Review : Aurora Kinase A (AURKA), Potential Target Therapies for Cancer Control and its Role in Oncogenicity

Cancer, a leading reason for mortality worldwide, arises due to irregularities in genetic and epigenetic modulation that disrupt the coordinated events of the cell cycle, resulting in dysregulated protein kinase expression and functionality. Aurora kinase A (AURKA), a crucial member of specific serine/threonine kinase family, holds significant importance in regulating maturation and stability in centrosome construction, microtubular stability, and the sequential regulation of chromosome segregation during the cell cycle. Dysregulation or overexpression of AURKA disrupts the delicate balance of the cell cycle, leading to the emergence of various cancers. The aberrant expression of AURKA and its downstream molecules plays a pivotal role in the pathogenesis of cancer. Furthermore, dysfunctions of AURKA are closely associated with therapeutic drug resistance observed in pancreatic cancer, breast cancer, and ovarian cancer. Experimental data strongly supports AURKA as a promising target for anticancer interventions. Modulation of its activity or suppression of its expression effectively hampers the progression of numerous cancer types. Hence, the employment of single AURKA inhibitors or their combination with other anti-cancer therapies has demonstrated remarkable outcomes in cancer prevention. This report aims to deliver the multifaceted role of AURKA, highlighting its contribution to oncogenicity, and discussing potential target therapies that have been clinically implemented to regulate its activity and impede tumor progression.

Introduction

The cell cycle is a highly regulated process crucial for cellular growth, division, and maintenance of genomic integrity. Among the various kinases involved in cell cycle control, Aurora kinase A (AURKA) emerges as a key player, regulating critical cellular events such as centrosome maturation, microtubule dynamics, and the precise segregation of chromosomes. This review provides an overview of AURKA's role in cell cycle regulation and highlights its significance in maintaining cellular homeostasis. The dysregulation of AURKA expression is frequently observed in numerous cancers, leading to abnormal cell cycle progression and oncogenic transformation. The aberrant activation of AURKA leads to disrupted cell cycle progression, contributing significantly to the development and progression of diverse cancer types. It elucidates the mechanisms underlying AURKA overexpression and its association with cancer development.

AURKA as a Prognostic Indicator
Clinical studies have revealed the prognostic value of AURKA expression in different cancer types. In this subsection we discuss the correlation between AURKA expression levels and patient prognosis, emphasizing its potential as a diagnostic and prognostic biomarker regarding cancer pathogenesis. The development of specific inhibitors targeting AURKA has opened new avenues for cancer treatment. In this essay we tried to emphasize an overview of the different classes of AURKA inhibitors and their mechanisms of action, highlighting their potential for therapeutic intervention. Combinatorial approaches, employing AURKA inhibitors in conjunction with other anti-cancer therapies, have shown promising results in preclinical and clinical settings. This review explores various combination strategies and their efficacy in combating cancer progression. It also provides an update on ongoing clinical trials investigating the efficacy and safety of AURKA-targeted therapies in cancer patients. It discusses the current status of these trials, their design, and potential implications for future treatment strategies. The rapid advancement of scientific knowledge has uncovered new insights into the intricate mechanisms involving AURKA and its downstream signaling pathways. It also highlights emerging research areas that hold promise for further understanding of AURKA's role in oncogenicity and the development of novel therapeutic approaches.

Aurora Kinase A and its Role in Cell Cycle Regulation

AURKA plays multiple critical roles during the cell cycle, particularly in the G2/M transition. Its activation orchestrates vital activities such as chromosomal alignment, centrosome maturation, and separation, spindle assembly checkpoint maintenance, nucleation of microtubule, bipolar spindle formation, cytoplasm division, and exit from the division stage. Autophosphorylation of AURKA at Thr288 initiates a cascade, activating polo-like kinase 1 (PLK1), which further triggers the activation of the PLK1-Cyclin B-CDK complex. The AURKA/PLK1 axis plays a pivotal role in regulating centrosome maturation, ensuring precise cell division.

