Tissue-Based Diagnostic Biomarkers of Aggressive Variant Prostate Cancer: A Narrative Review.

Prostate cancer (PC) is a common malignancy among elderly men, characterized by great heterogeneity in its clinical course, ranging from an indolent to a highly aggressive disease. The aggressive variant of prostate cancer (AVPC) clinically shows an atypical pattern of disease progression, similar to that of small cell PC (SCPC), and also shares the chemo-responsiveness of SCPC. The term AVPC does not describe a specific histologic subtype of PC but rather the group of tumors that, irrespective of morphology, show an aggressive clinical course, dictated by androgen receptor (AR) indifference. AR indifference represents an adaptive response to androgen deprivation therapy (ADT), driven by epithelial plasticity, an inherent ability of tumor cells to adapt to their environment by changing their phenotypic characteristics in a bi-directional way. The molecular profile of AVPC entails combined alterations in the tumor suppressor genes retinoblastoma protein 1 (RB1), tumor protein 53 (TP53), and phosphatase and tensin homolog (PTEN). The understanding of the biologic heterogeneity of castration-resistant PC (CRPC) and the need to identify the subset of patients that would potentially benefit from specific therapies necessitate the development of prognostic and predictive biomarkers. This review aims to discuss the possible pathophysiologic mechanisms of AVPC development and the potential use of emerging tissue-based biomarkers in clinical practice.

CUL4A Ubiquitin Ligase Is an Independent Predictor of Overall Survival in Pancreatic Adenocarcinoma.

BACKGROUND/AIM: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with dismal prognosis. Genomic instability due to defects in cell-cycle regulation/mitosis or deficient DNA-damage repair is a major driver of PDAC progression with clinical relevance. Deregulation of licensing of DNA replication leads to DNA damage and genomic instability, predisposing cells to malignant transformation. While overexpression of DNA replication-licensing factors has been reported in several human cancer types, their role in PDAC remains largely unknown. We aimed here to examine the expression and prognostic significance of the DNA replication-licensing factors chromatin licensing and DNA replication factor 1 (CDT1), cell-division cycle 6 (CDC6), minichromosome maintenance complex component 7 (MCM7) and also of the ubiquitin ligase regulator of CDT1, cullin 4A (CUL4A), in PDAC. MATERIALS AND METHODS: Expression levels of CUL4, CDT1, CDC6 and MCM7 were evaluated by immunohistochemistry in 76 formalin-fixed paraffin-embedded specimens of PDAC patients in relation to DNA-damage response marker H2AX, clinicopathological parameters and survival. We also conducted bioinformatics analysis of data from online available databases to corroborate our findings. RESULTS: CUL4A and DNA replication-licensing factors were overexpressed in patients with PDAC and expression of CDT1 positively correlated with H2AX. Expression of CUL4A and CDT1 positively correlated with lymph node metastasis. Importantly, elevated CUL4A expression was associated with reduced overall survival and was an independent indicator of poor prognosis on multivariate analysis. CONCLUSION: Our findings implicate CUL4A, CDT1, CDC6 and MCM7 in PDAC progression and identify CUL4A as an independent prognostic factor for this disease.

High-power laser lithotripsy - do we treat or harm? Histological evaluation of temperature effects in an in vivo study with thulium fiber laser.

Introduction: The aim of this study was to evaluate the possible histopathological alterations that occur in the kidneys due to a continuous temperature increase above 43°C for one hour of lithotripsy using a newly introduced thulium fiber laser (TFL). Material and methods: Two female pigs were used. After the insertion of a 9.5/11.5 ureteral access sheath, flexible ureteroscopy and laser lithotripsy for one hour were conducted. A TFL laser with a 200-µm fiber was used. The power setting used was 8 W (0.5 J × 16 Hz). A K-type thermocouple was inserted and fixed in the upper calyx of the right porcine kidney to record the temperature in the pelvicalyceal system during the laser activation. Second-look flexible nephroscopy followed by nephrectomy and pathohistological evaluation of the operated kidney was performed one week after the procedure in the first pig and 2 weeks after the surgery in the second pig. Results: Flexible nephroscopy did not reveal significant differences between the 2 porcine kidneys. Nevertheless, the histopathological report demonstrated severe alterations in the kidney of the first pig. Mild changes were reported in the kidney of the second pig. A significant improvement in inflammation and haemorrhagic lesions was demonstrated when comparing the 2 kidneys. Conclusions: The difference demonstrated between the 2 kidneys based on the histopathological report shows that the healing process is capable of improving severe to mild alterations within a one-week time frame. Two weeks after the surgery, only minor changes were observed, suggesting that even temperature increases above the threshold can be tolerated regarding renal damage.

