Mitotic Kinase Aurora Research Articles

Aurora B/AIR-2 regulates sister centromere resolution and CENP-A/HCP-3 organization to prevent merotelic attachments.

During cell division, the accurate capture of sister kinetochores that are built on the centromeres of chromosomes by microtubules emanating from opposite spindle poles governs faithful chromosome segregation. To ensure sister chromatids separate correctly, sister centromeres undergo resolution to achieve bi-polar orientation prior to microtubule attachments. Failure of centromere resolution increases the frequency of merotelic attachments, with microtubules from opposite poles attaching to the same sister kinetochore, causing lagging chromosome, aneuploidy, and even cancer progression. The Aurora B-mediated tension-sensing machinery to correct erroneous kinetochore-microtubule attachments has been well studied. However, preventative mechanisms to avoid merotelic attachments that occur in the earlier mitotic stage are poorly understood. In this study, we found that inactivation of mitotic kinase Aurora B/AIR-2 increases merotelic attachments in Caenorhabditis elegans. On one hand, Aurora B/AIR-2-deficient cells exhibited a delay in the occurrence of centromere resolution and a disruption in targeting condensin II components to chromatin. On the other hand, loss of Aurora B/AIR-2 results in an increased localization of centromeric proteins CENP-A/HCP-3 and M18BP1/KNL-2 as well as the kinetochore protein MIS-12 on chromatin, which may generate ectopic kinetochores causing erroneous attachments. To conclude, this study elucidated that Aurora B/AIR-2 regulates sister centromere resolution and CENP-A/HCP-3 deposition to actively prevent merotely and chromosome instability in cells.

Bipartite binding interface recruiting HP1 to chromosomal passenger complex at inner centromeres.

Maintenance of ploidy depends on the mitotic kinase Aurora B, the catalytic subunit of the chromosomal passenger complex (CPC) whose proficient activity is supported by HP1 enriched at inner centromeres. HP1 is known to associate with INCENP of the CPC in a manner that depends on the PVI motif conserved across HP1 interactors. Here, we found that the interaction of INCENP with HP1 requires not only the PVI motif but also its C-terminally juxtaposed domain. Remarkably, these domains conditionally fold the β-strand (PVI motif) and the α-helix from a disordered sequence upon HP1 binding and render INCENP with high affinity to HP1. This bipartite binding domain termed SSH domain (Structure composed of Strand and Helix) is necessary and sufficient to attain a predominant interaction of HP1 with INCENP. These results identify a unique HP1-binding module in INCENP that ensures enrichment of HP1 at inner centromeres, Aurora B activity, and thereby mitotic fidelity.

The people behind the papers - Nadia Manzi and Daniel Dickinson.

The mitotic kinase Aurora A has been shown to regulate the anterior-posterior polarity in developing Caenorhabditis elegans embryos. In a new study, Daniel Dickinson and colleagues find that Aurora A has temporally distinct roles in coordinating the localization of Partitioning defective (PAR) proteins to establish cell polarity during development. To find out more about the story behind the paper, we caught up with first author Nadia Manzi and corresponding author Daniel Dickinson, Assistant Professor at the University of Texas at Austin.

Aurora kinase A-mediated phosphorylation triggers structural alteration of Rab1A to enhance ER complexity during mitosis.

Morphological rearrangement of the endoplasmic reticulum (ER) is critical for metazoan mitosis. Yet, how the ER is remodeled by the mitotic signaling remains unclear. Here, we report that mitotic Aurora kinase A (AURKA) employs a small GTPase, Rab1A, to direct ER remodeling. During mitosis, AURKA phosphorylates Rab1A at Thr75. Structural analysis demonstrates that Thr75 phosphorylation renders Rab1A in a constantly active state by preventing interaction with GDP-dissociation inhibitor (GDI). Activated Rab1A is retained on the ER and induces the oligomerization of ER-shaping protein RTNs and REEPs, eventually triggering an increase of ER complexity. In various models, from Caenorhabditis elegans and Drosophila to mammals, inhibition of Rab1AThr75 phosphorylation by genetic modifications disrupts ER remodeling. Thus, our study reveals an evolutionarily conserved mechanism explaining how mitotic kinase controls ER remodeling and uncovers a critical function of Rab GTPases in metaphase.