Consequences of Aurora Kinase-A Dysregulation

Experimental evidence highlights the pivotal role of dysregulated AURKA expression in driving normal cells toward tumorigenic states. Abnormal AURKA expression has been observed across various solid malignancies, including neuroblastoma, colorectal, breast, ovarian, cervical, and prostate cancers. AURKA amplification leads to non-disjunction, resulting in aneuploidy, excessive centrosome numbers, abnormal centrosome development, and the formation of multipolar spindles. As a key regulator of the G2/M transition, aberrant AURKA expression disrupts cell cycle checkpoints, impairing the faithful progression of cells.

Furthermore, AURKA dysregulation influences downstream molecules, such as FOXK1, which is activated by Aurora-A through phosphorylation of the 37th serine residue of SOX8. This activation impacts the glycolytic pathway, leading to chemoresistance. Inhibition of Aurora-A in ovarian cancer cells has demonstrated reduced cell viability, as Aurora-A promotes glycolytic metabolism and delays cellular senescence. Additionally, dysregulated AURKA affects glycolysis by phosphorylating PKM2 at threonine 45, redirecting glycolytic intermediates toward alternative biosynthetic pathways.

AURKA Dysregulation in Cancer Pathogenesis

The dysregulated expression of AURKA contributes to various aspects of cancer pathogenesis. Notably, its overexpression in pancreatic ductal adenocarcinoma (PDAC) is associated with enhanced epithelial-to-mesenchymal transition (EMT), facilitating tumor metastasis. Inhibition of AURKA using small interfering RNA shows promise in reducing tumorigenicity and improving recurrence rates in PDAC cells. Moreover, AURKA overexpression in colorectal cancer correlates with upstream signaling molecule upregulation, such as PLK1, and downregulation of microtubule-associated protein 9 (MAP9).

In gastric cancer, AURKA and kinesin family member C1 (KIFC1) exhibit a clear association, with elevated levels of both proteins linked to tumor characteristics, including node involvement, distant metastasis, tumor size, and poor patient survival rates. Furthermore, AURKA overexpression in upper gastrointestinal adenocarcinomas triggers STAT3 phosphorylation and subsequent nuclear translocation, promoting aggressive metastasis. Increased β-Catenin levels and activation of GSK-3 and AKT, mediated by elevated AURKA expression, promote gastric cancer cell proliferation, metastasis, and migration.

AURKA dysregulation also influences hepatocellular carcinoma, glioblastoma stem cells, and breast cancer. In hepatocellular carcinoma, AURKA overexpression induces epithelial-to-mesenchymal transition and confers resistance to radiation therapy. AURKA phosphorylates BRCA1, impairing its ability to prevent centrosome re-duplication and increasing the risk of breast cancer. Additionally, AURKA deregulation in gastric cancer leads to the repression of downstream targets, including PUMA and p21/WAF1, thereby preventing apoptosis and promoting cell survival.

Exploring the Involvement of Aurora Kinase A (AURKA) in Necroptosis

Necroptosis, a programmed form of cell death, has emerged as a potential avenue for targeted cancer therapy. Recent studies have uncovered a possible association between necroptosis regulation and key pathways involving Aurora Kinase A (AURKA), glycogen synthase kinase 3 (GSK3), and Wnt/β-catenin. Specifically, the compound CCT137690 has been shown to induce necroptosis in established K-Ras-driven pancreatic ductal adenocarcinoma (PDAC), leading to increased production of immune cell death mediators, such as high mobility group box 1 (HMGB1) and adenosine triphosphate (ATP). Notably, AURKA and GSK3 have also been identified as predictive markers for poor outcomes in pancreatic cancer patients. These findings shed light on the intricate roles of necroptosis-associated genes and proteins in both early oncogenesis, with potential implications for cancer prevention, and later stages of tumor growth, paving the way for the development of targeted therapies.

Recent research suggests a potential interplay between necroptosis regulation and key signaling pathways involving AURKA, GSK3, and Wnt/β-catenin. Understanding the involvement of AURKA in necroptosis could provide valuable insights into the mechanisms underlying pancreatic carcinoma development and offer new opportunities for targeted interventions.

