RABL6A Is an Essential Driver of MPNSTs that Negatively Regulates the RB1 Pathway and Sensitizes Tumor Cells to CDK4/6 Inhibitors

Abstract

Purpose: Malignant peripheral nerve sheath tumors (MPNSTs) are deadly sarcomas that lack effective therapies. In most MPNSTs, the retinoblastoma (RB1) tumor suppressor is disabled by hyperactivation of cyclin dependent kinases (CDKs), commonly through loss of CDK inhibitory proteins such as p27(Kip1). RABL6A is an inhibitor of RB1 whose role in MPNSTs is unknown. To gain insight into MPNST development and establish new treatment options, we investigated RABL6A-RB1 signaling and CDK inhibitor-based therapy in MPNSTs. Experimental Design:We examined patient-matched MPNSTs and precursor lesions by RNA-Seq and IHC. Molecular and biological effects of silencing RABL6A and/or p27 in MPNST lines and normal human Schwann cells were determined. Tumor suppressive effects of CDK inhibitors were measured in MPNST cells and orthotopic tumors.Results: RABL6A was dramatically upregulated in human MPNSTs compared to precursor lesions, which correlated inversely with p27 levels. Silencing RABL6A caused MPNST cell death and G1 arrest that coincided with p27 upregulation, CDK downregulation and RB1 activation. The growth suppressive effects of RABL6A loss, and its regulation of RB1, were largely rescued by p27 depletion. Importantly, reactivation of RB1 using a CDK4/6 inhibitor (palbociclib) killed MPNST cells in vitro in a RABL6A-dependent manner and suppressed MPNST growth in vivo. Low-dose combination of drugs targeting multiple RB1 kinases (CDK4/6, CDK2) had enhanced anti-tumorigenic activity associated with potential MPNST cell redifferentiation.Conclusions: RABL6A is a new driver of MPNST pathogenesis that acts in part through p27-RB1 inactivation. Our results suggest RB1 targeted therapy with multiple pathway drugs may effectively treat MPNSTs.

CDKs in Sarcoma: Mediators of Disease and Emerging Therapeutic Targets

“…One study showed that cytoplasmic p27 KIP1 localization correlates with increased nuclear cyclin E1 expression and poor prognosis in MPNST patients [83]. Most recently, immunohistochemical analyses of patient matched PNFs and MPNSTs revealed dramatic upregulation of an oncogenic GTPase, RABL6A, in MPNSTs [84]. RABL6A is a newly recognized negative regulator of RB1 and p53 signaling [85][86][87].…”

“…RABL6A is a newly recognized negative regulator of RB1 and p53 signaling [85][86][87]. In MPNSTs, RABL6A was found to promote MPNST pathogenesis, in part, through its downregulation of p27 KIP1 , activation of CDKs, and inhibition of RB1 [84].…”

“…Second, monotherapy with CDK4/6 inhibitors is often over-ridden by CDK2-mediated phosphorylation of RB1, which can bypass the need for CDK4/6 and drive resistance to CDK4/6 inhibitors [190,197]. Studies in MPNSTs highlight this concept, where sustained CDK4/6 inhibition leads to acquired drug resistance that can be blunted by concomitant CDK2 inhibition [84]. Importantly, tumor cells have numerous mechanisms of resistance to CDK4/6 inhibitors at their disposal, which are both cell cycle specific (e.g., CCNE amplification and CDK2 hyperactivation) and independent of the cell cycle (e.g., activation of PI3K/AKT/mTOR signaling) (reviewed in [198]).…”

From Genes to -Omics: The Evolving Molecular Landscape of Malignant Peripheral Nerve Sheath Tumor

“…These investigations have confirmed or supplemented the genomic data by assaying downstream pathway effects in human MPNST. Several studies have broadly analyzed gene expression in human MPNST samples using microarray or RNAseq approaches [ 48 , 88 ]; these data can be examined in relation to known genetic changes to generate additional hypotheses for effects on downstream signaling pathways. Recent work compared gene expression in multiple functional pathways across pNF, ANF/ANNUBP, and MPNST and found that some ANNUBP share signaling pathway characteristics that more closely resemble pNF (e.g., ERK/MAPK) and others (e.g., AKT/mTOR) are more similar to MPNST [ 88 ].…”

“…Several studies have broadly analyzed gene expression in human MPNST samples using microarray or RNAseq approaches [ 48 , 88 ]; these data can be examined in relation to known genetic changes to generate additional hypotheses for effects on downstream signaling pathways. Recent work compared gene expression in multiple functional pathways across pNF, ANF/ANNUBP, and MPNST and found that some ANNUBP share signaling pathway characteristics that more closely resemble pNF (e.g., ERK/MAPK) and others (e.g., AKT/mTOR) are more similar to MPNST [ 88 ]. Phospho-proteome arrays may be used to investigate kinase signaling in relation to various genomic alterations or therapeutic interventions in MPNST; to date this has primarily been used in MPNST cell lines or animal models (see article by Grit et al in this Special Issue on Genomics and Models of Nerve Sheath Tumors) [ 89 ].…”