Design and synthesis of (E)-3-benzylideneindolin-2-one derivatives as potential allosteric inhibitors of Aurora A kinase

The mitotic kinase Aurora A, a pivotal regulator of cell cycle, is overexpressed in various cancers and has emerged as one of the most promising targets for anticancer drug discovery....

A Network of 17 Microtubule-Related Genes Highlights Functional Deregulations in Breast Cancer.

A wide panel of microtubule-associated proteins and kinases is involved in coordinated regulation of the microtubule cytoskeleton and may thus represent valuable molecular markers contributing to major cellular pathways deregulated in cancer. We previously identified a panel of 17 microtubule-related (MT-Rel) genes that are differentially expressed in breast tumors showing resistance to taxane-based chemotherapy. In the present study, we evaluated the expression, prognostic value and functional impact of these genes in breast cancer. We show that 14 MT-Rel genes (KIF4A, ASPM, KIF20A, KIF14, TPX2, KIF18B, KIFC1, AURKB, KIF2C, GTSE1, KIF15, KIF11, RACGAP1, STMN1) are up-regulated in breast tumors compared with adjacent normal tissue. Six of them (KIF4A, ASPM, KIF20A, KIF14, TPX2, KIF18B) are overexpressed by more than 10-fold in tumor samples and four of them (KIF11, AURKB, TPX2 and KIFC1) are essential for cell survival. Overexpression of all 14 genes, and underexpression of 3 other MT-Rel genes (MAST4, MAPT and MTUS1) are associated with poor breast cancer patient survival. A Systems Biology approach highlighted three major functional networks connecting the 17 MT-Rel genes and their partners, which are centered on spindle assembly, chromosome segregation and cytokinesis. Our studies identified mitotic Aurora kinases and their substrates as major targets for therapeutic approaches against breast cancer.

Amorphous silica nanoparticles cause abnormal cytokinesis and multinucleation through dysfunction of the centralspindlin complex and microfilaments

BackgroundWith the large-scale production and application of amorphous silica nanoparticles (aSiNPs), its adverse health effects are more worthy of our attention. Our previous research has demonstrated for the first time that aSiNPs induced cytokinesis failure, which resulted in abnormally high incidences of multinucleation in vitro, but the underlying mechanisms remain unclear. Therefore, the purpose of this study was firstly to explore whether aSiNPs induced multinucleation in vivo, and secondly to investigate the underlying mechanism of how aSiNPs caused abnormal cytokinesis and multinucleation.MethodsMale ICR mice with intratracheal instillation of aSiNPs were used as an experimental model in vivo. Human hepatic cell line (L-02) was introduced for further mechanism study in vitro.ResultsIn vivo, histopathological results showed that the rate of multinucleation was significantly increased in the liver and lung tissue after aSiNPs treatment. In vitro, immunofluorescence results manifested that aSiNPs directly caused microfilaments aggregation. Following mechanism studies indicated that aSiNPs increased ROS levels. The accumulation of ROS further inhibited the PI3k 110β/Aurora B pathway, leading to a decrease in the expression of centralspindlin subunits MKLP1 and CYK4 as well as downstream cytokines regulation related proteins Ect2, Cep55, CHMP2A and RhoA. Meanwhile, the particles caused abnormal co-localization of the key mitotic regulatory kinase Aurora B and the centralspindlin complex by inhibiting the PI3k 110β/Aurora B pathway. PI3K activator IGF increased the phosphorylation level of Aurora B and improved the relative ratio of the centralspindlin cluster. And ROS inhibitors NAC reduced the ratio of multinucleation, alleviated the PI3k 110β/Aurora B pathway inhibition, and then increased the expression of MKLP1, CYK4 and cytokinesis-related proteins, whilst NAC restored the clustering of the centralspindlin.ConclusionThis study demonstrated that aSiNPs led to multinucleation formation both in vivo and in vitro. ASiNPs exposure caused microfilaments aggregation and inhibited the PI3k 110β/Aurora B pathway through excessive ROS, which then hindered the centralspindlin cluster as well as restrained the expression of centralspindlin subunits and cytokinesis-related proteins, which ultimately resulted in cytokinesis failure and the formation of multinucleation.