Necroptosis Induction in Pancreatic Carcinoma

The compound CCT137690 has shown promising results in inducing necroptosis specifically in established K-Ras-driven PDAC. This induction of necroptosis is accompanied by an upregulation of immune cell death mediators, including HMGB1 and ATP. The activation of necroptosis in pancreatic carcinoma holds potential therapeutic implications, as it enhances antitumor T cell immunity, which is crucial for effective cancer control.

AURKA and GSK3 as Predictive Markers

In addition to their roles in cell cycle regulation and oncogenesis, AURKA and GSK3 have emerged as predictive markers for poor outcomes in pancreatic cancer patients. Their association with necroptosis suggests their involvement in the complex interplay between cell death pathways and cancer progression. Further investigation into the mechanistic links between AURKA, GSK3, and necroptosis may unravel novel therapeutic targets and strategies for combating pancreatic carcinoma.

Implications for Cancer Prevention and Targeted Therapies

The identification of necroptosis-related genes and proteins, including AURKA and GSK3, in both early oncogenesis and later stages of tumor growth underscores their multifaceted roles in cancer pathogenesis. Harnessing the potential of necroptosis induction as a therapeutic strategy, particularly in pancreatic carcinoma, holds promise for enhancing treatment outcomes. Developing targeted therapies that exploit the interplay between necroptosis regulation and key signaling pathways may offer novel approaches to combat this devastating disease.

The investigation into the involvement of AURKA in necroptosis provides valuable insights into the intricate connections between cell death pathways and pancreatic carcinoma. The induction of necroptosis and the predictive value of AURKA and GSK3 in pancreatic cancer patients highlight the potential of targeting these pathways for improved therapeutic outcomes. Further research is warranted to elucidate the underlying mechanisms and identify specific therapeutic interventions that exploit necroptosis regulation in the context of pancreatic carcinoma. Such advancements have the potential to revolutionize cancer treatment and pave the way for more effective personalized therapies.

Targeting Aurora Kinase A Overexpression for Cancer Therapy: Recent Advances and Promising Strategies

This bibliographic report provides a comprehensive review of the role of AURKA in cancer pathogenesis and highlights recent developments in targeting AURKA for effective therapeutic interventions. The utilization of AURKA inhibitors, particularly MLN8237 (Alisertib), has shown promising results in controlling AURKA overexpression and restraining cancer growth, specifically in pancreatic cancer. Studies have revealed that MLN8237 significantly impedes the proliferation and migration of pancreatic ductal adenocarcinoma (PDAC) cells derived from human patients. Furthermore, MLN8237 induces senescence and programmed cell death in pancreatic cancer cells, aligning with the known oncogenic functions of AURKA.

Combination strategies incorporating AURKA inhibitors have demonstrated synergistic effects in enhancing tumor suppression. The combination of AURKA inhibitors with H3K9 methyltransferase inhibitors, taxanes, RNA interference targeting Aurora kinase, or BCL2 protein inhibitors has exhibited enhanced efficacy in inducing tumor cell death. MLN8237 competes with Aurora Kinase A to bind with ATP binding sites and effectively inhibits Aurora-A, thereby maintaining proper spindle polarity in cells. Overall, MLN8237 (Alisertib) exhibits potent anti-proliferative, anti-metastatic, and pro-apoptotic properties in pancreatic cancer cells, suggesting its potential as a chemotherapeutic agent for PDAC.

Administration of MLN8237
MLN8237has also shown promising outcomes in the treatment of recurrent/refractory atypical teratoid rhabdoid tumors (ATRT). In a trial involving four ATRT patients, oral administration of MLN8237 for seven consecutive days resulted in disease stabilization and subsequent decline after three treatment cycles. Notably, the combination of siRNA and MLN8237 has demonstrated intriguing effects on neuroblastoma cells, inducing apoptosis in senescent neuroblastoma cells through the targeting of the AKT/STAT3 signaling pathway. These findings indicate the therapeutic potential of MLN8237 for ATRT and neuroblastoma treatment.

