been shown to prevent carcinogen-induced mammary cancer in rats and inhibit the growth of ER(+) human breast cancer xenografts in mice. Recently, studies using replica exchange molecular dynamics predicted that the TOVN region of AFPep might form a dynamically stable putative Type I beta-turn, and thus be biologically active without additional amino acids. The studies presented in this paper were performed to determine whether TOVN and other small analogs of AFPep would inhibit estrogen-stimulated cancer growth and exhibit a broad effective-dose range. These peptides contained nine or fewer amino acids, and were designed to bracket or include the putative pharmacophoric region (TOVN) of AFPep. Biological activities of these peptides were evaluated using an immature mouse uterine growth inhibition assay, a T47D breast cancer cell proliferation assay, and an MCF-7 breast cancer xenograft assay. TOVN had very weak antiestrogenic activity in comparison to AFPep's activity, whereas TOVNO had antiestrogenic and anticancer activities similar to AFPep. OVNO, which does not form a putative Type I beta-turn, had virtually no antiestrogenic and anticancer activities. A putative proteolytic cleavage product of AFPep, TOVNOGNEK, significantly inhibited E(2)-stimulated growth in vivo and in vitro over a wider dose range than AFPep or TOVNO. We conclude that TOVNO has anticancer potential, that TOVNOGNEK is as effective as AFPep in suppressing growth of human breast cancer cells, and that it does so over a broader effective-dose range.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Moduladores de Receptor Estrogênico/síntese química , Moduladores de Receptor Estrogênico/farmacologia , alfa-Fetoproteínas/síntese química , alfa-Fetoproteínas/farmacologia , Sequência de Aminoácidos , Animais , Antineoplásicos/química , Sítios de Ligação , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Moduladores de Receptor Estrogênico/química , Feminino , Humanos , Camundongos , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/farmacologia , Transplante Heterólogo , Útero/efeitos dos fármacos , Útero/crescimento & desenvolvimento , alfa-Fetoproteínas/químicaRESUMO
Parity profoundly reduces breast cancer (BC) risk later in life. It has been reasoned that hormones (either estradiol E2 or estriol E3), progesterone (P) or human chorionic gonadotropin (hCG) in the serum of pregnant women might lead to that reduction in risk. These agents have been shown to reduce BC incidence in nonpregnant rats. We investigated the hypothesis that exogenously added E2, E3, P, or hCG are not the proximal effectors of risk reduction, but that they elicit alpha-fetoprotein (alphaFP) from the nonpregnant liver, and that cFP is the proximal agent by which reduction of BC risk is obtained. Methylnitrosourea (MNU)-exposed animals were treated with saline, E3, E2 + P, E3 + P, hCG, or were allowed to experience pregnancy, and AFP levels were measured in the serum and subsequent tumor incidence was recorded. Human HepG2 liver cells in culture were treated with E3, E2 + P, P, or hCG and elicited AFP was measured in the media. The HepG2 culture media containing elicited AFP was assessed for its ability to inhibit proliferation of T47D cells when applied to these human BC cells in culture, and to inhibit the estrogen-induced phosphorylation of the estrogen receptor in T47D cells. For each condition in the prevention studies, hormone treatment reduced the incidence of BC to an extent similar to that reported by the original studies. In each condition, alphaFP levels in serum were elevated over that in control animals. In culture, treatment of human liver cells with E3, E2 + P, or hCG, but not P alone, led to increased levels of AFP in the media. Media containing hCG-elicited AFP inhibited the estrogen-stimulated proliferation of T47D cells in culture, and inhibited phosphorylation of the estrogen receptor, whereas, estrogens and hCG did not inhibit the growth of these tumor cells in culture. In conclusion, since the hormones of pregnancy elicit alphaFP from the liver, and alphaFP but not the hormones of pregnancy has direct antitumor properties, it is concluded that alphaFP is the proximal agent through which reduction in BC incidence is realized from the experience of pregnancy.