Aurora A polyubiquitinates the BRCA1-interacting protein OLA1 to promote centrosome maturation

The BRCA1-interacting protein Obg-like ATPase 1 (OLA1) functions in centriole duplication. In this study, we show the role of the mitotic kinase Aurora A in the reduction of centrosomal OLA1. Aurora A binds to and polyubiquitinates OLA1, targeting it for proteasomal degradation. NIMA-related kinase 2 (NEK2) phosphorylates the T124 residue of OLA1, increases binding of OLA1 to Aurora A and OLA1 polyubiquitination by Aurora A, and reduces centrosomal OLA1 in G2 phase. The kinase activity of Aurora A suppresses OLA1 polyubiquitination. The decrease in centrosomal OLA1 caused by Aurora A-mediated polyubiquitination promotes the recruitment of pericentriolar material proteins in G2 phase. The E3 ligase activity of Aurora A is critical for centrosome amplification induced by its overexpression. The results suggest a dual function of Aurora A as an E3 ubiquitin ligase and a kinase in the regulation of centrosomal OLA1, which is essential for proper centrosome maturation in G2 phase.

Phospho PTEN mediated dephosphorylation of mitotic kinase PLK1 and Aurora Kinase A prevents aneuploidy and preserves genomic stability

PTEN, dual phosphatase tumor suppressor protein, is found to be frequently mutated in various cancers. Post-translational modification of PTEN is important for its sub-cellular localization and catalytic functions. But how these modifications affect cytological damage and aneuploidy is not studied in detail. We focus on the role of phosphatase activity along with C-terminal phosphorylation of PTEN in perspective of cytological damage like micronucleus, nuclear bud, and nuclear bridge formation. Our data suggest that wild-type PTEN, but not phospho-mutant PTEN significantly reduces cytological damage in PTEN null PC3 cells. In case of phosphatase-dead PTEN, cytological damage markers are increased during 24h recovery after DNA damage. When we use phosphorylation and phosphatase-dead dual mutant PTEN, the extent of different cytological DNA damage parameters are similar to phosphatase-dead PTEN. We also find that both of those activities are essential for maintaining chromosome numbers. PTEN null cells exhibit significantly aberrant γ-tubulin pole formation during metaphase. Interestingly, we observed that p-PTEN localized to spindle poles along with PLK1 and Aurora Kinase A. Further depletion of phosphorylation and phosphatase activity of PTEN increases the expression of p-Aurora Kinase A (T288) and p-PLK1 (T210), compared to cells expressing wild-type PTEN. Again, wild-type PTEN but not phosphorylation-dead mutant is able to physically interact with PLK1 and Aurora Kinase A. Thus, our study suggests that the phosphorylation-dependent interaction of PTEN with PLK1 and Aurora Kinase A causes dephosphorylation of those mitotic kinases and by lowering their hyperphosphorylation status, PTEN prevents aberrant chromosome segregation in metaphase.

PP6 regulation of Aurora A-TPX2 limits NDC80 phosphorylation and mitotic spindle size.

Amplification of the mitotic kinase Aurora A or loss of its regulator protein phosphatase 6 (PP6) have emerged as drivers of genome instability. Cells lacking PPP6C, the catalytic subunit of PP6, have amplified Aurora A activity, and as we show here, enlarged mitotic spindles which fail to hold chromosomes tightly together in anaphase, causing defective nuclear structure. Using functional genomics to shed light on the processes underpinning these changes, we discover synthetic lethality between PPP6C and the kinetochore protein NDC80. We find that NDC80 is phosphorylated on multiple N-terminal sites during spindle formation by Aurora A-TPX2, exclusively at checkpoint-silenced, microtubule-attached kinetochores. NDC80 phosphorylation persists until spindle disassembly in telophase, is increased in PPP6C knockout cells, and is Aurora B-independent. An Aurora-phosphorylation-deficient NDC80-9A mutant reduces spindle size and suppresses defective nuclear structure in PPP6C knockout cells. In regulating NDC80 phosphorylation by Aurora A-TPX2, PP6 plays an important role in mitotic spindle formation and size control and thus the fidelity of cell division.