Colorectal cancer (CRC) has been associated with dysregulated expression of both Haspin and Aurora A kinase. Clinical trials exploring dual inhibition of Aurora A and Haspin kinases, utilizing MLN8237 (AURKA inhibitor) and CHR6494 (Haspin kinase inhibitor), have shown promise in inducing programmed cell death, inhibiting tumor proliferation, and disrupting critical pathways involved in CRC metastasis. Overexpression of Haspin in human CRC cells results in elevated levels of survivin, a protein associated with cell survival. Conversely, inhibition of Haspin gene expression reduces survivin levels, leading to decreased cell survival. Additionally, the combination of MLN8237 and CHR6494 reduces survivin protein levels, enhances p53 production, and promotes apoptosis via the p53-Bax-caspase-3 pathway, ultimately preventing proliferation and carcinogenesis in CRC cells by inducing mitotic arrest and mitotic catastrophe.

AKI603 Inhibits Breast Cancer Stem Cells
Furthermore, AKI603, an AURKA inhibitor, when used in conjunction with Thiostrepton, a FOXM1 inhibitor, synergistically decreases the self-renewal capacity of breast cancer stem cells (CSCs) and overcomes therapeutic resistance. This synergistic effect is attributed to the disruption of the positive feedback loop between AURKA and FOXM1. Additionally, palmatine alkaloid, an isoquinoline alkaloid derived from a common plant, exhibits positive AURKA inhibitory properties and demonstrates the ability to induce G2/M arrest and apoptosis in colon cancer cells by blocking AURKA.

Natural compound gossypin has been identified as an inhibitor of AURKA expression and has exhibited anticancer effects on gastric cancer cells. Gossypin treatment effectively inhibits cell proliferation and migration through its inhibition of AURKA. This inhibition leads to the activation of caspases, cleavage of PARP, increased expression of CYT-C, reduced expression of BCL-XL, and subsequent induction of apoptosis.

Finally, the efficacy of MLN8237 and Adavosertib in suppressing head and neck squamous cell cancer (HNSCC) in patients without human papillomavirus infection has been established. By targeting critical proteins involved in DNA damage response and cell cycle control, these inhibitors effectively inhibit cell growth and enhance treatment responsiveness in HNSCC patients.

So, targeting AURKA overexpression represents a promising therapeutic strategy for cancer treatment. AURKA inhibitors, including both synthetic and natural compounds, have demonstrated efficacy in inhibiting tumor growth, and metastasis and overcoming therapeutic resistance in various cancer types. Further research and clinical trials are necessary to optimize AURKA-targeted therapies and improve patient outcomes.
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Experimental Tools and Techniques
To check Aurora kinase A inhibitor MLN8237 which suppresses pancreatic cancer growth, the researchers employed various techniques and tools to investigate the role of Aurora kinase in pancreatic cancer and evaluate the efficacy of the specific AURKA inhibitor MLN8237.

1. Cell Culture and Transwell Assay: Human primary pancreatic stellate cells (HPSCs) and different pancreatic cancer cell lines (BxPC-3, ASPC-1, PANC-1, Mia Paca-2) obtained from American Type Culture Collection (ATCC) were cultured under recommended conditions. To monitor migration of cells, Transwell assays were executed.

2. Western Blotting: The researchers conducted Western blotting analysis to examine protein expression levels. Antibodies used in these experiments included Aurora kinase-A, phosphorylated Aurora kinase-A, DEP-1, cleaved poly(ADP-ribose) polymerase (c-PARP), Caspase-3, and GAPDH.

3. Immunohistochemistry Staining: Normal human and pancreatic cancer tissues were acquired from resection specimens. Immunohistochemistry staining was performed using antibodies against Aurora kinase-A, alpha-Smooth Muscle Actin (aSMA), and cleaved Caspase 3.

4. BrdU Incorporation and Cell Count: BrdU incorporation kit was used to assess cell proliferation, and cell numbers were determined using the Cell Counting Kit 8 (WST-8).