Assuntos
Neoplasias da Mama/prevenção & controle , Proliferação de Células/efeitos dos fármacos , Estrogênios/uso terapêutico , alfa-Fetoproteínas/metabolismo , Animais , Neoplasias da Mama/induzido quimicamente , Neoplasias da Mama/metabolismo , Carcinógenos , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Gonadotropina Coriônica/uso terapêutico , Estradiol/uso terapêutico , Estriol/uso terapêutico , Feminino , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Metilnitrosoureia , Gravidez , Progesterona/uso terapêutico , Ratos , Substâncias para o Controle da Reprodução/uso terapêutico , Fatores de Risco , Células Tumorais CultivadasRESUMO
This study was aimed to obtain additional information on the activity of a cyclized 9-amino acid peptide (cP) containing the active site of alpha fetoprotein, which inhibits the estrogen-stimulated proliferation of tumor cells in culture and of xenografts in immunodeficient mice. Breast cancer cells cultured in the presence of 2 nM estradiol were exposed to cP for different periods and their proliferation, estradiol binding parameters, clustering tendency and expression of E-cadherin and p21Cip1 were analyzed by biochemical and cell biology methods. The proliferation of MCF7 cells was significantly decreased by the addition of 2 microg/ml cP to the medium. cP did not increase cell death rate nor alter the number of binding sites for estradiol nor the endogenous aromatase activity of MCF7 cells. cP also decreased the proliferation of estrogen-dependent ZR75-1 cells but had no effect on estrogen-independent MDA-MB-231 cells. An increased nuclear p21Cip1 expression detected after cP treatment suggests that cP slows MCF7 cell proliferation via this regulator. We propose that cP could represent a novel breast cancer therapeutic agent whose mechanism of action is different from that of tamoxifen or of inhibitors of aromatase.
Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Estradiol/farmacologia , Glândulas Mamárias Humanas/efeitos dos fármacos , alfa-Fetoproteínas/farmacologia , Animais , Antineoplásicos/química , Western Blotting , Neoplasias da Mama/tratamento farmacológico , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/biossíntese , Inibidor de Quinase Dependente de Ciclina p21/efeitos dos fármacos , Feminino , Imunofluorescência , Humanos , Camundongos , Camundongos Nus , Peptídeos/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto , alfa-Fetoproteínas/químicaRESUMO
Breast cancer is the most common cancer among women, and tamoxifen is the preferred drug for estrogen receptor-positive breast cancer treatment. Many of these cancers are intrinsically resistant to tamoxifen or acquire resistance during treatment. Consequently, there is an ongoing need for breast cancer drugs that have different molecular targets. Previous work has shown that 8-mer and cyclic 9-mer peptides inhibit breast cancer in mouse and rat models, interacting with an unsolved receptor, while peptides smaller than eight amino acids did not. We show that the use of replica exchange molecular dynamics predicts the structure and dynamics of active peptides, leading to the discovery of smaller peptides with full biological activity. Simulations identified smaller peptide analogues with the same conserved reverse turn demonstrated in the larger peptides. These analogues were synthesized and shown to inhibit estrogen-dependent cell growth in a mouse uterine growth assay, a test showing reliable correlation with human breast cancer inhibition.