BCL-xL inhibition potentiates cancer therapies by redirecting the outcome of p53 activation from senescence to apoptosis.

Cancer therapies trigger diverse cellular responses, ranging from apoptotic death to acquisition of persistent therapy-refractory states such as senescence. Tipping the balance toward apoptosis could improve treatment outcomes regardless of therapeutic agent or malignancy. We find that inhibition of the mitochondrial protein BCL-xL increases the propensity of cancer cells to die after treatment with a broad array of oncology drugs, including mitotic inhibitors and chemotherapy. Functional precision oncology and omics analyses suggest that BCL-xL inhibition redirects the outcome of p53 transcriptional response from senescence to apoptosis, which likely occurs via caspase-dependent down-modulation of p21 and downstream cytostatic proteins. Consequently, addition of a BCL-2/xL inhibitor strongly improves melanoma response to the senescence-inducing drug targeting mitotic kinase Aurora kinase A (AURKA) in mice and patient-derived organoids. This study shows a crosstalk between the mitochondrial apoptotic pathway and cell cycle regulation that can be targeted to augment therapeutic efficacy in cancers with wild-type p53.

Revisiting degron motifs in human AURKA required for its targeting by APC/CFZR1.

Mitotic kinase Aurora A (AURKA) diverges from other kinases in its multiple active conformations that may explain its interphase roles and the limited efficacy of drugs targeting the kinase pocket. Regulation of AURKA activity by the cell is critically dependent on destruction mediated by the anaphase-promoting complex (APC/CFZR1) during mitotic exit and G1 phase and requires an atypical N-terminal degron in AURKA called the "A-box" in addition to a reported canonical D-box degron in the C-terminus. Here, we find that the reported C-terminal D-box of AURKA does not act as a degron and instead mediates essential structural features of the protein. In living cells, the N-terminal intrinsically disordered region of AURKA containing the A-box is sufficient to confer FZR1-dependent mitotic degradation. Both in silico and in cellulo assays predict the QRVL short linear interacting motif of the A-box to be a phospho-regulated D-box. We propose that degradation of full-length AURKA also depends on an intact C-terminal domain because of critical conformational parameters permissive for both activity and mitotic degradation of AURKA.

Discovery and Optimization of Seven-Membered Lactam-Based Compounds to Phenocopy the Inhibition of the Aurora Kinase B.

We used mechanism-informed phenotypic screening to identify and optimize compounds that phenocopy the genetic depletion of the mitotic aurora kinase B (AURKB) kinase. After assaying nine aryl fused seven-membered lactam compounds, we identified a hit compound 6a that was subsequently optimized to five lead compounds with low nanomolar activity, represented by the lead compound 6v (19 nM). With excellent drug-like properties, these compounds reproduced the loss of function in phenotypes of AURKB and exhibited potent cytotoxic activities in various cancer cell lines. Collectively, these data support that seven-membered lactam-based analogs might be valuable for further development as a new type of antimitotic agents for the treatment of cancer.

