Effect of liposome-encapsulated alpha- or beta-interferon on breast cancer cell lines.

Experimental evidence and clinical studies have indicated that interferons (IFN) inhibit proliferation in a wide panel of neoplasms, including breast cancer. However, the antitumor activity of IFN requires the continuous presence of high concentrations of the drug and is associated with side effects. To explore the potential of liposomes as an IFN delivery system, we compared the effect of free or liposome-encapsulated alpha-IFN and beta-IFN on the growth of two breast cancer cell lines (MCF7 and MDA-MB231). Cells were cultured in the presence of IFN at different concentrations (500, 1000, 2000 IU/ml) or in the presence of multilamellar liposomes (phosphatidylcholine-phosphatidylserine at a molar ratio of 7:3) containing saline buffer, alpha-IFN or beta-IFN. Additional control groups consisted of cells cultured with alpha-IFN or beta-IFN plus empty liposomes. Empty liposomes were not cytotoxic and did not interfere with IFN activity. In both cell lines liposomes encapsulating alpha-IFN (at the highest lipid:drug ratio) inhibited cell growth in a manner similar to that of free alpha-IFN, whereas liposomes encapsulating beta-IFN showed slightly, lower inhibition than free beta-IFN, this was more evident in MCF7 cells. The present results indicate that liposomes encapsulating alpha-IFN or beta-IFN were effective on the growth of both breast cancer cell lines, which are characterized by a different estrogen responsiveness, and that they might be a useful carrier system for the delivery of high doses of IFN.

Simultaneous but not sequential treatment with sodium butyrate improves the antiproliferative effect of alpha- or beta-interferon on a breast cancer cell line.

Clinical evidence indicates that alpha- and beta-interferon (alpha-IFN, beta-IFN) are only partially effective in human breast cancer. To improve their effectiveness, it has been proposed that differentiation inducers, such as sodium butyrate (NaB), be used to increase the IFN sensitivity of tumors. Therefore, we assessed concomitant or sequential combinations of low/intermediate concentrations of alpha-IFN or beta-IFN (10, 50 and 100 IU/ml) with a low concentration (0.1 mM) of NaB on a human breast cancer cell line (MDA-MB231), which exhibited a moderate sensitivity to IFN. Moreover, to verify the capability of NaB to potentiate IFN effectiveness by increasing IFN receptor (IFN-R) concentration, we investigated the effect of NaB on the synthesis of IFN-R. The concomitant presence of NaB and alpha-IFN or beta-IFN significantly improved the antiproliferative effect of the corresponding IFN alone. Conversely, the sequential treatment NaB-IFN did not enhance the inhibitory activity of the cytokine, although NaB was able to induce the expression of IFN-R. More likely, NaB induced the expression of some component of the IFN system, such as Stat1, Stat2 or p48, whose higher availability in the cytoplasmic compartment promotes formation of the multimeric transcription factor ISGF3, which induces the transcription of IFN-stimulated genes.

Effect of sodium butyrate on human breast cancer cell lines.

We have investigated the effects exerted by sodium butyrate (NaBu) on the growth and cell cycle perturbations of four human breast cancer cell lines (MCF7, T47D, MDA-MB231 and BT20) with different steroid receptor profiles. Moreover, since one of the supposed mechanisms of action for NaBu activity involves the induction of apoptosis, we have studied the effects of NaBu on DNA fragmentation by agarose gel electrophoresis and flow cytometry. In all investigated cell lines, NaBu exerted a time- and dose-dependent inhibition of growth and caused a maximum inhibitory effect (85% to 90%) at the concentration of 2.5 mM. The inhibition was already evident after 3 days of treatment. The antiproliferative effect of NaBu was associated with a persistent block of cells in the G2M phase. The block was associated with apoptosis only in oestrogen-receptor positive cell lines. The inhibiting effect of NaBu in hormone-dependent and independent cell lines and its ability to induce apoptosis through a cell cycle perturbation in hormone-dependent cell lines may have important implications in the treatment of human tumours including breast cancer.

Combined effect of tamoxifen or interferon-beta and 4-hydroxyphenylretinamide on the growth of breast cancer cell lines.