Assuntos
Desenho de Fármacos , Moduladores de Receptor Estrogênico , Modelos Moleculares , Oligopeptídeos , alfa-Fetoproteínas/química , Sequência de Aminoácidos , Animais , Moduladores de Receptor Estrogênico/síntese química , Moduladores de Receptor Estrogênico/química , Moduladores de Receptor Estrogênico/farmacologia , Feminino , Camundongos , Dados de Sequência Molecular , Oligopeptídeos/síntese química , Oligopeptídeos/química , Oligopeptídeos/farmacologia , Tamanho do Órgão/efeitos dos fármacos , Útero/efeitos dos fármacosRESUMO
BACKGROUND: We have synthesized a cyclic nonapeptide (AFPep) that is effective, after being administered by parenteral routes, for the treatment or the prevention of breast cancer. To test the hypothesis that AFPep remains safe and efficacious after oral administration, three different whole-animal bioassays were utilized, and the mechanism by which AFPep functions was investigated. METHODS: Using a human breast cancer xenograft model in mice for therapeutic activity, a carcinogen-induced breast cancer model in rats for prevention efficacy, and a mouse uterus growth inhibition model of anti-estrogenic activity, AFPep was administered by oral gavage (p.o.) and its effects compared to those following intraperitoneal (i.p.) and subcutaneous (s.c.) administration. Toxicity studies evaluated body weights and organ weights in mice and rats receiving AFPep. Preliminary mechanistic studies were carried out in T47D human breast cancer cells growing in culture and evaluated the effect of AFPep on estrogen-stimulated cell growth, phosphorylation of the estrogen receptor (ER), and on level of ER-related kinases. RESULTS: Orally administered AFPep stopped the growth of human tumor xenografts in mice, decreased the incidence and multiplicity of breast cancers in carcinogen-exposed rats, and inhibited the estrogen-stimulated growth of mouse uteri. In each of these systems, orally administered AFPep produced an effect similar to that obtained for AFPep administered by either i.p or s.c. routes. In rodents, no evidence of toxicity was seen for the peptide, even at very high doses. In culture, AFPep inhibited the estrogen-stimulated growth, but not the basal growth, of T47D cells, and it inhibited the estrogen-stimulated phosphorylation of Serine 118 in the ER of these cells, which was not explainable by early changes in ER-related kinases. CONCLUSIONS: Chronic oral administration of AFPep appears to be safe and effective for the treatment or prevention of breast cancer in animal models.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Peptídeos Cíclicos/uso terapêutico , alfa-Fetoproteínas , Administração Oral , Animais , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Humanos , Masculino , Camundongos , Camundongos SCID , Transplante de Neoplasias , Transplante HeterólogoRESUMO
PURPOSE: alpha-Fetoprotein (AFP) is a protein of pregnancy associated with a decrease in lifetime risk of breast cancer in parous women. A synthetic, cyclic nonapeptide has been developed that mimics the antioncogenic active site of AFP. To test the hypothesis that the AFP-derived peptide (AFPep) can prevent breast cancer, the N-methyl-N-nitrosourea-induced breast cancer model was used in rats. EXPERIMENTAL DESIGN: AFPep was given daily by injection beginning 10 days after N-methyl-N-nitrosourea treatment and continued for 23 days (a time designed to mimic pregnancy) or for other times to assess efficacy as a function of drug duration. Tumor incidence, multiplicity, and latency were noted as end points. At necropsy, pathology analysis of tumors and major organs were obtained. RESULTS: AFPep prevented cancer in a dose-dependent fashion. Significantly longer mean tumor-free days (P < 0.02), lower tumor incidence (P = 0.004), and lower tumor multiplicity were observed for AFPep-treated groups. No evidence of host toxicity as measured by body weight, cage activity, fur texture, and organ weights (liver, uterus, heart, kidney, and spleen) were found in animals treated with AFPep. Mechanistic studies using transplantable human breast cancer xenografts showed that the peptide interfered with estrogen-dependent breast cancer growth inhibited the phosphorylation of the estrogen receptor and activated phosphorylation of p53. CONCLUSIONS: AFPep is a well-tolerated, mechanistically novel, chemopreventive agent in models of breast cancer and warrants further development for the prevention and treatment of this disease in humans.