Modulation of oxidative phosphorylation augments antineoplastic activity of mitotic aurora kinase inhibition

Uncontrolled mitosis is one of the most important features of cancer, and mitotic kinases are thought to be ideal targets for anticancer therapeutics. However, despite numerous clinical attempts spanning decades, clinical trials for mitotic kinase-targeting agents have generally stalled in the late stages due to limited therapeutic effectiveness. Alisertib (MLN8237) is a promising oral mitotic aurora kinase A (AURKA, Aurora-A) selective inhibitor, which is currently under several clinical evaluations but has failed in its first Phase III trial due to inadequate efficacy. In this study, we performed genome-wide CRISPR/Cas9-based screening to identify vulnerable biological processes associated with alisertib in breast cancer MDA-MB-231 cells. The result indicated that alisertib treated cancer cells are more sensitive to the genetic perturbation of oxidative phosphorylation (OXPHOS). Mechanistic investigation indicated that alisertib treatment, as well as other mitotic kinase inhibitors, rapidly reduces the intracellular ATP level to generate a status that is highly addictive to OXPHOS. Furthermore, the combinational inhibition of mitotic kinase and OXPHOS by alisertib, and metformin respectively, generates severe energy exhaustion in mitotic cells that consequently triggers cell death. The combination regimen also enhanced tumor regression significantly in vivo. This suggests that targeting OXPHOS by metformin is a potential strategy for promoting the therapeutic effects of mitotic kinase inhibitors through the joint targeting of mitosis and cellular energy homeostasis.

Bub1 and CENP-U redundantly recruit Plk1 to stabilize kinetochore-microtubule attachments and ensure accurate chromosome segregation.

Bub1 is required for the kinetochore/centromere localization of two essential mitotic kinases Plk1 and Aurora B. Surprisingly, stable depletion of Bub1 by ∼95% in human cells marginally affects whole chromosome segregation fidelity. We show that CENP-U, which is recruited to kinetochores by the CENP-P and CENP-Q subunits of the CENP-O complex, is required to prevent chromosome mis-segregation in Bub1-depleted cells. Mechanistically, Bub1 and CENP-U redundantly recruit Plk1 to kinetochores to stabilize kinetochore-microtubule attachments, thereby ensuring accurate chromosome segregation. Furthermore, unlike its budding yeast homolog, the CENP-O complex does not regulate centromeric localization of Aurora B. Consistently, depletion of Bub1 or CENP-U sensitizes cells to the inhibition of Plk1 but not Aurora B kinase activity. Taken together, our findings provide mechanistic insight into the regulation of kinetochore function, which may have implications for targeted treatment of cancer cells with mutations perturbing kinetochore recruitment of Plk1 by Bub1 or the CENP-O complex.

Suv420 enrichment at the centromere limits Aurora B localization and function.

Centromere structure and function are defined by the epigenetic modification of histones at centromeric and pericentromeric chromatin. The constitutive heterochromatin found at pericentromeric regions is highly enriched for H3K9me3 and H4K20me3. Although mis-expression of the methyltransferase enzymes that regulate these marks, Suv39 and Suv420, is common in disease, the consequences of such changes are not well understood. Our data show that increased centromere localization of Suv39 and Suv420 suppresses centromere transcription and compromises localization of the mitotic kinase Aurora B, decreasing microtubule dynamics and compromising chromosome alignment and segregation. We find that inhibition of Suv420 methyltransferase activity partially restores Aurora B localization to centromeres and that restoration of the Aurora B-containing chromosomal passenger complex to the centromere is sufficient to suppress mitotic errors that result when Suv420 and H4K20me3 is enriched at centromeres. Consistent with a role for Suv39 and Suv420 in negatively regulating Aurora B, high expression of these enzymes corresponds with increased sensitivity to Aurora kinase inhibition in human cancer cells, suggesting that increased H3K9 and H4K20 methylation may be an underappreciated source of chromosome mis-segregation in cancer. This article has an associated First Person interview with the first author of the paper.

