5 research outputs found
Identification and Characterization of Dual Inhibitors of the USP25/ 28 Deubiquitinating Enzyme Subfamily
The
ubiquitin proteasome system is widely postulated to be a new
and important field of drug discovery for the future, with the ubiquitin
specific proteases (USPs) representing one of the more attractive
target classes within the area. Many USPs have been linked to critical
axes for therapeutic intervention, and the finding that USP28 is required
for c-Myc stability suggests that USP28 inhibition may represent a
novel approach to targeting this so far undruggable oncogene. Here,
we describe the discovery of the first reported inhibitors of USP28,
which we demonstrate are able to bind to and inhibit USP28, and while
displaying a dual activity against the closest homologue USP25, these
inhibitors show a high degree of selectivity over other deubiquitinases
(DUBs). The utility of these compounds as valuable probes to investigate
and further explore cellular DUB biology is highlighted by the demonstration
of target engagement against both USP25 and USP28 in cells. Furthermore,
we demonstrate that these inhibitors are able to elicit modulation
of both the total levels and the half-life of the c-Myc oncoprotein
in cells and also induce apoptosis and loss of cell viability in a
range of cancer cell lines. We however observed a narrow therapeutic
index compared to a panel of tissue-matched normal cell lines. Thus,
it is hoped that these probes and data presented herein will further
advance our understanding of the biology and tractability of DUBs
as potential future therapeutic targets
FLCN, a novel autophagy component, interacts with GABARAP and is regulated by ULK1 phosphorylation
Birt-Hogg-Dubé (BHD) syndrome is a rare autosomal dominant condition caused by mutations in the FLCN gene and characterized by benign hair follicle tumors, pneumothorax, and renal cancer. Folliculin (FLCN), the protein product of the FLCN gene, is a poorly characterized tumor suppressor protein, currently linked to multiple cellular pathways. Autophagy maintains cellular homeostasis by removing damaged organelles and macromolecules. Although the autophagy kinase ULK1 drives autophagy, the underlying mechanisms are still being unraveled and few ULK1 substrates have been identified to date. Here, we identify that loss of FLCN moderately impairs basal autophagic flux, while re-expression of FLCN rescues autophagy. We reveal that the FLCN complex is regulated by ULK1 and elucidate 3 novel phosphorylation sites (Ser406, Ser537, and Ser542) within FLCN, which are induced by ULK1 overexpression. In addition, our findings demonstrate that FLCN interacts with a second integral component of the autophagy machinery, GABA(A) receptor-associated protein (GABARAP). The FLCN-GABARAP association is modulated by the presence of either folliculin-interacting protein (FNIP)-1 or FNIP2 and further regulated by ULK1. As observed by elevation of GABARAP, sequestome 1 (SQSTM1) and microtubule-associated protein 1 light chain 3 (MAP1LC3B) in chromophobe and clear cell tumors from a BHD patient, we found that autophagy is impaired in BHD-associated renal tumors. Consequently, this work reveals a novel facet of autophagy regulation by ULK1 and substantially contributes to our understanding of FLCN function by linking it directly to autophagy through GABARAP and ULK1
A role of autophagy in PTP4A3-driven cancer progression
Autophagy, a "self-eating" cellular process, has dual roles in promoting and suppressing tumor growth, depending on cellular context. PTP4A3/PRL-3, a plasma membrane and endosomal phosphatase, promotes multiple oncogenic processes including cell proliferation, invasion, and cancer metastasis. In this study, we demonstrate that PTP4A3 accumulates in autophagosomes upon inhibition of autophagic degradation. Expression of PTP4A3 enhances PIK3C3-BECN1-dependent autophagosome formation and accelerates LC3-I to LC3-II conversion in an ATG5-dependent manner. PTP4A3 overexpression also enhances the degradation of SQSTM1, a key autophagy substrate. These functions of PTP4A3 are dependent on its catalytic activity and prenylation-dependent membrane association. These results suggest that PTP4A3 functions to promote canonical autophagy flux. Unexpectedly, following autophagy activation, PTP4A3 serves as a novel autophagic substrate, thereby establishing a negative feedback-loop that may be required to fine-tune autophagy activity. Functionally, PTP4A3 utilizes the autophagy pathway to promote cell growth, concomitant with the activation of AKT. Clinically, from the largest ovarian cancer data set (GSE 9899, n = 285) available in GEO, high levels of expression of both PTP4A3 and autophagy genes significantly predict poor prognosis of ovarian cancer patients. These studies reveal a critical role of autophagy in PTP4A3-driven cancer progression, suggesting that autophagy could be a potential Achilles heel to block PTP4A3-mediated tumor progression in stratified patients with high expression of both PTP4A3 and autophagy genes
Identification and Structure-Guided Development of Pyrimidinone Based USP7 Inhibitors
Ubiquitin
specific protease 7 (USP7, HAUSP) has become an attractive
target in drug discovery due to the role it plays in modulating Mdm2
levels and consequently p53. Increasing interest in USP7 is emerging
due to its potential involvement in oncogenic pathways as well as
possible roles in both metabolic and immune disorders in addition
to viral infections. Potent, novel, and selective inhibitors of USP7
have been developed using both rational and structure-guided design
enabled by high-resolution cocrystallography. Initial hits were identified
via fragment-based screening, scaffold-hopping, and hybridization
exercises. Two distinct subseries are described along with associated
structure–activity relationship trends, as are initial efforts
aimed at developing compounds suitable for <i>in vivo</i> experiments. Overall, these discoveries will enable further research
into the wider biological role of USP7