title: Specific CP110 Phosphorylation Sites Mediate Anaphase Catastrophe after CDK2 Inhibition: Evidence for Cooperation with USP33 Knockdown.
abstract: Chromosomal instability (CIN) is a hallmark of solid tumor biology and is implicated in carcinogenesis. Preferentially eliminating malignant cells by targeting CIN and aneuploidy is an attractive antineoplastic strategy. We previously reported that CDK2 antagonism causes lung cancer cells to undergo anaphase catastrophe and apoptosis through inhibition of phosphorylation of the centrosomal protein CP110. Cells with activating KRAS mutations were particularly sensitive to CDK2 inhibition due to downregulation of CP110 protein levels. This study investigated mechanisms of CDK2 antagonism that mediate anaphase catastrophe via changes in CP110 protein expression and how activated KRAS affects CP110 levels in lung cancers. Site-directed mutagenesis revealed candidate CDK phosphorylation sites of CP110 (residues Ser 170 and Thr 194) critical for conferring anaphase catastrophe by altering centrosome clustering in mitosis. Intriguingly, KRAS mutation can promote CP110 protein degradation by upregulating the ubiquitin ligase SCF(cyclinF), which targets CP110 protein for destabilization. Finally, CDK2 inhibitor response was enhanced when combined with knockdown of the deubiquitinase USP33 that in turn accelerates CP110 protein degradation. Thus, this study provides molecular pharmacologic insights into how CP110 expression regulates response to CDK2 inhibition. An improved understanding of in vitro antineoplastic mechanisms of combining CDK2 antagonism with induced CP110 repression provides a rationale for exploring clinical consequences of this strategy. Taken together, preclinical findings obtained from combining CDK2 inhibition with USP33 repression have implications for treating patients with non-small cell lung cancers.
authors: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Shanhu
authors: S
authors: Hu
authors: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Yun
authors: Y
authors: Lu
authors: Department of Biochemistry, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Bernardo
authors: B
authors: Orr
authors: Department of Biochemistry, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Kristina
authors: K
authors: Godek
authors: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Lisa Maria
authors: LM
authors: Mustachio
authors: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Masanori
authors: M
authors: Kawakami
authors: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: David
authors: D
authors: Sekula
authors: Department of Biochemistry, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire. Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Duane A
authors: DA
authors: Compton
authors: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire.
authors: Sarah
authors: S
authors: Freemantle
authors: Department of Pharmacology and Toxicology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire. Norris Cotton Cancer Center, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire. Department of Medicine, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire, and Dartmouth-Hitchcock Medical Center, Lebanon, New Hampshire. edmitrovsky@mdanderson.org.
authors: Ethan
authors: E
authors: Dmitrovsky
mesh_terms: D000705:Anaphase / Q000187:drug effects* / Q000235:genetics; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D018385:Centrosome / Q000187:drug effects / Q000378:metabolism; D051357:Cyclin-Dependent Kinase 2 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016213:Cyclins / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000378:metabolism; D011687:Purines / Q000494:pharmacology; D034622:RNA Interference; D000077546:Roscovitine; D012694:Serine / Q000235:genetics / Q000378:metabolism; D013912:Threonine / Q000235:genetics / Q000378:metabolism; D043222:Ubiquitin Thiolesterase / Q000235:genetics / Q000378:metabolism*
chemical_list: C503669:CCNF protein, human; C467559:CCP110 protein, human; D018797:Cell Cycle Proteins; D016213:Cyclins; C117307:KRAS protein, human; D008869:Microtubule-Associated Proteins; D010750:Phosphoproteins; D047428:Protein Kinase Inhibitors; D011687:Purines; D000077546:Roscovitine; D013912:Threonine; D012694:Serine; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; C449799:USP33 protein, human; D043222:Ubiquitin Thiolesterase; D016283:Proto-Oncogene Proteins p21(ras)