Aurora B regulates PP1γ–Repo-Man interactions to maintain the chromosome condensation state

The mitotic kinase Aurora B regulates the condensation of chromatin into chromosomes by phosphorylating chromatin proteins during early mitosis, whereas the phosphatase PP1γ performs the opposite function. The roles of Aurora B and PP1γ must be tightly coordinated to maintain chromosomes at a high phosphorylation state, but the precise mechanisms regulating their function remain largely unclear. Here, mainly through immunofluorescence microscopy and co-immunoprecipitation assays, we find that dissociation of PP1γ from chromosomes is essential for maintaining chromosome phosphorylation. We uncover that PP1γ is recruited to mitotic chromosomes by its regulatory subunit Repo-Man in the absence of Aurora B activity and that Aurora B regulates dissociation of PP1γ by phosphorylating and disrupting PP1γ-Repo-Man interactions on chromatin. Overexpression of Repo-Man mutants that cannot be phosphorylated or inhibition of Aurora B kinase activity resulted in the retention of PP1γ on chromatin and prolonged the chromatin condensation process; a similar outcome was caused by the ectopic targeting of PP1γ to chromatin. Together, our findings reveal a novel regulation mechanism of chromatin condensation in which Aurora B counteracts PP1γ activity by releasing PP1γ from Repo-Man and may have important implications for understanding the regulations of dynamic structural changes of the chromosomes in mitosis.

Abstract 3011: Phase I study of TT-00420, a multiple kinase inhibitor, in patients with triple negative breast cancers and other advanced solid tumors

Abstract Background: TT-00420, a multikinase inhibitor, targets cell proliferation, angiogenesis, and immunomodulatory pathways by inhibiting the mitotic kinases Aurora A/B, Janus kinases (JAK) involved in cytokine signalling, and receptor tyrosine kinases involved in angiogenesis (FGFRs and VEGFRs). These pathways are important in the pathogenesis of triple negative breast cancer (TNBC) and TT-00420 has shown preclinical efficacy against multiple subtypes of TNBC. Methods: This phase I, open-label, first-in-human study is enrolling adult patients with advanced or metastatic TNBC and other advanced solid tumors. The study contains a dose escalation phase (N=22), followed by dose expansion in two parallel cohorts, a TNBC cohort (N=22) and a selected advanced tumor (SAT) cohort (N=22). The provisional dose range for dose escalation is 1 mg/d to 20 mg/d. Dose escalation is guided by Bayesian modeling with overdose control until a dose recommended for dose expansion is determined. Adverse events (AE) are evaluated per CTCAE v5.0 criteria, and tumor response is evaluated per RECIST v1.1 criteria. Patients receive a single daily oral administration of TT-00420 continuously for 28-day cycles. The primary endpoint is to evaluate dose limiting toxicity (DLT) and identify a maximum tolerated dose (MTD). Secondary objectives include evaluating the efficacy and pharmacokinetic profile of TT-00420. Results: As of the data cut-off date on September 26, 2019, 11 patients have received TT-00420 treatment in 4 dose levels, 1 mg/d (N=1), 3 mg/d (N=1), 5 mg/d (N=3), and 8 mg/d (N=6). No DLTs have been observed at any of the tested dose levels. Commonly reported suspected AEs across all tested dose levels were grades 1 or 2 diarrhea (n=4, 36.4%), myalgia (n=2, 18.2%) and hypertension (n=2, 18.2%). Two suspected serious adverse events reported were grade 2 hypertension (8 mg/d) and grade 3 nausea (5 mg/d). No grade 3 AEs have occurred in more than one subject. Stable disease has been observed in 4 patients, including a metastatic TNBC patient treated at 8 mg/d, who received 9 lines of prior antineoplastic therapy. Per RECIST v1.1 criteria, this TNBC patient had a 21% decrease in target lesions observed on restaging after two cycles of TT-00420 treatment. Interestingly, significant clinical improvement in the skin lesions were observed as well. More detailed data from this TNBC patient and other patients treated in the study will be presented. Enrollment in dose escalation is currently ongoing. Clinical trial information: NCT03654547 Citation Format: Sarina A. Piha-Paul, Ecaterina Ileana-Dumbrava, Filip Janku, Daniel D. Karp, Funda Meric-Bernstam, Jordi Rodon, Brenda Ngo, Peng Peng, Frank Wu. Phase I study of TT-00420, a multiple kinase inhibitor, in patients with triple negative breast cancers and other advanced solid tumors [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3011.

