RESUMEN
Trastuzumab is by far the drug of choice for treatment of human epidermal growth factor receptor 2 (Her2) overexpressing breast cancer patients. However, frequently, the therapy remains ineffective due to the induced drug resistance. In spite of various reported mechanisms, we hypothesize that the acquired resistance to trastuzumab might be attributed to the failure of the drug to activate phosphatase and tensin homolog (PTEN) mainly due to the high level of reduced thioredoxin-1 protein among the resistant cells. In the present study, the effect(s) of PX-12, a Trx-1 inhibitor, was examined on proliferation of breast cancer cells which are unresponsive to trastuzumab. Treatment of the cells with PX-12 (5 µM) and trastuzumab (10 µg/ml) reduced cells viabilities, p-Akt, and Bcl2 levels while increasing the levels of reactive oxygen species (ROS) and p-JNK with consequent higher levels of G1 arrest and apoptosis among the resistant cells compared to parental trastuzumab sensitive cells. The most significant observation was that PX-12/trastuzumab co-treatment enhanced the cell membrane localization of PTEN which is believed to be the active biological form of the signal. Our data confirmed that Trx-1 inhibition is required for chemosensitization of resistant breast cancer cells to anti-Her2 therapy, and this approach might offer an alternative clinical strategy for preventing acquired resistance.
Asunto(s)
Antineoplásicos/farmacología , Resistencia a Antineoplásicos , Fosfohidrolasa PTEN/metabolismo , Tiorredoxinas/metabolismo , Trastuzumab/farmacología , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Membrana Celular/metabolismo , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Disulfuros/farmacología , Resistencia a Antineoplásicos/genética , Humanos , Imidazoles/farmacología , Modelos Biológicos , Unión Proteica/efectos de los fármacos , Transporte de Proteínas , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Receptor ErbB-2/antagonistas & inhibidores , Tiorredoxinas/genéticaRESUMEN
Several mammalian enzymes are anchored to the outer surface of the plasma membrane by a covalently attached glycosylphosphatidylinositol (GPI) structure. These include acetylcholinesterase, alkaline phosphatase (AP) and 5'-nucleotidase among other enzymes. Recently, it has been reported that these membrane enzymes can be released into the serum by the GPI-dependent phospholipase D under various medical disturbances such as cancer and/or by chemical and physical manipulation of the biological systems. Treatment of MCF-7 cells with two consecutive effective concentrations of 3-hydrogenkwadaphnin (3-HK, 3 nM) for 48 h enhanced membrane AP activity by almost 330% along with a 40% reduction in the AP activity of the cell culture medium. In addition, our data indicate that 3-HK is capable of inducing mainly the tissue-nonspecific alkaline phosphatase (TNAP) isoenzyme, along with enhancing its thermostability. These findings, besides establishing a correlation between the antiproliferative activity of 3-HK and the extent of plasma membrane AP activity, might assist in the development of new diagnostic tools for following cancer medical treatments.
Asunto(s)
Fosfatasa Alcalina/metabolismo , Antineoplásicos Fitogénicos/farmacología , Diterpenos/farmacología , Fosfatasa Alcalina/antagonistas & inhibidores , Antineoplásicos Fitogénicos/aislamiento & purificación , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/patología , Línea Celular Tumoral , Membrana Celular/efectos de los fármacos , Membrana Celular/enzimología , Supervivencia Celular/efectos de los fármacos , Diterpenos/aislamiento & purificación , Inhibidores Enzimáticos/farmacología , Femenino , Glicosilfosfatidilinositoles/metabolismo , Humanos , Microdominios de Membrana/efectos de los fármacos , Microdominios de Membrana/enzimología , Fitoterapia , Thymelaeaceae/químicaRESUMEN
Regarding the world wide success of artificial chaperone-assisted protein refolding technique and based on its well worked-out mechanism, it is anticipated that the lipid moieties of glycosylphosphatidylinositol (GPI) group, which is present in some membrane proteins, might interfere with the capturing step of the technique. To find an answer, we evaluated the chemical denaturation and also the refolding behavior of insoluble and soluble alkaline phosohatase (ALP), with or without GPI group, respectively. The results indicated that the presence of GPI in the enzyme increased the stability of the protein against chemical denaturation while it decreased its refolding yield by the artificial chaperone refolding technique. The lower refolding yield, compared to soluble ALP (sALP), might be due to a less efficient stripping step caused by new interactions imparted to the refolding elements of the system especially those among the hydrophobic tails of GPI and the capturing agent of the technique. These new interactions will interrupt the kinetics of detergent stripping from the captured molecules by the stripping agent (i.e., cyclodextrins). This situation will lead to higher intermolecular hydrophobic interactions among the refolding protein intermediates leading to their higher misfolding and aggregation.