Assuntos
Alquilantes/toxicidade , Neoplasias Mamárias Experimentais/induzido quimicamente , Neoplasias Mamárias Experimentais/prevenção & controle , Metilnitrosoureia/toxicidade , Fragmentos de Peptídeos/uso terapêutico , alfa-Fetoproteínas/uso terapêutico , Animais , Sítios de Ligação , Feminino , Humanos , Incidência , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos SCID , Fosforilação , Ratos , Ratos Sprague-Dawley , Receptores de Estrogênio/metabolismo , Receptores de Progesterona/metabolismo , Distribuição Tecidual , Transplante Heterólogo , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/metabolismo , alfa-Fetoproteínas/farmacocinéticaRESUMO
DSCR1 (adapt78) is a stress-inducible gene and cytoprotectant. Its protein product, DSCR1 (Adapt78), also referred to as MCIP1, inhibits intracellular calcineurin, a phosphatase that mediates many cellular responses to calcium. Exposure of human U251 and HeLa cells to hydrogen peroxide led to a rapid hyperphosphorylation of DSCR1 (Adapt78). Inhibitor and agonist studies revealed that a broad range of kinases were not responsible for DSCR1 (Adapt78) hyperphosphorylation, including ERK1/2, although parallel activation of the latter was observed. Phosphorylation of both DSCR1 (Adapt78) and ERK1/2 was attenuated by inhibitors of tyrosine phosphatase, suggesting the common upstream involvement of tyrosine dephosphorylation. The hyperphosphorylation electrophoretic shift in DSCR1 (Adapt78) mobility was also observed with other oxidizing agents (peroxynitrite and menadione) but not nonoxidants. Calcium ionophores strongly induced the levels of both hypo- and hyper-phosphorylated DSCR1 (Adapt78) but did not alter phosphorylation status. Calcium-dependent growth factor- and angiotensin II-stimulation also induced both DSCR1 (Adapt78) species. Phosphorylation of either or both serines in a 13-amino acid peptide made to a calcineurin-interacting conserved region of DSCR1 (Adapt78) attenuated inhibition of calcineurin. These data indicate that DSCR1 (Adapt78) protein is a novel, early stage oxidative stress-activated phosphorylation target and newly identified calcium-inducible protein, and suggest that these response mechanisms may contribute to the known cytoprotective and calcineurin-inhibitory activities of DSCR1 (Adapt78).
Assuntos
Inibidores de Calcineurina , Cálcio/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas Musculares/metabolismo , Estresse Oxidativo , Angiotensina II/metabolismo , Antifibrinolíticos/farmacologia , Astrocitoma/patologia , Calcineurina/metabolismo , Divisão Celular/efeitos dos fármacos , Proteínas de Ligação a DNA , Ensaio de Desvio de Mobilidade Eletroforética , Inibidores Enzimáticos/farmacologia , Substâncias de Crescimento/metabolismo , Células HeLa , Humanos , Peróxido de Hidrogênio/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular , Ionóforos/farmacologia , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Oxidantes/farmacologia , Ácido Peroxinitroso/farmacologia , Fosforilação/efeitos dos fármacos , Proteínas Tirosina Fosfatases/antagonistas & inibidores , Vitamina K 3/farmacologiaRESUMO
An 8-mer peptide (EMTOVNOG) derived from alpha-fetoprotein was compared with tamoxifen for activity against growth of human breast cancer xenografts implanted in immune-deficient mice. Both peptide and tamoxifen prevented growth of estrogen-receptor-positive MCF-7 and T47D human breast cancer xenografts. A subline of MCF-7, made resistant to tamoxifen by a 6-month exposure to this drug in culture, was found to be resistant to tamoxifen in vivo. Peptide completely prevented the xenograft growth of this tamoxifen-resistant subline of MCF-7. Neither peptide nor tamoxifen was effective in slowing the xenograft growth of the estrogen-receptor-negative MDA-MB-231 human breast cancer. A worrisome side effect of tamoxifen is its hypertrophic effect on the uterus. In this study, tamoxifen was shown to stimulate the growth of the immature mouse uterus in vivo, and the peptide significantly inhibited tamoxifen's uterotrophic effect. The mechanism of action of peptide is different from that of tamoxifen in that the peptide does not interfere with the binding of [(3)H]estradiol to the estrogen receptor. In conclusion, alpha-fetoprotein-derived peptide appears to be a novel agent that interferes with the growth of tamoxifen-sensitive as well as tamoxifen-resistant estrogen-receptor-positive human breast cancers; it inhibits the uterotrophic side effect of tamoxifen and, thus, it may be useful in combination with or in place of tamoxifen for treatment of estrogen-receptor-positive human breast cancers.