Publications by authors named "Alan P Fields"

76 Publications

FoxM1 insufficiency hyperactivates Ect2-RhoA-mDia1 signaling to drive cancer.

Nat Cancer 2020 Oct 12;1(10):1010-1024. Epub 2020 Oct 12.

Department of Biochemistry and Molecular Biology, Mayo Clinic Rochester, MN, USA.

FoxM1 activates genes that regulate S-G2-M cell-cycle progression and, when overexpressed, is associated with poor clinical outcome in multiple cancers. Here we identify FoxM1 as a tumor suppressor in mice that, through its N-terminal domain, binds to and inhibits Ect2 to limit the activity of RhoA GTPase and its effector mDia1, a catalyst of cortical actin nucleation. FoxM1 insufficiency impedes centrosome movement through excessive cortical actin polymerization, thereby causing the formation of non-perpendicular mitotic spindles that missegregate chromosomes and drive tumorigenesis in mice. Importantly, low expression correlates with RhoA GTPase hyperactivity in multiple human cancer types, indicating that suppression of the newly discovered Ect2-RhoAmDia1 oncogenic axis by FoxM1 is clinically relevant. Furthermore, by dissecting the domain requirements through which FoxM1 inhibits Ect2 GEF activity, we provide mechanistic insight for the development of pharmacological approaches that target protumorigenic RhoA activity.
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http://dx.doi.org/10.1038/s43018-020-00116-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8623810PMC
October 2020

Protein kinase C and SRC signaling define reciprocally related subgroups of glioblastoma with distinct therapeutic vulnerabilities.

Cell Rep 2021 Nov;37(8):110054

Department of Cancer Biology, Mayo Clinic, Jacksonville, FL 32224, USA. Electronic address:

We report that atypical protein kinase Cι (PKCι) is an oncogenic driver of glioblastoma (GBM). Deletion or inhibition of PKCι significantly impairs tumor growth and prolongs survival in murine GBM models. GBM cells expressing elevated PKCι signaling are sensitive to PKCι inhibitors, whereas those expressing low PKCι signaling exhibit active SRC signaling and sensitivity to SRC inhibitors. Resistance to the PKCι inhibitor auranofin is associated with activated SRC signaling and response to a SRC inhibitor, whereas resistance to a SRC inhibitor is associated with activated PKCι signaling and sensitivity to auranofin. Interestingly, PKCι- and SRC-dependent cells often co-exist in individual GBM tumors, and treatment of GBM-bearing mice with combined auranofin and SRC inhibitor prolongs survival beyond either drug alone. Thus, we identify PKCι and SRC signaling as distinct therapeutic vulnerabilities that are directly translatable into an improved treatment for GBM.
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http://dx.doi.org/10.1016/j.celrep.2021.110054DOI Listing
November 2021

Aberrant expression and subcellular localization of ECT2 drives colorectal cancer progression and growth.

Cancer Res 2021 Nov 4. Epub 2021 Nov 4.

Department of Pharmacology, University of North Carolina at Chapel Hill

ECT2 is an activator of RHO GTPases that is essential for cytokinesis. Additionally, ECT2 was identified as an oncoprotein when expressed ectopically in NIH/3T3 fibroblasts. However, oncogenic activation of ECT2 resulted from N-terminal truncation, and such truncated ECT2 proteins have not been found in cancer patients. In this study, we observed elevated expression of full-length ECT2 protein in preneoplastic colon adenomas, driven by increased ECT2 mRNA abundance and associated with APC tumor suppressor loss. Elevated ECT2 levels were detected in the cytoplasm and nucleus of colorectal cancer (CRC) tissue, suggesting cytoplasmic mislocalization as one mechanism of early oncogenic ECT2 activation. Importantly, elevated nuclear ECT2 correlated with poorly differentiated tumors, and a low cytoplasmic:nuclear ratio of ECT2 protein correlated with poor patient survival, suggesting that nuclear and cytoplasmic ECT2 play distinct roles in CRC. Depletion of ECT2 reduced anchorage-independent cancer cell growth and invasion independent of its function in cytokinesis, and loss of Ect2 extended survival in a KrasG12D Apc-null colon cancer mouse model. Expression of ECT2 variants with impaired nuclear localization or guanine nucleotide exchange catalytic activity failed to restore cancer cell growth or invasion, indicating that active, nuclear ECT2 is required to support tumor progression. Nuclear ECT2 promoted ribosomal DNA transcription and ribosome biogenesis in CRC. These results support a driver role for both cytoplasmic and nuclear ECT2 overexpression in CRC and emphasize the critical role of precise subcellular localization in dictating ECT2 function in neoplastic cells.
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http://dx.doi.org/10.1158/0008-5472.CAN-20-4218DOI Listing
November 2021

Editorial.

Authors:
Alan P Fields

Adv Biol Regul 2021 05 26;80:100770. Epub 2020 Nov 26.

Monica Flynn Jacoby Professor of Cancer Research, Department of Cancer Biology, Mayo Clinic Griffin Cancer Research, Building 4500 San Pablo Road, Jacksonville, 32224, Florida, United States. Electronic address:

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http://dx.doi.org/10.1016/j.jbior.2020.100770DOI Listing
May 2021

Recurrent copy number gains drive PKCι expression and PKCι-dependent oncogenic signaling in human cancers.

Adv Biol Regul 2020 12 12;78:100754. Epub 2020 Sep 12.

