Publications
Year of publication 2017
Targeting Conformational Activation of CDK2 Kinase.
Biotechnology journal : 12 : 1600531 : DOI : 10.1002/biot.201600531 Learn moreSummary
Cyclin-dependent kinases constitute attractive pharmacological targets for cancer therapeutics, yet inhibitors in clinical trials target the ATP-binding pocket of the CDK and therefore suffer from limited selectivity and emergence of resistance. The more recent development of allosteric inhibitors targeting conformational plasticity of protein kinases offers promising perspectives for therapeutics. In particular tampering with T-loop dynamics of CDK2 kinase would provide a selective means of inhibiting this kinase, by preventing its conformational activation. To this aim we engineered a fluorescent biosensor that specifically reports on conformational changes of CDK2 activation loop and is insensitive to ATP or ATP-competitive inhibitors, which constitutes a highly sensitive probe for identification of selective T-loop modulators. This biosensor was successfully applied to screen a library of small chemical compounds leading to discovery of a family of quinacridine analogs, which potently inhibit cancer cell proliferation, and promote accumulation of cells in S phase and G2. These compounds bind CDK2/ Cyclin A, inhibit its kinase activity, compete with substrate binding, but not with ATP, and dock onto the T-loop of CDK2. The best compound also binds CDK4 and CDK4/Cyclin D1, but not CDK1. The strategy we describe opens new doors for the discovery of a new class of allosteric CDK inhibitors for cancer therapeutics.
Fold upExploring the mechanism of inhibition of human telomerase by cysteine-reactive compounds.
FEBS letters : 591 : 863-874 : DOI : 10.1002/1873-3468.12589 Learn moreSummary
Telomerase is an almost universal cancer target that consists minimally of a core protein (hTERT) and an RNA (hTR). Some inhibitors of this enzyme are thought to function by the covalent binding to one or several cystein residues; however, this inhibition mechanism has never been investigated because of the difficulty in producing telomerase. In the present study, we use a recent method to produce recombinant hTERT to analyse the effect of cysteine reactive inhibitors on telomerase. Using mass-spectrometry (MS) and mutagenesis analysis, we identify several targeted residues in separated domains of the hTERT protein and show that cysteine-reactive reagents abolish the interaction with the CR4/5 region of hTR. This article is protected by copyright. All rights reserved.
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CD45 phosphatase is crucial for human and murine acute myeloid leukemia maintenance through its localization in lipid rafts.
Oncotarget : 7 : 64785-64797 : DOI : 10.18632/oncotarget.11622 Learn moreSummary
CD45 is a pan-leukocyte protein with tyrosine phosphatase activity involved in the regulation of signal transduction in hematopoiesis. Exploiting CD45 KO mice and lentiviral shRNA, we prove the crucial role that CD45 plays in acute myeloid leukemia (AML) development and maintenance. We discovered that CD45 does not colocalize with lipid rafts on murine and human non-transformed hematopoietic cells. Using a mouse model, we proved that CD45 positioning within lipid rafts is modified during their oncogenic transformation to AML. CD45 colocalized with lipid rafts on AML cells, which contributes to elevated GM-CSF signal intensity involved in proliferation of leukemic cells. We furthermore proved that the GM-CSF/Lyn/Stat3 pathway that contributes to growth of leukemic cells could be profoundly affected, by using a new plasma membrane disrupting agent, which rapidly delocalized CD45 away from lipid rafts. We provide evidence that this mechanism is also effective on human primary AML samples and xenograft transplantation. In conclusion, this study highlights the emerging evidence of the involvement of lipid rafts in oncogenic development of AML and the targeting of CD45 positioning among lipid rafts as a new strategy in the treatment of AML.
Fold upRespiratory syncytial virus infection in macaques is not suppressed by intranasal sprays of pyrimidine biosynthesis inhibitors.
Antiviral research : 58-62 : DOI : 10.1016/j.antiviral.2015.11.006 Learn moreSummary
There is imperious need for efficient therapies against ubiquitous and life-threatening respiratory viruses, foremost among them being the human respiratory syncytial virus (hRSV). Several research groups who performed functional screens for broad-spectrum antivirals identified compounds targeting the de novo pyrimidine biosynthesis pathway. Despite their strong antiviral activity in vitro, whether such antimetabolites are effective in vivo remains highly controversial. Here, we evaluated two potent pyrimidine biosynthesis inhibitors developed in our laboratory, IPPA17-A04 and GAC50, in a model of mild hRSV-infection in cynomolgus macaques. In this model, hRSV replication is restricted to the epithelium of the upper respiratory tract, and is compatible with a topical treatment by intranasal sprays. The local administration of palivizumab, a neutralizing anti-hRSV antibody used in clinics, significantly reduced virus replication. In contrast, pyrimidine biosynthesis inhibitors did not show any inhibitory effect on hRSV growth when delivered topically as experimented in our model. Our results should help to better define the potential applications of this class of antimetabolites in the treatment of viral infections.
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