Publications by authors named "Anna Aakula"

6 Publications

  • Page 1 of 1

Phosphoproteome and drug-response effects mediated by the three protein phosphatase 2A inhibitor proteins CIP2A, SET, and PME-1.

J Biol Chem 2020 03 18;295(13):4194-4211. Epub 2020 Feb 18.

Turku Bioscience Centre, University of Turku and Åbo Akademi University, 20500 Turku, Finland; Institute of Biomedicine, University of Turku, 20500 Turku, Finland. Electronic address:

Protein phosphatase 2A (PP2A) critically regulates cell signaling and is a human tumor suppressor. PP2A complexes are modulated by proteins such as cancerous inhibitor of protein phosphatase 2A (CIP2A), protein phosphatase methylesterase 1 (PME-1), and SET nuclear proto-oncogene (SET) that often are deregulated in cancers. However, how they impact cellular phosphorylation and how redundant they are in cellular regulation is poorly understood. Here, we conducted a systematic phosphoproteomics screen for phosphotargets modulated by siRNA-mediated depletion of CIP2A, PME-1, and SET (to reactivate PP2A) or the scaffolding A-subunit of PP2A (PPP2R1A) (to inhibit PP2A) in HeLa cells. We identified PP2A-modulated targets in diverse cellular pathways, including kinase signaling, cytoskeleton, RNA splicing, DNA repair, and nuclear lamina. The results indicate nonredundancy among CIP2A, PME-1, and SET in phosphotarget regulation. Notably, PP2A inhibition or reactivation affected largely distinct phosphopeptides, introducing a concept of nonoverlapping phosphatase inhibition- and activation-responsive sites (PIRS and PARS, respectively). This phenomenon is explained by the PPP2R1A inhibition impacting primarily dephosphorylated threonines, whereas PP2A reactivation results in dephosphorylation of clustered and acidophilic sites. Using comprehensive drug-sensitivity screening in PP2A-modulated cells to evaluate the functional impact of PP2A across diverse cellular pathways targeted by these drugs, we found that consistent with global phosphoproteome effects, PP2A modulations broadly affect responses to more than 200 drugs inhibiting a broad spectrum of cancer-relevant targets. These findings advance our understanding of the phosphoproteins, pharmacological responses, and cellular processes regulated by PP2A modulation and may enable the development of combination therapies.
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http://dx.doi.org/10.1074/jbc.RA119.011265DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7105317PMC
March 2020

PP2A inhibition is a druggable MEK inhibitor resistance mechanism in KRAS-mutant lung cancer cells.

Sci Transl Med 2018 07;10(450)

Turku Centre for Biotechnology, University of Turku and Åbo Akademi University, 20520 Turku, Finland.

Kinase inhibitor resistance constitutes a major unresolved clinical challenge in cancer. Furthermore, the role of serine/threonine phosphatase deregulation as a potential cause for resistance to kinase inhibitors has not been thoroughly addressed. We characterize protein phosphatase 2A (PP2A) activity as a global determinant of KRAS-mutant lung cancer cell resistance across a library of >200 kinase inhibitors. The results show that PP2A activity modulation alters cancer cell sensitivities to a large number of kinase inhibitors. Specifically, PP2A inhibition ablated mitogen-activated protein kinase kinase (MEK) inhibitor response through the collateral activation of AKT/mammalian target of rapamycin (mTOR) signaling. Combination of mTOR and MEK inhibitors induced cytotoxicity in PP2A-inhibited cells, but even this drug combination could not abrogate MYC up-regulation in PP2A-inhibited cells. Treatment with an orally bioavailable small-molecule activator of PP2A DT-061, in combination with the MEK inhibitor AZD6244, resulted in suppression of both p-AKT and MYC, as well as tumor regression in two KRAS-driven lung cancer mouse models. DT-061 therapy also abrogated MYC-driven tumorigenesis. These data demonstrate that PP2A deregulation drives MEK inhibitor resistance in KRAS-mutant cells. These results emphasize the need for better understanding of phosphatases as key modulators of cancer therapy responses.
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http://dx.doi.org/10.1126/scitranslmed.aaq1093DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8335581PMC
July 2018

Systematic Identification of MicroRNAs That Impact on Proliferation of Prostate Cancer Cells and Display Changed Expression in Tumor Tissue.