Department of Cancer Cell Biology, Mayo Clinic Florida, Jacksonville, FL, 32224, USA. Electronic address:

PRKCI is frequently overexpressed in multiple human cancers, and PKCι expression is often prognostic for poor patient survival, indicating that elevated PKCι broadly plays an oncogenic role in the cancer phenotype. PKCι drives multiple oncogenic signaling pathways involved in transformed growth, and transgenic mouse models have revealed that PKCι is a critical oncogenic driver in both lung and ovarian cancers. We now report that recurrent 3q26 copy number gain (CNG) is the predominant genetic driver of PRKCI mRNA expression in all major human cancer types exhibiting such CNGs. In addition to PRKCI, CNG at 3q26 leads to coordinate CNGs of ECT2 and SOX2, two additional 3q26 genes that collaborate with PRKCI to drive oncogenic signaling and tumor initiation in lung squamous cell carcinoma. Interestingly however, whereas 3q26 CNG is a strong driver of PRKCI mRNA expression across all tumor types examined, it has differential effects on ECT2 and SOX2 mRNA expression. In some tumors types, particularly those with squamous histology, all three 3q26 oncogenes are coordinately overexpressed as a consequence of 3q26 CNG, whereas in other cancers only PRKCI and ECT2 mRNA are coordinately overexpressed. This distinct pattern of expression of 3q26 genes corresponds to differences in genomic signatures reflective of activation of specific PKCι oncogenic signaling pathways. In addition to highly prevalent CNG, some tumor types exhibit monoallelic loss of PRKCI. Interestingly, many tumors harboring monoallelic loss of PRKCI express significantly lower PRKCI mRNA and exhibit evidence of WNT/β-catenin signaling pathway activation, which we previously characterized as a major oncogenic pathway in a newly described, PKCι-independent molecular subtype of lung adenocarcinoma. Finally, we show that CNG-driven activation of PKCι oncogenic signaling predicts poor patient survival in many major cancer types. We conclude that CNG and monoallelic loss are the major determinants of tumor PRKCI mRNA expression across virtually all tumor types, but that tumor-type specific mechanisms determine whether these copy number alterations also drive expression of the collaborating 3q26 oncogenes ECT2 and SOX2, and the oncogenic PKCι signaling pathways activated through the collaborative action of these genes. Our analysis may be useful in identifying tumor-specific predictive biomarkers and effective PKCι-targeted therapeutic strategies in the multitude of human cancers harboring genetic activation of PRKCI.
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http://dx.doi.org/10.1016/j.jbior.2020.100754DOI Listing
December 2020

Protein kinase Cι promotes UBF1-ECT2 binding on ribosomal DNA to drive rRNA synthesis and transformed growth of non-small-cell lung cancer cells.

J Biol Chem 2020 06 29;295(24):8214-8226. Epub 2020 Apr 29.

Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center, Jacksonville, Florida, USA

Epithelial cell-transforming sequence 2 (ECT2) is a guanine nucleotide exchange factor for Rho GTPases that is overexpressed in many cancers and involved in signal transduction pathways that promote cancer cell proliferation, invasion, and tumorigenesis. Recently, we demonstrated that a significant pool of ECT2 localizes to the nucleolus of non-small-cell lung cancer (NSCLC) cells, where it binds the transcription factor upstream binding factor 1 (UBF1) on the promoter regions of ribosomal DNA (rDNA) and activates rDNA transcription, transformed cell growth, and tumor formation. Here, we investigated the mechanism by which ECT2 engages UBF1 on rDNA promoters. Results from ECT2 mutagenesis indicated that the tandem BRCT domain of ECT2 mediates binding to UBF1. Biochemical and MS-based analyses revealed that protein kinase Cι (PKCι) directly phosphorylates UBF1 at Ser-412, thereby generating a phosphopeptide-binding epitope that binds the ECT2 BRCT domain. Lentiviral shRNA knockdown and reconstitution experiments revealed that both a functional ECT2 BRCT domain and the UBF1 Ser-412 phosphorylation site are required for UBF1-mediated ECT2 recruitment to rDNA, elevated rRNA synthesis, and transformed growth. Our findings provide critical molecular insight into ECT2-mediated regulation of rDNA transcription in cancer cells and offer a rationale for therapeutic targeting of UBF1- and ECT2-stimulated rDNA transcription for the management of NSCLC.
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http://dx.doi.org/10.1074/jbc.RA120.013175DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7294082PMC
June 2020

Chromosome 3q26 Gain Is an Early Event Driving Coordinated Overexpression of the PRKCI, SOX2, and ECT2 Oncogenes in Lung Squamous Cell Carcinoma.

Cell Rep 2020 01;30(3):771-782.e6

Department of Cancer Biology, Mayo Clinic Florida, Jacksonville, FL 32224, USA. Electronic address:

Lung squamous cell carcinoma (LSCC) is a prevalent form of lung cancer exhibiting distinctive histological and genetic characteristics. Chromosome 3q26 copy number gain (CNG) is a genetic hallmark of LSCC present in >90% of tumors. We report that 3q26 CNGs occur early in LSCC tumorigenesis, persist during tumor progression, and drive coordinate overexpression of PRKCI, SOX2, and ECT2. Overexpression of PRKCI, SOX2, and ECT2 in the context of Trp53 loss is sufficient to transform mouse lung basal stem cells into tumors with histological and genomic features of LSCC. Functionally, PRKCI and SOX2 collaborate to activate an extensive transcriptional program that enforces a lineage-restricted LSCC phenotype, whereas PRKCI and ECT2 collaborate to promote oncogenic growth. Gene signatures indicative of PKCι-SOX2 and PKCι-ECT2 signaling activity are enriched in the classical subtype of human LSCC and predict distinct therapeutic vulnerabilities. Thus, the PRKCI, SOX2, and ECT2 oncogenes represent a multigenic driver of LSCC.
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http://dx.doi.org/10.1016/j.celrep.2019.12.071DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7238436PMC
January 2020

Oncogenic protein kinase Cι signaling mechanisms in lung cancer: Implications for improved therapeutic strategies.