5. Mouse Orthotopic and Genetic Pancreatic Cancer Models: Male nude mice were used to establish orthotopic pancreatic cancer models by injecting MIA PaCa-2 cells into the pancreas tail. Genetic pancreatic cancer models were developed using p53/LSL/Pdx-Cre mice. MLN8237 and Gemcitabine hydrochloride were administered via intraperitoneal injection. Saline was used as a control.

6. Statistical Analysis: Statistical analysis was performed using GraphPad Prism 5 software. Values were expressed as mean ± SEM, and statistical significance was determined using unpaired two-tailed Student's t-test and one-way ANOVA followed by the Newman-Keuls test.

So, they actually utilized these techniques and tools to investigate the expression of Aurora kinase-A in pancreatic ductal adenocarcinoma (PDAC) and evaluate the efficacy of MLN8237 in inhibiting proliferation, migration, and tumor burden in both in vitro cell line models and in vivo mouse models of pancreatic cancer.

Now, To check the Co-inhibition of Aurora A and Haspin kinases, which enhances survivin blockage and p53 induction for mitotic catastrophe and apoptosis in human colorectal cancer, following techniques were used.

1. Cell Culture and Treatment: Colorectal cancer (CRC) cell lines were cultured and treated with inhibitors targeting Aurora A kinase (AURKA) and Haspin kinase (Haspin). The specific inhibitors used were MLN8237 (an AURKA inhibitor) and CHR6494 (a Haspin kinase inhibitor).

2. Cell Viability and Proliferation Assays: Cell viability and proliferation were assessed using various assays such as the MTT assay, colony formation assay, and EdU incorporation assay. These assays helped evaluate the effects of the inhibitors on cell growth and survival.

3. Flow Cytometry Analysis: Flow cytometry was used to analyze cell cycle distribution and apoptotic cell death. DNA content staining with propidium iodide and Annexin V staining were performed to determine cell cycle progression and apoptosis induction, respectively.

4. Immunoblotting: Immunoblotting analysis was conducted to examine protein expression levels. Antibodies against specific proteins of interest were used, including survivin, p53, Bcl-2 family proteins (Bcl-2, Bcl-xL, Bax), caspase-3, and phosphorylated histone H3.

5. Immunofluorescence Staining: Immunofluorescence staining was performed to visualize the localization and expression of specific proteins in CRC cells. Antibodies targeting survivin and p53 were used for immunofluorescence staining.

6. Gene Silencing: RNA interference (RNAi) techniques were employed to silence the expression of AURKA, Haspin, and p53 genes in CRC cells. Small interfering RNA (siRNA) molecules targeting these genes were transfected into cells to assess the impact on cell survival and apoptosis.

7. Statistical Analysis: Statistical analysis was carried out using appropriate methods to determine the significance of the observed results.

8. Annexin V-FITC staining: It was performed to identify and quantify apoptotic cells. FITC-conjugated Annexin V, a protein that has a high affinity for phosphatidylserine, was used to label exposed phosphatidylserine on the outer membrane of apoptotic cells. This allows the distinction between viable cells, early apoptotic cells (phosphatidylserine externalization), and late apoptotic/necrotic cells (phosphatidylserine externalization and compromised cell membrane integrity).

After staining with Annexin V-FITC, the cells were analyzed by flow cytometry to determine the percentage of cells in each apoptotic state. The Annexin V-FITC assay provided quantitative data on the induction of apoptosis in colorectal cancer cells treated with the AURKA and Haspin kinase inhibitors.

This assay helped evaluate the efficacy of the inhibitors in promoting apoptosis, which is a critical process for inhibiting cancer cell growth and promoting tumor regression. So, the aim was to explore potential strategies for enhancing the effectiveness of targeted therapies in CRC treatment.