Abstract B20: Patient-derived organoids demonstrate synergistic effect of co-targeting Aurora kinase and prosurvival BCL2 family proteins

Abstract Many melanomas are resistant to standard-of-care immune checkpoint blockade and inhibitors of mutated BRAF. New effective therapies that work via immune and BRAF-independent mechanisms are urgently needed. We have previously demonstrated that an inhibitor of mitotic kinase Aurora A (AURKAi) can induce a distinct type of stable cell cycle known as senescence in melanoma tumors independent of their genetic background. Here we hypothesized that combination of AURKAi with a drug that selectively kills senescent cells, such as BCL2 inhibitor (BCLi) navitoclax, will be synthetically lethal to melanoma cells. Melanoma patient-derived organoids (PDOs) were used to test efficacy of AURKAi and BCLi combination treatment. To generate organoids, patient-derived tumor cells were cultured ex vivo in 3D matrix. Drug response was evaluated using fluorescent viability assay. Fifteen out of twenty PDOs were highly sensitive to AURKAi and BCLi combination treatment, exhibiting robust induction of cell death. No cell death was observed when either drug was added individually. NextGen DNA sequencing indicated that all sensitive PDOs retained wild-type TP53, while resistant tumors had mutations in this gene. Knockout or knockdown of TP53 abrogated AURKAi and BCLi-induced cell death in melanoma cells, suggesting that transcription factor p53 encoded by TP53 is a key mediator of drug response. Mechanistically, p53 was induced by AURKAi treatment independent of BCLi. However, single-agent AURKAi treatment resulted in activation of p53-mediated cell cycle arrest based on increased expression of CDK inhibitor p21 and decreased expression of proliferation markers. In contrast, addition of BCLi to AURKAi activated proapoptotic p53 target BAX and induced cleavage of PARP and caspases 3, 7, and 8. Treatment with pan-caspase inhibitor abrogated cell death, suggesting caspase-dependent apoptosis. Electron microscopy revealed morphologic signs of apoptosis in combination-treated cells, including cell blebbing, apoptotic bodies, and nuclear fragmentation. This suggests that BCLi shifts cell fate decision in AURKAi-treated cells towards apoptosis. In conclusion, combination treatment with AURKAi and BCl2i induced p53-dependent apoptosis. Preclinical evaluation of this drug combination in PDO model demonstrated robust efficacy against melanomas with wild-type p53. Since p53 mutations are relatively rare in melanoma, a combination of AURKAi and BCLi presents a promising therapeutic option for this deadly disease. Citation Format: Vijaya Bharti, Ashlyn Blevins, Vivian L. Weiss, Sheau C Chen, Roman Uzhachenko, Ann Richmond, Anna E. Vilgelm. Patient-derived organoids demonstrate synergistic effect of co-targeting Aurora kinase and prosurvival BCL2 family proteins [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr B20.

Aurora B Kinase Promotes CHIP-Dependent Degradation of HIF1α in Prostate Cancer Cells.

Hypoxia is a major factor in tumor progression and resistance to therapies, which involves elevated levels of the transcription factor HIF1α. Here, we report that prostate tumor xenografts express high levels of HIF1α and show greatly enhanced growth in response to knockdown of the E3 ligase CHIP (C-terminus of Hsp70-interacting protein). In multiple human prostate cancer cell lines under hypoxia, taxol treatment induces the degradation of HIF1α, and this response is abrogated by knockdown of CHIP, but not by E3 ligase VHL or RACK1. HIF1α degradation is accompanied by loss of function, evidenced by reduced expression of HIF1α-dependent genes. CHIP-dependent HIF1α degradation also occurs in cells arrested in mitosis by nocodazole instead of taxol. Mitotic kinase Aurora B activity is required for taxol-induced HIF1α degradation. Purified Aurora B directly phosphorylates HIF1α at multiple sites, and these modifications enhance its polyubiquitination by CHIP in a purified reconstituted system. Our results show how activation of Aurora B promotes CHIP-dependent degradation of HIF1α in prostate cancer cells. This new knowledge may affect the use of mitotic kinase inhibitors and open new approaches for treatment of hypoxic prostate tumors.