Adv Biol Regul 2020 01 25;75:100656. Epub 2019 Sep 25.

From the Department of Cancer Cell Biology, Mayo Clinic Florida, Jacksonville, FL, 32224, USA. Electronic address:

Protein Kinase Cι (PKCι) is a major oncogene involved in the initiation, maintenance and progression of numerous forms of human cancer. In the lung, PKCι is necessary for the maintenance of the transformed phenotype of the two major forms of non-small cell lung cancer (NSCLC), lung adenocarcinoma (LADC) and lung squamous cell carcinoma (LSCC). In addition, PKCι is necessary for both LADC and LSCC tumorigenesis by establishing and maintaining a highly aggressive stem-like, tumor-initiating cell phenotype. Interestingly however, while PKCι signaling in these two major lung cancer subtypes shares some common elements, it also drives distinct, sub-type specific pathways. Furthermore, recent analysis has revealed both PKCι-dependent and PKCι-independent pathways to LADC development. Herein, we discussion our current knowledge of oncogenic PKCι signaling in LADC and LSCC, and discuss these findings in the context of how they may inform strategies for improved therapeutic intervention in these deadly diseases.
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http://dx.doi.org/10.1016/j.jbior.2019.100656DOI Listing
January 2020

Protein Kinase Cι and Wnt/β-Catenin Signaling: Alternative Pathways to Kras/Trp53-Driven Lung Adenocarcinoma.

Cancer Cell 2019 08 1;36(2):156-167.e7. Epub 2019 Aug 1.

Department of Cancer Biology, Mayo Clinic Florida, 4500 San Pablo Road, Griffin Cancer Research Building, Room 212, Jacksonville, FL 32224, USA. Electronic address:

We report that mouse LSL-Kras;Trp53 (KP)-mediated lung adenocarcinoma (LADC) tumorigenesis can proceed through both PKCι-dependent and PKCι-independent pathways. The predominant pathway involves PKCι-dependent transformation of bronchoalveolar stem cells (BASCs). However, KP mice harboring conditional knock out Prkci alleles (KPI mice) develop LADC tumors through PKCι-independent transformation of Axin2 alveolar type 2 (AT2) stem cells. Transformed growth of KPI, but not KP, tumors is blocked by Wnt pathway inhibition in vitro and in vivo. Furthermore, a KPI-derived genomic signature predicts sensitivity of human LADC cells to Wnt inhibition, and identifies a distinct subset of primary LADC tumors exhibiting a KPI-like genotype. Thus, LADC can develop through both PKCι-dependent and PKCι-independent pathways, resulting in tumors exhibiting distinct oncogenic signaling and pharmacologic vulnerabilities.
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http://dx.doi.org/10.1016/j.ccell.2019.07.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6693680PMC
August 2019

Protein kinase C: A versatile oncogene in the lung.

Mol Cell Oncol 2018 10;5(5):e1190886. Epub 2018 May 10.

Department of Cancer Biology, Mayo Clinic Florida Jacksonville, Florida.

We have recently demonstrated that protein kinase C (PKC) promotes a stem-like, tumor-initiating cell phenotype in -driven lung adenocarcinoma by activating a novel ELF3-NOTCH3 signaling axis. Combined PKC and NOTCH inhibition was identified as a novel strategy for the treatment of -driven lung adenocarcinoma.
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http://dx.doi.org/10.1080/23723556.2016.1190886DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6154840PMC
May 2018

Oncogenic Ect2 signaling regulates rRNA synthesis in NSCLC.

Small GTPases 2019 09 5;10(5):388-394. Epub 2017 Jul 5.

a Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center , Jacksonville , FL , USA.

The Rho GTPase family members Rac1, Cdc42 and RhoA play key contributory roles in the transformed phenotype of human cancers. Epithelial Cell Transforming Sequence 2 (Ect2), a guanine nucleotide exchange factor (GEF) for these Rho GTPases, has also been implicated in a variety of human cancers. We have shown that Ect2 is frequently overexpressed in both major forms of non-small cell lung cancer (NSCLC), lung adenocarcinoma (LADC) and lung squamous cell carcinoma (LSCC), which together make up approximately 70% of all lung cancer diagnoses. Furthermore, we have found that Ect2 is required for multiple aspects of the transformed phenotype of NSCLC cells including transformed growth and invasion and tumorigenesis . More recently, we showed that a major mechanism by which Ect2 drives -mediated LADC transformation is by regulating rRNA (rRNA) synthesis. However, it remains unclear whether Ect2 plays a similar role in ribosome biogenesis in LSCC. Here we demonstrate that Ect2 expression correlates positively with expression of ribosome biogenesis genes and with pre-ribosomal 45S RNA abundance in primary LSCC tumors. Furthermore, we demonstrate that Ect2 functionally regulates rRNA synthesis in LSCC cells. Based on these data, we propose that inhibition of Ect2-mediated nucleolar signaling holds promise as a potential therapeutic strategy for improved treatment of both LADC and LSCC.
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http://dx.doi.org/10.1080/21541248.2017.1335274DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6748369PMC
September 2019

Ect2-Dependent rRNA Synthesis Is Required for KRAS-TRP53-Driven Lung Adenocarcinoma.