Eur Urol 2016 06 20;69(6):1120-8. Epub 2015 Oct 20.

Institute for Molecular Medicine Finland (FIMM), University of Helsinki, Helsinki, Finland; VTT Technical Research Centre of Finland, Medical Biotechnology, Turku, Finland(1).

Background: Systematic approaches to functionally identify key players in microRNA (miRNA)-target networks regulating prostate cancer (PCa) proliferation are still missing.

Objective: To comprehensively map miRNA regulation of genes relevant for PCa proliferation through phenotypic screening and tumor expression data.

Design, Setting, And Participants: Gain-of-function screening with 1129 miRNA molecules was performed in five PCa cell lines, measuring proliferation, viability, and apoptosis. These results were integrated with changes in miRNA expression from two cohorts of human PCa (188 tumors in total). For resulting miRNAs, the predicted targets were collected and analyzed for patterns with gene set enrichment analysis, and for their association with biochemical recurrence free survival.

Outcome Measurements And Statistical Analysis: Rank product statistical analysis was used to evaluate miRNA effects in phenotypic screening and for expression differences in the prostate tumor cohorts. Expression data were analyzed using the significance analysis of microarrays (SAM) method and the patient material was subjected to Kaplan-Meier statistics.

Results And Limitations: Functional screening identified 25 miRNAs increasing and 48 miRNAs decreasing cell viability. Data integration resulted in 14 miRNAs, with aberrant expression and effect on proliferation. These miRNAs are predicted to regulate >3700 genes, of which 28 were found up-regulated and 127 down-regulated in PCa compared with benign tissue. Seven genes, FLNC, MSRB3, PARVA, PCDH7, PRNP, RAB34, and SORBS1, showed an inverse association to their predicted miRNA, and were identified to significantly correlate with biochemical recurrence free survival in PCa patients.

Conclusions: A systematic in vitro screening approach combined with in vivo expression and gene set enrichment analysis provide unbiased means for revealing novel miRNA-target links, possibly driving the oncogenic processes in PCa.

Patient Summary: This study identified novel regulatory molecules, which impact on PCa proliferation and are aberrantly expressed in clinical tumors. Thus, our study reveals regulatory nodes with potential for therapy.
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http://dx.doi.org/10.1016/j.eururo.2015.09.019DOI Listing
June 2016

MicroRNA-135b regulates ERα, AR and HIF1AN and affects breast and prostate cancer cell growth.

Mol Oncol 2015 Aug 21;9(7):1287-300. Epub 2015 Mar 21.

Institute for Molecular Medicine Finland, FIMM, Helsinki, Finland.

MicroRNAs (miRNAs) regulate a wide range of cellular signaling pathways and biological processes in both physiological and pathological states such as cancer. We have previously identified miR-135b as a direct regulator of androgen receptor (AR) protein level in prostate cancer (PCa). We wanted to further explore the relationship of miR-135b to hormonal receptors, particularly estrogen receptor α (ERα). Here we show that miR-135b expression is lower in ERα-positive breast tumors as compared to ERα-negative samples in two independent breast cancer (BCa) patient cohorts (101 and 1302 samples). Additionally, the miR-135b expression is higher in AR-low PCa patient samples (47 samples). We identify ERα as a novel miR-135b target by demonstrating miR-135b binding to the 3'UTR of the ERα and decreased ERα protein and mRNA level upon miR-135b overexpression in BCa cells. MiR-135b reduces proliferation of ERα-positive BCa cells MCF-7 and BT-474 as well as AR-positive PCa cells LNCaP and 22Rv1 when grown in 2D. To identify other genes regulated by miR-135b we performed gene expression studies and found a link to the hypoxia inducible factor 1α (HIF1α) pathway. We show that miR-135b influences the protein level of the inhibitor for hypoxia inducible factor 1α (HIF1AN) and is able to bind to HIF1AN 3'UTR. Our study demonstrates that miR-135b regulates ERα, AR and HIF1AN protein levels through interaction with their 3'UTR regions, and proliferation in ERα-positive BCa and AR-positive PCa cells.
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http://dx.doi.org/10.1016/j.molonc.2015.03.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5528813PMC
August 2015

miR-183 in prostate cancer cells positively regulates synthesis and serum levels of prostate-specific antigen.

