Synergistic effect of Nutlin-3 combined with MG-132 on schwannoma cells through restoration of merlin and p53 tumour suppressors

Abstract

BackgroundThe great majority of sporadic vestibular schwannomas (VSs) are due to the mutations of the NF2 gene encoding merlin. Sporadic VSs exhibit variable growth patterns and only a small fraction of the tumours are fast-growing; however, the underlying mechanisms remain undefined.MethodsDNA sequencing and dosage analysis were used to identify the NF2 mutation status in sporadic schwannomas. The expression and sub-cellular localization of merlin and p53-MDM2 were assessed by immunoblotting, qRT-PCR and immunofluorescence. In vitro and in vivo studies were performed to reveal the effects of Nutlin-3 (a MDM2 inhibitor) and/or MG-132(a proteasome inhibitor) on schwannomas. The proliferation of schwannoma cells was assessed by CCK-8 assay, EdU staining and Flow cytometry analysis.FindingsDouble genetic hits of NF2 tended to occur in fast-growing tumours, characterized by the absence of merlin. The deregulation of p53-MDM2 was demonstrated to mediate merlin-deficient tumour growth, characterized by a nuclear accumulation of stabilized MDM2, contributing to a nuclear export of p53 for degradation. Nutlin-3 blocked the proliferation of schwannoma cells via a cooperative recovery of merlin and p53, accompanied by the shuttling of both proteins from the cytoplasm to the nucleus. We further demonstrated a difference in the sensitivity to Nutlin-3 between schwannoma cells with and without merlin expression. Nutlin-3 combined with MG-132 narrowed this between-group difference and triggered stronger inhibitory effects on the growth of schwannomas through coordinated reactivation of p53.InterpretationThese findings present treatment strategies directed on the pathogenesis of sporadic schwannomas.FundNational Natural Science Foundation of China.

Helping the Released Guardian: Drug Combinations for Supporting the Anticancer Activity of HDM2 (MDM2) Antagonists

“…The effect of MG-132 (one of the early proteasome inhibitors) and nutlin-3 on schwannoma cells has been assessed by Chen and colleagues. They have demonstrated that the differences in the expression of merlin, which is a tumor suppressor protein, in cancer cultures explain the functional diversity in response to nutlin-3 treatment [98]. It has been proved that dual therapy of nutlin-3 /MG-132 narrows the differences in the sensitivity to nutlin-3 in schwannoma cells.…”

“…It has been proved that dual therapy of nutlin-3 /MG-132 narrows the differences in the sensitivity to nutlin-3 in schwannoma cells. In addition, in vivo experiment with human xenograft model of schwannoma has revealed that the small-molecules work together to impose apoptosis [98].…”

Therapy of Sporadic and NF2-Related Vestibular Schwannoma

“…Weak or non-bands were found in 'two-hit' tumors, showing that the inactivation of both alleles is necessary for the total loss of merlin expression. At least one somatic mutation occurred in more than 85% of the sporadic VS cases that affected the function of the NF2 gene [33].…”

“…The term "hit" means a DNA mutation in a cell [32]. The "two-hit" inactivation of NF2 leads to the loss of merlin [33]. This theory also involves the loss of chromosome 22 and biallelic genetic alterations of the NF2 gene are seen in the majority of the sporadic VS cases [26].…”

“…Weak or non-bands were found in 'two-hit' tumors, showing that the inactivation of both alleles is necessary for the total loss of merlin expression. At least one somatic mutation occurred in more than 85% of the sporadic VS cases that affected the function of the NF2 gene [33].Another possible mechanism for the inactivation of NF2 in sporadic VS is epigenetic modification. Cytosine phosphate guanine (CpG) islands are the stretches of the regions of the genome.…”