Cancer Cell 2017 02 19;31(2):256-269. Epub 2017 Jan 19.

Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center, Griffin Cancer Research Building, Room 212, 4500 San Pablo Road, Jacksonville, FL 32224, USA. Electronic address:

The guanine nucleotide exchange factor (GEF) epithelial cell transforming sequence 2 (Ect2) has been implicated in cancer. However, it is not clear how Ect2 causes transformation and whether Ect2 is necessary for tumorigenesis in vivo. Here, we demonstrate that nuclear Ect2 GEF activity is required for Kras-Trp53 lung tumorigenesis in vivo and that Ect2-mediated transformation requires Ect2-dependent rDNA transcription. Ect2 activates rRNA synthesis by binding the nucleolar transcription factor upstream binding factor 1 (UBF1) on rDNA promoters and recruiting Rac1 and its downstream effector nucleophosmin (NPM) to rDNA. Protein kinase Cι (PKCι)-mediated Ect2 phosphorylation stimulates Ect2-dependent rDNA transcription. Thus, Ect2 regulates rRNA synthesis through a PKCι-Ect2-Rac1-NPM signaling axis that is required for lung tumorigenesis.
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http://dx.doi.org/10.1016/j.ccell.2016.12.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5310966PMC
February 2017

A proof-of-concept trial of protein kinase C iota inhibition with auranofin for the paclitaxel-induced acute pain syndrome.

Support Care Cancer 2017 03 12;25(3):833-838. Epub 2016 Nov 12.

Cancer Biology, Mayo Clinic, Jacksonville, FL, USA.

Purpose: Paclitaxel causes the paclitaxel-induced acute pain (PIAP) syndrome. Based on preclinical data, we hypothesized that the protein kinase C (PKC) iota inhibitor, auranofin (a gold salt used for other pain conditions), palliates this pain.

Methods: In a randomized, double-blinded manner, patients who had suffered this syndrome were assigned a one-time dose of auranofin 6 mg orally on day #2 of the chemotherapy cycle (post-paclitaxel) versus placebo. Patients completed the Brief Pain Inventory and a pain diary on days 2 through 8 and at the end of the cycle. The primary endpoint was pain scores, as calculated by area under the curve, in response to "Please rate your pain by circling the one number that best describes your pain at its worse in the last 24 hours."

Results: Thirty patients were enrolled. For the primary endpoint, mean area under the curve of 55 units (standard deviation 19) and 61 units (standard deviation 22) were observed in auranofin-treated and placebo-exposed patients, respectively (p = 0.44). On day 8 and at the end of the cycle, pain scores in auranofin-treated patients were more favorable, although differences were not statistically significant.

Conclusions: In the dose schedule studied, auranofin did not palliate the PIAP syndrome, but delayed beneficial trends suggest further study for this indication.
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http://dx.doi.org/10.1007/s00520-016-3467-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5269609PMC
March 2017

SOX2 Determines Lineage Restriction: Modeling Lung Squamous Cell Carcinoma in the Mouse.

Cancer Cell 2016 10;30(4):505-507

Department of Cancer Biology, Mayo Clinic College of Medicine, Jacksonville, FL 32224, USA. Electronic address:

In this issue of Cancer Cell, Ferone et al. demonstrate that SOX2 not only drives lung tumor formation but also restricts tumor lineage to squamous cell carcinoma (LSCC), regardless of cell of origin. This novel LSCC model should facilitate identification of key oncogenic drivers and treatment strategies for this lung cancer subtype.
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http://dx.doi.org/10.1016/j.ccell.2016.09.012DOI Listing
October 2016

Oncogenic PKCι decides tumor-initiating cell fate.

Cell Cycle 2016 09 17;15(18):2383-4. Epub 2016 Jun 17.

a Department of Cancer Biology , Mayo Clinic , Jacksonville , FL , USA.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5026803PMC
http://dx.doi.org/10.1080/15384101.2016.1194624DOI Listing
September 2016

Targeting oncogenic protein kinase Cι for treatment of mutant KRAS LADC.

Small GTPases 2017 01 31;8(1):58-64. Epub 2016 May 31.

a Department of Cancer Biology , Mayo Clinic , Jacksonville , FL , USA.

Lung cancer is the leading cause of cancer death in the US with ∼124,000 new cases annually, and a 5 y survival rate of ∼16%. Mutant KRAS-driven lung adenocarcinoma (KRAS LADC) is a particularly prevalent and deadly form of lung cancer. Protein kinase Cι (PKCι) is an oncogenic effector of KRAS that activates multiple signaling pathways that stimulate transformed growth and invasion, and maintain a KRAS LADC tumor-initiating cell (TIC) phenotype. PKCι inhibitors used alone and in strategic combination show promise as new therapeutic approaches to treatment of KRAS LADC. These novel drug combinations may improve clinical management of KRAS LADC.
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http://dx.doi.org/10.1080/21541248.2016.1194953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5331898PMC
January 2017

Stabilin-1 is expressed in human breast cancer and supports tumor growth in mammary adenocarcinoma mouse model.

Oncotarget 2016 May;7(21):31097-110

Institute for Transfusion Medicine and Immunology, Medical Faculty Mannheim, Ruprecht-Karls University of Heidelberg, Mannheim, Germany.