Eur Urol 2015 Oct 30;68(4):581-8. Epub 2014 Dec 30.

Department of Laboratory Medicine, Lund, Division of Clinical Chemistry, Lund University, Malmö, Sweden; Department of Laboratory Medicine, Lund, Division of Translational Cancer Research, Lund University, Lund, Sweden. Electronic address:

Background: Factors affecting serum prostate-specific antigen (PSA) levels in men are clinically important, but apart from effects mediated through the androgen receptor, they are poorly understood.

Objective: To investigate whether microRNA (miRNA) affects the synthesis and serum levels of PSA.

Design, Setting, And Participants: Reporter assays with PSA and KLK2 3' untranslated regions (UTRs) to confirm posttranscriptional regulation was followed by high-throughput screening of the effect of 1129 miRNAs on PSA levels using reverse phase protein arrays (RPPAs) to identify individual regulatory miRNAs. The candidate miRNAs were investigated further in vitro by Western blot, immunofluorometrics, activity assays, quantitative reverse transcriptase polymerase chain reaction, reporter assays, and growth assays. Prostate levels of miR-183 were compared with PSA transcript and serum PSA levels in prostate cancer cohorts.

Outcome Measurements And Statistical Analysis: RankProd was used to evaluate the RPPAs, and the Student t test was used for the in vitro experiments. The Spearman and Cuzick tests were used in the patient material, and overall survival was analysed by Kaplan-Meier and log-rank analysis.

Results And Limitations: Gain-of-function screenings identified 32 miRNAs that increase PSA levels. One of these, miR-183, was found to bind the 3' UTR of PSA directly and increase both protein and messenger RNA levels. Prostatic levels of miR-183 and serum PSA showed correlation in a cohort of 74 men. In addition, miR-183 promotes cellular growth in vitro and correlates to clinical parameters such as World Health Organisation grade and clinical progression.

Conclusions: The synthesis and serum levels of PSA are directly affected by miR-183 and may be a factor to consider when PSA values are evaluated in clinical settings.

Patient Summary: These findings offer novel insights into the regulation of prostate-specific antigen and may eventually affect clinical decision making in prostate cancer.
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http://dx.doi.org/10.1016/j.eururo.2014.12.025DOI Listing
October 2015

Systematic analysis of microRNAs targeting the androgen receptor in prostate cancer cells.

Cancer Res 2011 Mar 22;71(5):1956-67. Epub 2011 Feb 22.

Medical Biotechnology, VTT Technical Research Centre of Finland, Turku, Finland.

Androgen receptor (AR) is expressed in all stages of prostate cancer progression, including in castration-resistant tumors. Eliminating AR function continues to represent a focus of therapeutic investigation, but AR regulatory mechanisms remain poorly understood. To systematically characterize mechanisms involving microRNAs (miRNAs), we conducted a gain-of function screen of 1129 miRNA molecules in a panel of human prostate cancer cell lines and quantified changes in AR protein content using protein lysate microarrays. In this way, we defined 71 unique miRNAs that influenced the level of AR in human prostate cancer cells. RNA sequencing data revealed that the 3'UTR of AR (and other genes) is much longer than currently used in miRNA target prediction programs. Our own analyses predicted that most of the miRNA regulation of AR would target an extended 6 kb 3'UTR. 3'UTR-binding assays validated 13 miRNAs that are able to regulate this long AR 3'UTR (miR-135b, miR-185, miR-297, miR-299-3p, miR-34a, miR-34c, miR-371-3p, miR-421, miR-449a, miR-449b, miR-634, miR-654-5p, and miR-9). Fifteen AR downregulating miRNAs decreased androgen-induced proliferation of prostate cancer cells. In particular, analysis of clinical prostate cancers confirmed a negative correlation of miR-34a and miR-34c expression with AR levels. Our findings establish that miRNAs interacting with the long 3'UTR of the AR gene are important regulators of AR protein levels, with implications for developing new therapeutic strategies to inhibit AR function and androgen-dependent cell growth.
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http://dx.doi.org/10.1158/0008-5472.CAN-10-2421DOI Listing
March 2011
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