Evaluation of Antitumoral activity of bone targeted drugs/conventional chemotherapies and identification of biomarkers for the selection of patients with breast cancer for the bone targeted therapy in adjuvant setting

Bone metastases are responsible for different clinical complications defined as skeletal-related events (SREs) such as pathologic fractures, spinal cord compression, hypercalcaemia, bone marrow infiltration and severe bone pain requiring palliative radiotherapy. The general aim of these three years research period was to improve the management of patients with bone metastases through two different approaches of translational research. Firstly in vitro preclinical tests were conducted on breast cancer cells and on indirect co-colture of cancer cells and osteoclasts to evaluate bone targeted therapy singly and in combination with conventional chemotherapy. The study suggests that zoledronic acid has an antitumor activity in breast cancer cell lines. Its mechanism of action involves the decrease of RAS and RHO, as in osteoclasts. Repeated treatment enhances antitumor activity compared to non-repeated treatment. Furthermore the combination Zoledronic Acid + Cisplatin induced a high antitumoral activity in the two triple-negative lines MDA-MB-231 and BRC-230. The p21, pMAPK and m-TOR pathways were regulated by this combined treatment, particularly at lower Cisplatin doses. A co-colture system to test the activity of bone-targeted molecules on monocytes-breast conditioned by breast cancer cells was also developed. Another important criticism of the treatment of breast cancer patients, is the selection of patients who will benefit of bone targeted therapy in the adjuvant setting. A retrospective case-control study on breast cancer patients to find new predictive markers of bone metastases in the primary tumors was performed. Eight markers were evaluated and TFF1 and CXCR4 were found to discriminate between patients with relapse to bone respect to patients with no evidence of disease. In particular TFF1 was the most accurate marker reaching a sensitivity of 63% and a specificity of 79%. This marker could be a useful tool for clinicians to select patients who could benefit for bone targeted therapy in adjuvant setting.

Ruolo dei recettori degli estrogeni nella trasduzione del segnale nella neoplasia ovarica

Oestrogen induction of cell proliferation is critical in carcinogenesis of gynaecologic tissues. The effects of oestrogens are mediated by Oestrogen receptor (ER) ERα and ERβ, which are members of the nuclear steroid receptor superfamily. The balance between the ERα/ERβ levels seems critical during carcinogenesis due to their different role in proliferation and apoptosis. SERMs are a class of drugs targeting ERs used especially in the treatment of breast cancer, that despite their usefulness, cause side effects. Therefore, it’s important to develop new active molecules without side effects. In a previous work Andreani et al.(2007) investigated the antitumor activity of a new class of indole-derivatives in 60 different human cancer cell lines. In particular they noted that compound named 3L was able to induce a strong antiproliferative effect in cell lines derived from breast, cervix, ovary ,CNS and colon. The aim of this thesis is to characterize the biological effect in ovarian carcinoma cells (IGROV-1), colon adenocarcinoma cells (HT29), cervix adenocarcinoma cells (HelaS3) and breast cancer cells (MCF7). Among the effect exerted on the other cell lines, the most interesting is the cytostatic effect on IGROV-1. In order to identify the 3L molecular target we monitored the 3L concentration in the IGROV-1 nuclear fractions. The analysis revealed that the drug localizes in the nucleus starting from 6 hrs after treatment, suggesting a nuclear target. The stimulation with oestrogen did not increase the proliferation rate in 3L treated cells, suggesting a possible involvement with oestrogen receptors. Due to the 3L fluorescent properties, we demonstrated a colocalization between the ER and the 3L compound. In particular, a chromatin binding assay revealed the presence of a 3L-ERβ complex bound to DNA, interaction that may be the cause of the observed antiproliferative effect.