Stabilin-1 is a multifunctional scavenger receptor expressed on alternatively-activated macrophages. Stabilin-1 mediates phagocytosis of "unwanted-self" components, intracellular sorting, and endocytic clearance of extracellular ligands including SPARC that modulates breast cancer growth. The expression of stabilin-1 was found on tumor-associated macrophages (TAM) in mouse and human cancers including melanoma, lymphoma, glioblastoma, and pancreatic insulinoma. Despite its tumor-promoting role in mouse models of melanoma and lymphoma the expression and functional role of stabilin-1 in breast cancer was unknown. Here, we demonstrate that stabilin-1 is expressed on TAM in human breast cancer, and its expression is most pronounced on stage I disease. Using stabilin-1 knockout (ko) mice we show that stabilin-1 facilitates growth of mouse TS/A mammary adenocarcinoma. Endocytosis assay on stabilin-1 ko TAM demonstrated impaired clearance of stabilin-1 ligands including SPARC that was capable of inducing cell death in TS/A cells. Affymetrix microarray analysis on purified TAM and reporter assays in stabilin-1 expressing cell lines demonstrated no influence of stabilin-1 expression on intracellular signalling. Our results suggest stabilin-1 mediated silent clearance of extracellular tumor growth-inhibiting factors (e.g. SPARC) as a mechanism of stabilin-1 induced tumor growth. Silent clearance function of stabilin-1 makes it an attractive candidate for delivery of immunomodulatory anti-cancer therapeutic drugs to TAM.
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http://dx.doi.org/10.18632/oncotarget.8857DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5058742PMC
May 2016

Protein Kinase Cι Drives a NOTCH3-dependent Stem-like Phenotype in Mutant KRAS Lung Adenocarcinoma.

Cancer Cell 2016 Mar;29(3):367-378

Department of Cancer Biology, Mayo Clinic Cancer Center, Jacksonville, FL 32224, USA. Electronic address:

We report that the protein kinase Cι (PKCι) oncogene controls expression of NOTCH3, a key driver of stemness, in KRAS-mediated lung adenocarcinoma (LADC). PKCι activates NOTCH3 expression by phosphorylating the ELF3 transcription factor and driving ELF3 occupancy on the NOTCH3 promoter. PKCι-ELF3-NOTCH3 signaling controls the tumor-initiating cell phenotype by regulating asymmetric cell division, a process necessary for tumor initiation and maintenance. Primary LADC tumors exhibit PKCι-ELF3-NOTCH3 signaling, and combined pharmacologic blockade of PKCι and NOTCH synergistically inhibits tumorigenic behavior in vitro and LADC growth in vivo demonstrating the therapeutic potential of PKCι-ELF3-NOTCH3 signal inhibition to more effectively treat KRAS LADC.
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http://dx.doi.org/10.1016/j.ccell.2016.02.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4795153PMC
March 2016

The chromosome 3q26 OncCassette: A multigenic driver of human cancer.

Adv Biol Regul 2016 Jan 23;60:47-63. Epub 2015 Dec 23.

Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center, Jacksonville, FL, United States.

Recurrent copy number variations (CNVs) are genetic alterations commonly observed in human tumors. One of the most frequent CNVs in human tumors involves copy number gains (CNGs) at chromosome 3q26, which is estimated to occur in >20% of human tumors. The high prevalence and frequent occurrence of 3q26 CNG suggest that it drives the biology of tumors harboring this genetic alteration. The chromosomal region subject to CNG (the 3q26 amplicon) spans from chromosome 3q26 to q29, a region containing ∼200 protein-encoding genes. The large number of genes within the amplicon makes it difficult to identify relevant oncogenic target(s). Whereas a number of genes in this region have been linked to the transformed phenotype, recent studies indicate a high level of cooperativity among a subset of frequently amplified 3q26 genes. Here we use a novel bioinformatics approach to identify potential driver genes within the recurrent 3q26 amplicon in lung squamous cell carcinoma (LSCC). Our analysis reveals a set of 35 3q26 amplicon genes that are coordinately amplified and overexpressed in human LSCC tumors, and that also map to a major LSCC susceptibility locus identified on mouse chromosome 3 that is syntenic with human chromosome 3q26. Pathway analysis reveals that 21 of these genes exist within a single predicted network module. Four 3q26 genes, SOX2, ECT2, PRKCI and PI3KCA occupy the hub of this network module and serve as nodal genes around which the network is organized. Integration of available genetic, genomic, biochemical and functional data demonstrates that SOX2, ECT2, PRKCI and PIK3CA are cooperating oncogenes that function within an integrated cell signaling network that drives a highly aggressive, stem-like phenotype in LSCC tumors harboring 3q26 amplification. Based on the high level of genomic, genetic, biochemical and functional integration amongst these 4 3q26 nodal genes, we propose that they are the key oncogenic targets of the 3q26 amplicon and together define a "3q26 OncCassette" that mediates 3q26 CNG-driven tumorigenesis. Genomic analysis indicates that the 3q26 OncCassette also operates in other major tumor types that exhibit frequent 3q26 CNGs, including head and neck squamous cell carcinoma (HNSCC), ovarian serous cancer and cervical cancer. Finally, we discuss how the 3q26 OncCassette represents a tractable target for development of novel therapeutic intervention strategies that hold promise for improving treatment of 3q26-driven cancers.
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http://dx.doi.org/10.1016/j.jbior.2015.10.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4729592PMC
January 2016

A small molecule inhibitor of atypical protein kinase C signaling inhibits pancreatic cancer cell transformed growth and invasion.

Oncotarget 2015 Jun;6(17):15297-310

Department of Cancer Biology, Mayo Clinic, Jacksonville, FL, USA.