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Conclusion

The administration of AURKA inhibitors has revealed significant toxicities, raising concerns regarding their clinical utility. Undesirable effects, such as central nervous system dysregulation resulting in symptoms like drowsiness and dizziness, have been observed. These adverse events are believed to be linked to the interaction of AURKA inhibitors with GABA-A receptors. Therefore, it is crucial to explore and evaluate novel inhibitors that can offer positive clinical outcomes with minimal harm. Additionally, investigating the potential of combination therapies involving AURKA inhibitors and other oncogene inhibitors holds promise and merits extensive investigation as it may yield improved treatment responses. Furthermore, it is essential to recognize that achieving successful outcomes in cancer therapy necessitates the downregulation of downstream targets regulated by AURKA. Understanding the duration of effectiveness of AURKA inhibitors in inhibiting cancer cells and sustaining long-term remission is also critical. To comprehensively elucidate the precise mechanisms and limitations of existing inhibitors, well-designed preclinical studies and clinical trials are imperative. These endeavors will facilitate the development of new compounds, analogs, and derivatives that can enhance the long-term efficacy and safety profiles of AURKA inhibitors.

In summary, while AURKA inhibitors offer promise in cancer treatment, their associated toxicities and the need for sustained therapeutic response necessitate further investigation. Exploring combination therapies, understanding downstream targets, and optimizing the duration of effectiveness are crucial aspects that require attention. Continued research efforts, including preclinical and clinical studies, will pave the way for the development of improved AURKA inhibitors and ultimately contribute to enhanced patient outcomes in cancer therapy.

References

1. Co-inhibition of Aurora A and Haspin kinases enhances survivin blockage and p53 induction for mitotic catastrophe and apoptosis in human colorectal cancer. Chien-I Lina, Zan-Chu Chena, Chien-Hung Chena, Yun-Hsuan Changa, Tsai-Chia Leea, Tsai-Tai Tanga, Tzu-Wei Yua, Chih-Man Yangb, Ming-Chang Tsaic, Chi-Chou Huangd, Tzu-Wei Yangb, Chun-Che Linb, Rou-Hsin Wanga, Guang-Yuh Chioua, Yuh-Jyh Jonga,. s.l. : Elsevier, Science Direct, Biochemical Pharmacology , 2022, Elsevier, Science Direct, Biochemical Pharmacology.

2. Inhibition of Aurora kinase A activity enhances the antitumor response of beta-catenin blockade in human adrenocortical cancer cells. Andrea Gutierrez Mariaa, Kleiton Silva Borgesb, R.C.P. Lirab, Carolina Hassib Thom ́ec, Annabel Berthona, Ludivine Drougata, Katja Kiseljak-Vassiliadesd, Margaret E. Wiermand, Fabio R. Faucza, Vitor Marcel Façac, Luiz Gonzaga Toneb, Constantin. 2021, Elsevier, Science Direct, Molecular and Cellular Endocrinology, p. 10.

3. Inhibition of Aurora Kinase A Induces Necroptosis in Pancreatic Carcinoma, Gastroenterology. Author manuscript; available in PMC 2018 November 01. Yangchun Xie, Shan Zhu, Meizuo Zhong, Minghua Yang, Xiaofan Sun, Jinbao Liu, Guido Kroemer, Michael Lotze, Herbert J. Zeh III, Rui Kang, and Daolin Tang. 2017, HHS Public Access, Department of Health and Human Services, USA, p. 26.

4. Aurora A–mediated pyruvate kinase M2 phosphorylation. Ya Jiang, Ting Wang, Dandan Sheng, Chaoqiang Han, Tian Xu, Peng Zhang, Weiyi You, Weiwei Fan, Zhiyong Zhang, Tengchuan Jin, Xiaotao Duan, Xiao Yuan, Xing Liu, Kaiguang Zhang, Ke Ruan, Jue Shi, Jing Guo, Aoxing Cheng and Zhenye Yang. s.l. : Elsevier, Science Direct, Journal of Biological Chemistry , 2022.

5. Aurora kinase an inhibitor MLN8237 supresses pancreatic cancer growth. Yuebo Zhang, Young Ma, Ying Wang, Debabrata Mukhopadhyay, Yan Bi. 2022, Science Direct, Elsevier, p. 7.