To improve the effectiveness of 4-hydroxyphenylretinamide (4-HPR), an analogue of retinoic acid used in chemoprevention and treatment of breast cancer, we investigated the effect of concomitant administration of 4-HPR (0.1, 1 microM) and tamoxifen (TAM, 0.1, 1 microM), or 4-HPR and interferon-beta (IFN-beta, 10, 100, 500 IU/ml) on the growth of four cell lines (MCF7, T47D, MDA-MB231 and BT20) characterized by a different steroid receptor profile. A high concentration of 4-HPR caused a significant inhibitory effect not only on the estrogen receptor-positive cell lines (MCF7 and T47D), but also on one (BT20) of the two estrogen receptor-negative cell lines. IFN-beta displayed a dose-dependent inhibitory effect in all cell lines, but it was most evident in MCF7 cells. In all cell lines, the combination of 4-HPR (0.1 microM) and TAM (1 microM) or IFN-beta (500 IU/ml) generally caused additive or synergistic effects. In particular, the finding that in estrogen receptor-negative MDA-MB231 cells 4-HPR (which at 1 microM was singly ineffective) in combination with TAM at 1 microM or any concentration of IFN-beta produced a synergistic effect suggests that the compound could act through a pathway independent of specific receptors for retinoids. Our results indicate that intrinsic characteristics of cells can influence responsiveness to 4-HPR, TAM and IFN-beta, singly or in association, ever within cell lines with similar steroid receptor profiles. Thus, more attention should be payed to the biological characteristics of the single tumor in order to help choose the best combination of drugs to be applied.

The effect of ICI 164384 and beta-interferon on the growth and steroid receptor profile of breast cancer cell lines.

We investigated the effect of a concomitant treatment of ICI 164384 and B-interferon (beta-IFN) on the growth of estrogen-receptor-positive (ER+) and estrogen-receptor-negative (ER-) breast cancer cell lines and on their steroid receptor profiles. ICI 164384 reduced cell proliferation not only in ER+ but also in ER- cell lines and completely suppressed the stimulation induced by estradiol (E2) in hormone-sensitive cell lines, MCF7 and T47D. When associated with beta-IFN, ICI 164384 increased the inhibitory effect exerted by the low concentration of beta-IFN. Moreover, ICI 164384, singly or in association with beta-IFN, did not affect ER and PgR concentration in ER- cell lines, whereas in ER+ cell lines we observed an almost total disappearance of ER and PgR. In conclusion, our study seems to indicate that, although beta-IFN is able to control the proliferation of hormone-sensitive and hormone-independent subclones, it does not further improve the antiproliferative activity of ICI 164384. In contrast, the presence of ICI 164384, which does not induce the selection of resistant subclones under the same experimental conditions in which TAM does, may improve the efficacy of low concentration of beta-IFN and prevent the development of a secondary TAM-induced resistance.

Tamoxifen and beta-interferon: effect of simultaneous or sequential treatment on breast cancer cell lines.

The presence of steroid receptors in breast cancer is usually considered as a good indicator for responsiveness to antiestrogen treatment. However, not all estrogen-receptor (ER) positive tumors respond to tamoxifen, probably owing to the natural heterogeneity of breast cancer, which after tamoxifen treatment can result in an overgrowth of selected ER-negative clones. Since in a previous study we found that beta-interferon exerts an antiproliferative activity on several breast cancer cell lines, regardless of their steroid receptor status, we addressed our attention to the possibility of improving tamoifen efficacy by the addition of beta-interferon. Thus, we investigated the effect of concomitant or sequential treatment with tamoxifen and beta-interferon on breast cancer cell lines (MCF7, T47D, MDA-MB231, BT20) characterized by a different steroid receptor status. The results showed that beta-interferon caused a cell growth inhibition additional to that produced by tamoxifen, regardless of the receptor status of the cell lines. These findings indicate the potential of the combined treatment with tamoxifen and beta-interferon due to the direct effect of beta-interferon on ER-negative cells and the potentiation of tamoxifen activity in ER-positive cells, perhaps through the induction of steroid receptor synthesis.

The effect of alpha-, beta- and gamma-interferon on the growth of breast cancer cell lines.

We investigated the antiproliferative effect of different concentrations (10, 100, 1000 IU/ml) of alpha-, beta- and gamma-interferons (IFN) on breast cancer cell lines. Cell lines were treated with IFN, in the absence or in the presence of estradiol for 9 days, and the effect on growth was evaluated as variation in DNA content. Inhibitory effect varied as a function of the type of interferon and cell line. Alpha-IFN and gamma-IFN were effective only at 1000 IU/ml and not in all cell lines, whereas a maximum effect was observed for beta-IFN regardless of the steroid receptor status of cell lines. In fact, a significant growth inhibition was observed at the intermediate concentration of 100 IU/ml in all but MDA-MB231 cell lines. Moreover, in both estrogen-receptor positive cell lines, beta-IFN counteracted growth stimulation induced by estradiol and showed a strong antiestrogenic activity. In conclusion, our results show that beta-IFN is the most active among the IFNs tested and suggest its usefulness in the treatment of all breast cancers, irrespective of their steroid receptor status.