Pancreatic cancer is highly resistant to current chemotherapies. Identification of the critical signaling pathways that mediate pancreatic cancer transformed growth is necessary for the development of more effective therapeutic treatments. Recently, we demonstrated that protein kinase C iota (PKCι) and zeta (PKCζ) promote pancreatic cancer transformed growth and invasion, by activating Rac1→ERK and STAT3 signaling pathways, respectively. However, a key question is whether PKCι and PKCζ play redundant (or non-redundant) roles in pancreatic cancer cell transformed growth. Here we describe the novel observations that 1) PKCι and PKCζ are non-redundant in the context of the transformed growth of pancreatic cancer cells; 2) a gold-containing small molecule known to disrupt the PKCι/Par6 interaction, aurothiomalate, also disrupts PKCζ/Par6 interaction; 3) aurothiomalate inhibits downstream signaling of both PKCι and PKCζ, and blocks transformed growth of pancreatic cancer cells in vitro; and 4) aurothiomalate inhibits pancreatic cancer tumor growth and metastasis in vivo. Taken together, these data provide convincing evidence that an inhibitor of atypical PKC signaling inhibits two key oncogenic signaling pathways, driven non-redundantly by PKCι and PKCζ, to significantly reduce tumor growth and metastasis. Our results demonstrate that inhibition of atypical PKC signaling is a promising therapeutic strategy to treat pancreatic cancer.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4558152PMC
http://dx.doi.org/10.18632/oncotarget.3812DOI Listing
June 2015

Protein kinase D1 drives pancreatic acinar cell reprogramming and progression to intraepithelial neoplasia.

Nat Commun 2015 Feb 20;6:6200. Epub 2015 Feb 20.

Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center, Mayo Clinic, Jacksonville, Florida 32224, USA.

The transdifferentiation of pancreatic acinar cells to a ductal phenotype (acinar-to-ductal metaplasia, ADM) occurs after injury or inflammation of the pancreas and is a reversible process. However, in the presence of activating Kras mutations or persistent epidermal growth factor receptor (EGF-R) signalling, cells that underwent ADM can progress to pancreatic intraepithelial neoplasia (PanIN) and eventually pancreatic cancer. In transgenic animal models, ADM and PanINs are initiated by high-affinity ligands for EGF-R or activating Kras mutations, but the underlying signalling mechanisms are not well understood. Here, using a conditional knockout approach, we show that protein kinase D1 (PKD1) is sufficient to drive the reprogramming process to a ductal phenotype and progression to PanINs. Moreover, using 3D explant culture of primary pancreatic acinar cells, we show that PKD1 acts downstream of TGFα and Kras, to mediate formation of ductal structures through activation of the Notch pathway.
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http://dx.doi.org/10.1038/ncomms7200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4394184PMC
February 2015

Molecular pathways: novel approaches for improved therapeutic targeting of Hedgehog signaling in cancer stem cells.

Clin Cancer Res 2015 Feb;21(3):505-13

Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center, Jacksonville, Florida.

The Hedgehog (Hh) signaling pathway is critical for embryonic development. In adult tissues, Hh signaling is relatively quiescent with the exception of roles in tissue maintenance and repair. Aberrant activation of Hh signaling is implicated in multiple aspects of transformation, including the maintenance of the cancer stem cell (CSC) phenotype. Preclinical studies indicate that CSCs from many tumor types are sensitive to Hh pathway inhibition and that Hh-targeted therapeutics block many aspects of transformation attributed to CSCs, including drug resistance, relapse, and metastasis. However, to date, Hh inhibitors, specifically those targeting Smoothened [such as vismodegib, BMS-833923, saridegib (IPI-926), sonidegib/erismodegib (LDE225), PF-04449913, LY2940680, LEQ 506, and TAK-441], have demonstrated good efficacy as monotherapy in patients with basal cell carcinoma and medulloblastoma, but have shown limited activity in other tumor types. This lack of success is likely due to many factors, including a lack of patient stratification in early trials, cross-talk between Hh and other oncogenic signaling pathways that can modulate therapeutic response, and a limited knowledge of Hh pathway activation mechanisms in CSCs from most tumor types. Here, we discuss Hh signaling mechanisms in the context of human cancer, particularly in the maintenance of the CSC phenotype, and consider new therapeutic strategies that hold the potential to expand considerably the scope and therapeutic efficacy of Hh-directed anticancer therapy.
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http://dx.doi.org/10.1158/1078-0432.CCR-14-0507DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4316382PMC
February 2015

A mixed-methods feasibility trial of protein kinase C iota inhibition with auranofin in asymptomatic ovarian cancer patients.

Oncology 2015 6;88(4):208-13. Epub 2014 Dec 6.

Department of Oncology, Mayo Clinic, Rochester, Minn., USA.

Purpose: This trial was undertaken (1) to determine the feasibility of enrolling asymptomatic ovarian cancer patients with CA-125 elevation in a trial with the protein kinase C iota (PKCι) inhibitor auranofin and (2) to understand patients' perceptions of CA-125 monitoring.

Methods: Asymptomatic ovarian cancer patients with CA-125 elevation received 3 mg auranofin orally twice per day and were evaluated. The patients participated in interviews about CA-125 monitoring.

