Maspin enhances cisplatin chemosensitivity in bladder cancer T24 and 5637 cells and correlates with prognosis of muscle-invasive bladder cancer patients receiving cisplatin based neoadjuvant chemotherapy

Maspin is a 42-kDa protein associated with various tumor related processes such as the inhibition of cell migration, cell invasion, angiogenesis, as well as improvement in cell adhesion and the induction of programmed cell death, thus classifying it as a tumor suppressor [15–17]. Emerging evidence shows that Maspin expression was preserved in superficial bladder cancers and its expression associated with good prognosis. Kramer [18] investigated specimens from 162 non-muscle invasive bladder cancer patients treated by transurethral resection. They showed positive Maspin expression rate was 75.9 % and had a longer PFS compared with negative expression (46 vs 18 months, respectively). And Acikalin, Blandamura also concluded similar results on Maspin expression and superficial bladder cancer [19, 20]. In our study, there were 23 (37.1 %) cases of Maspin (+) in 62 MIBC cases. The OS and PFS rate among patients with Maspin (?) were significantly lower than these among patients with Maspin (+) expression (P?=?0.0036 and 0.0005, respectively). This expression rate was lower than the reported results in superficial bladder cancer while it correlated with the Beecken’s study [10], who showed Maspin was significantly diminished in MIBC and patients with high Maspin expression had relatively good prognosis.

Aa a tumor suppressor, Maspin has been found to increase cell apoptosis induced by chemo-regents in several types of cancers. Jiang et al. found Maspin transfected MDA-MB-435 breast carcinoma cells displayed increased sensitivity to staurosporine induced apoptosis [21]. Ben et al. testified Maspin could mediate E2F1-induced apoptosis of U2OS and SAOS2 osteosarcoma cells to doxorubicin and cisplatin [22]. Additional supporting evidence using prostate cancer cells had also reported that Maspin induced apoptosis sensitivity to chemical reagents [23], as well as increased the rate of apoptosis related to inhibition of proteosomal pathways [24].

In addition to surgical treatment, chemotherapy is an important strategy for the therapy of MIBC. Cisplatin based chemotherapy is widely used in bladder cancer treatment [25]. However, a significant proportion of patients develop disease progression after an initial response to chemotherapy. The drug resistance affect the therapeutic efficacy of bladder cancer treatment [26]. Recently, several molecules and reagents had been found to enhance chemosensitivity in bladder cancer, such as miR-101, TFAP2a, LRIG1, 5-Azacytidine, LY294002, et al. [27–32]. But the crosstalk between Maspin and cisplatin based chemosensitiviy is rare till now. In the present study, our results showed that Maspin overexpression increased the cell apoptosis and enhanced chemosensitivity of both T24 and 5637 cells to cisplatin, with the IC50 value decreasing from 23.32?±?4.30 mg/l to 15.30?±?2.10 mg/l and from 4.91?±?0.42 mg/l to 2.14?±?0.21 mg/l, respectively. Further more, in MIBC patients treating with NACT plus RC-PLND, Maspin expression was correlated with the prognosis, showing the OS and PFS rates were much longer in Maspin(+) group.

Bcl-2, a 26-kDa integral, is a powerful endogenous anti-apoptosis protein that in response to a variety of cell death signals [33]. Research showed it negatively regulated the activation of Caspase-3, the key executioner of apoptosis, which functions as an effector of mammalian cell death pathways. Downexpression of Bcl-2 promote Caspase activities and apoptosis [34]. The present study results show upregulation of the Maspin expression in T24 and 5637 cells, resulting in a decrease in the Bcl-2 level, with a concurrent increase in the Caspase-3 activation. Recent evidence indicates that AKT is frequently activated in many types of human cancer, and activated PI3K/AKT/mTOR signaling pathway is a significant contributor to tumor progression and poor prognosis, including bladder cancer [35]. Moreover, AKT overexpression is often associated with resistance to cisplatin chemotherapy, such as in ovarian and lung cancer, et al. [36, 37]. Sun et al. testified MK2206, a highly potent and selective allosteric AKT inhibitor, could promote the cisplatin-induced cytotoxicity and apoptosis in urothelial cancer cell lines through inhibiting the Akt pathway [38]. Wu and his colleagues found LY294002, a PI3K inhibitor, could suppress proliferation and sensitize doxorubicin chemotherapy in bladder cancer EJ cells. Our results showed that compared with control group, though AKT expression did’t have significant changes, p-AKT, PI3K and mTOR expression were significantly decreased in Maspin group. So, the above results suggest upregulating Maspin expression could enhance cisplatin chemosensitivity and was induced by cell apoptosis through PI3K/AKT/mTOR signaling pathway in T24 and 5637 cells.

As concluded by Berardi, there are three strategies to re-activate or increase Maspin expression [8]. The first one is designing transcription factors, to hit the system that inhibits the expression of maspin, such as artificial transcription factors (ATFs) and chromatin remodeling drugs [39]. In our previous study, we found a DNA methyltransferase inhibitor, 5-aza-2?-deoxycytidine (5-Aza-CdR), could reactivate Maspin expression and inhibit the proliferation, migration, and invasion of T24 cells, which may serve as a potential strategy for the treatment of bladder cancer [40]. The second one is identifying natural substances that can determine the activation and the expression of Maspin, such as curcumin, a hydrophobic polyphenol derived from turmeric [41]. And Last but not least, gene therapy capable of up-regulating the Maspin in cancer cell, such as Maspin DNA-liposome therapy and adeno-associated virus (AAV, serotype 2) vector encoding Maspin [42, 43].