Results: Ten patients were enrolled in slightly over 6 months, exceeding our anticipated accrual rate. Four manifested stable CA-125 levels for 1 month or longer. The median progression-free survival was 2.8 months (95% CI: 1.3-3.8); auranofin was well tolerated. One patient had baseline and monthly CA-125 levels of 5,570, 6,085, 3,511, and 2,230 U/ml, respectively, stopped auranofin because of radiographic progression at 3 months, and manifested an increase in CA-125 to 7,168 U/ml approximately 3 months later. Patient interviews revealed (1) the important role of CA-125 in cancer monitoring, (2) ardent advocacy of CA-125 testing, and (3) an evolution toward CA-125 assuming a life of its own.

Conclusions: This study showed the feasibility of enrolling asymptomatic ovarian cancer patients with CA-125 elevation in a trial with auranofin. One patient had a decline in CA-125, suggesting that PKCι inhibition merits further study in ovarian cancer.
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http://dx.doi.org/10.1159/000369257DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4536897PMC
June 2015

The PRKCI and SOX2 oncogenes are coamplified and cooperate to activate Hedgehog signaling in lung squamous cell carcinoma.

Cancer Cell 2014 Feb;25(2):139-51

Department of Cancer Biology, Mayo Clinic Cancer Center, Jacksonville, FL 32224, USA. Electronic address:

We report that two oncogenes coamplified on chromosome 3q26, PRKCI and SOX2, cooperate to drive a stem-like phenotype in lung squamous cell carcinoma (LSCC). Protein kinase Cι (PKCι) phosphorylates SOX2, a master transcriptional regulator of stemness, and recruits it to the promoter of Hedgehog (Hh) acyltransferase (HHAT) that catalyzes the rate-limiting step in Hh ligand production. PKCι-mediated SOX2 phosphorylation is required for HHAT promoter occupancy, HHAT expression, and maintenance of a stem-like phenotype. Primary LSCC tumors coordinately overexpress PKCι, SOX2, and HHAT and require PKCι-SOX2-HHAT signaling to maintain a stem-like phenotype. Thus, PKCι and SOX2 are genetically, biochemically, and functionally linked in LSCC, and together they drive tumorigenesis by establishing a cell-autonomous Hh signaling axis.
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http://dx.doi.org/10.1016/j.ccr.2014.01.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3949484PMC
February 2014

Utility and applications of orthotopic models of human non-small cell lung cancer (NSCLC) for the evaluation of novel and emerging cancer therapeutics.

Curr Protoc Pharmacol 2013 Oct 8;62:14.27.1-14.27.17. Epub 2013 Oct 8.

Department of Cancer Biology, Mayo Clinic College of Medicine, Jacksonville, Florida.

Lung cancer is a leading cause of cancer deaths worldwide. Despite advances in chemotherapy, radiation therapy, and surgery, lung cancer continues to have a low 5-year survival rate, highlighting a dire need for more effective means of prevention, diagnosis, prognosis, and treatment. Mouse models that recapitulate the clinical features of advanced human lung cancer are critical for testing novel therapeutic approaches. This unit describes a highly reproducible, easy-to-establish orthotopic murine model of lung cancer, provides methods for in vivo imaging and monitoring of tumor growth, and discusses the usefulness of this model for translational lung cancer research and the development of therapeutic strategies.
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http://dx.doi.org/10.1002/0471141755.ph1427s62DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4362678PMC
October 2013

Atypical protein kinase Cι as a human oncogene and therapeutic target.

Biochem Pharmacol 2014 Mar 11;88(1):1-11. Epub 2013 Nov 11.

Department of Cancer Biology, Mayo Clinic Comprehensive Cancer Center, 45400 San Pablo Road, Jacksonville, FL 32224, USA. Electronic address:

Protein kinase inhibitors represent a major class of targeted therapeutics that has made a positive impact on treatment of cancer and other disease indications. Among the promising kinase targets for further therapeutic development are members of the Protein Kinase C (PKC) family. The PKCs are central components of many signaling pathways that regulate diverse cellular functions including proliferation, cell cycle, differentiation, survival, cell migration, and polarity. Genetic manipulation of individual PKC isozymes has demonstrated that they often fulfill distinct, nonredundant cellular functions. Participation of PKC members in different intracellular signaling pathways reflects responses to varying extracellular stimuli, intracellular localization, tissue distribution, phosphorylation status, and intermolecular interactions. PKC activity, localization, phosphorylation, and/or expression are often altered in human tumors, and PKC isozymes have been implicated in various aspects of transformation, including uncontrolled proliferation, migration, invasion, metastasis, angiogenesis, and resistance to apoptosis. Despite the strong relationship between PKC isozymes and cancer, to date only atypical PKCiota has been shown to function as a bona fide oncogene, and as such is a particularly attractive therapeutic target for cancer treatment. In this review, we discuss the role of PKCiota in transformation and describe mechanism-based approaches to therapeutically target oncogenic PKCiota signaling in cancer.
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http://dx.doi.org/10.1016/j.bcp.2013.10.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3944347PMC
March 2014

PKCι maintains a tumor-initiating cell phenotype that is required for ovarian tumorigenesis.

Mol Cancer Res 2013 Dec 30;11(12):1624-35. Epub 2013 Oct 30.

Griffin Cancer Research Building, Rm. 212, Mayo Clinic College of Medicine, 4500 San Pablo Road, Jacksonville, FL 32224.

Unlabelled: Protein kinase Cι (PKCι) has oncogenic potential and is an attractive therapeutic target for treatment of lung cancer, particularly those tumors that express elevated PKCι. However, whether PKCι is a viable target in ovarian cancer is unknown, and virtually nothing is known about the mechanism by which PKCι drives ovarian tumorigenesis. Here, it is demonstrated that PKCι maintains a tumor-initiating cell (TIC) phenotype that drives ovarian tumorigenesis. A highly tumorigenic population of cells from human ovarian cancer cell lines exhibit cancer stem-like TIC properties, including self-renewal, clonal expansion, expression of stem-related genes, enhanced transformed growth in vitro, and aggressive tumor-initiating activity in vivo. Genetic disruption of PKCι inhibits the proliferation, clonal expansion, anchorage-independent growth, and enhanced tumorigenic properties of ovarian TICs. Biochemical analysis demonstrates that PKCι acts through its oncogenic partner Ect2 to activate a MEK/ERK signaling axis that drives the ovarian TIC phenotype. Genomic analysis reveals that PKCι and Ect2 are coordinately amplified and overexpressed in the majority of primary ovarian serous tumors, and these tumors exhibit evidence of an active PKCι-Ect2 signaling axis in vivo. Finally, this study reveals that auranofin, a potent and selective inhibitor of oncogenic PKCι signaling, inhibits the tumorigenic properties of ovarian TIC cells in vitro and in vivo. These data demonstrate that PKCι is required for a TIC phenotype in ovarian cancer, and that auranofin is an attractive therapeutic option to target deadly ovarian TICs in ovarian cancer patients.

Implications: PKCι drives a tumor-initiating cell phenotype in ovarian cancer cells that can be therapeutically targeted with auranofin, a small molecule inhibitor of PKCι signaling.
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http://dx.doi.org/10.1158/1541-7786.MCR-13-0371-TDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3888959PMC
December 2013

Protein kinase C zeta regulates human pancreatic cancer cell transformed growth and invasion through a STAT3-dependent mechanism.

PLoS One 2013 28;8(8):e72061. Epub 2013 Aug 28.

Department of Cancer Biology, Mayo Clinic, Jacksonville, Florida, United States of America.

Pancreatic cancer is a very aggressive disease with few therapeutic options. In this study, we investigate the role of protein kinase C zeta (PKCζ) in pancreatic cancer cells. PKCζ has been shown to act as either a tumor suppressor or tumor promoter depending upon the cellular context. We find that PKCζ expression is either maintained or elevated in primary human pancreatic tumors, but is never lost, consistent with PKCζ playing a promotive role in the pancreatic cancer phenotype. Genetic inhibition of PKCζ reduced adherent growth, cell survival and anchorage-independent growth of human pancreatic cancer cells in vitro. Furthermore, PKCζ inhibition reduced orthotopic tumor size in vivo by inhibiting tumor cell proliferation and increasing tumor necrosis. In addition, PKCζ inhibition reduced tumor metastases in vivo, and caused a corresponding reduction in pancreatic cancer cell invasion in vitro. Signal transducer and activator of transcription 3 (STAT3) is often constitutively active in pancreatic cancer, and plays an important role in pancreatic cancer cell survival and metastasis. Interestingly, inhibition of PKCζ significantly reduced constitutive STAT3 activation in pancreatic cancer cells in vitro and in vivo. Pharmacologic inhibition of STAT3 mimicked the phenotype of PKCζ inhibition, and expression of a constitutively active STAT3 construct rescued the transformed phenotype in PKCζ-deficient cells. We conclude that PKCζ is required for pancreatic cancer cell transformed growth and invasion in vitro and tumorigenesis in vivo, and that STAT3 is an important downstream mediator of the pro-carcinogenic effects of PKCζ in pancreatic cancer cells.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0072061PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3756013PMC
May 2014

Phase I dose escalation study of the PKCι inhibitor aurothiomalate for advanced non-small-cell lung cancer, ovarian cancer, and pancreatic cancer.

Anticancer Drugs 2013 Nov;24(10):1079-83

aDepartment of Medical Oncology, Mayo Clinic, Rochester, Minnesota bDepartment of Cancer Biology, Mayo Clinic, Jacksonville, Florida cDepartment of Internal Medicine, Greater Baltimore Medical Center, Baltimore, Maryland dDepartment of Medicine, Roswell Park Cancer Institute, Buffalo, New York, USA.

Protein kinase C iota (PKCι) is overexpressed in non-small-cell lung cancer, ovarian, and pancreatic cancers, where it plays a critical role in oncogenesis. The gold compound aurothiomalate (ATM) has been shown to inhibit PKCι signaling and exerts potent antitumor activity in preclinical models. We sought to determine the maximum tolerated dose (MTD) of ATM. We conducted a phase I dose escalation trial of ATM in patients with non-small-cell lung cancer, ovarian or pancreatic cancer. Patients received ATM intramuscularly weekly for three cycles (cycle duration 4 weeks) at 25, 50, or 75 mg in a 3+3 design. The dose was not escalated for individual patients. Blood samples were analyzed for elemental gold levels. Patients were evaluated every 4 weeks for toxicity and every 8 weeks for response. Fifteen patients were enrolled in this study. Six patients were treated at 25 mg, seven at 50 mg, and two at 75 mg. There was one dose-limiting toxicity at 25 mg (hypokalemia), one at 50 mg (urinary tract infection), and none at 75 mg. There were three grade 3 hematologic toxicities. The recommended MTD of ATM is 50 mg. Patients received treatment for a median of two cycles (range 1-3). There appeared to be a dose-related accumulation of steady-state plasma concentrations of gold consistent with linear pharmacokinetics. In summary, this phase I study was successful in identifying ATM 50 mg intramuscularly weekly as the MTD. Future clinical investigations targeting PKCι are currently in progress.
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http://dx.doi.org/10.1097/CAD.0000000000000009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3937851PMC
November 2013
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