Elucidating the role of PRMTs in prostate cancer using open access databases and a patient cohort dataset.

Protein arginine methylation is an understudied epigenetic mechanism catalyzed by enzymes known as Protein Methyltransferases of Arginine (PRMTs), while the opposite reaction is performed by Jumonji domain- containing protein 6 (JMJD6). There is increasing evidence that PRMTs are deregulated in prostate cancer (PCa). In this study, the expression of two PRMT members, PRMT2 and PRMT7 as well as JMJD6, a demethylase, was analyzed in PCa. Initially, we retrieved data from The Cancer Genome Atlas (TCGA) project and the Gene Expression Omnibus (GEO) database to explore the differential expression of various PRMT family members in patients with PCa and then applied immunohistochemistry in a patient cohort across the spectrum of PCa, including non-neoplastic prostate tissue and lymph node metastatic foci. The results from the TCGA analysis revealed that PRMT7, PRMT6 and PRMT3 expression increased while PRMT2, PRMT9 and JMJD6 levels decreased in the tumor compared to non-neoplastic prostate. Results from the GEO datasets were similar, albeit not identical with the TCGA results, with PRMT7 and PRMT3 being upregulated and PRMT2 and JMJD6 being downregulated in the tumor compared to non-neoplastic tissue in some of them. In addition, PRMT7 levels decreased with stage and grade progression in the TCGA analysis. In the patient cohort, both PRMTs and JMJD6 were overexpressed in PCa compared to non-neoplastic tissue, and nuclear PRMT2 and JMJD6 were upregulated in lymph node metastasis, too. PRMT7 and JMJD6 expression were upregulated with the progression of stage and JMJD6 was also increased with the elevation of grade. After androgen ablation therapy, nuclear expression of PRMT7 and JMJD6 were elevated compared to untreated tumors. PRMT2, PRMT7 and JMD6 were also correlated with markers of EMT and cell cycle regulators. Finally, our findings indicate that PRMTs and JMJD6 are involved in prostate cancer progression and revealed a potential interplay of PRMTs with EMT mediators, underscoring the need for therapeutic targeting of arginine methylation in prostate cancer.

Ras suppressor-1 (RSU1) exerts a tumor suppressive role with prognostic significance in lung adenocarcinoma.

Ras suppressor-1 (RSU1), originally described as a suppressor of Ras oncogenic transformation, localizes to focal adhesions interacting with the ILK-PINCH-PARVIN (IPP) complex that exerts a well-established oncogenic role in cancer. However, RSU1 implication in lung cancer is currently unknown. Our study aims to address the role of RSU1 in lung adenocarcinoma (LUADC). We here show that RSU1 protein expression by immunohistochemistry is downregulated in LUADC human tissue samples and represents a significant prognostic indicator. In silico analysis of gene chip and RNA seq data validated our findings. Depletion of RSU1 by siRNA in lung cancer cells promotes anchorage-independent cell growth, cell motility and epithelial to mesenchymal transition (EMT). Silencing of RSU1 also alters IPP complex expression in lung cancer cells. The p29 RSU1 truncated isoform is detected in lung cancer cells, and its expression is downregulated upon RSU1 silencing, whereas it is overexpressed upon ILK overexpression. These findings suggest that RSU1 exerts a tumor suppressive role with prognostic significance in LUADC.

Clearance of Metal Particles After Percutaneous Nephrolithotomy with Trilogy Lithotripter.

Purpose: To evaluate the clearance of metal particles produced and released in the pelvicaliceal system (PCS) during percutaneous nephrolithotomy (PCNL) with the use of the Swiss Lithoclast® Trilogy dual-energy (EMS Urology, Nyon, Switzerland) lithotripter. Methods: An experimental in vivo study and a clinical investigation of case series were conducted. An in vivo porcine model with two pigs for lithotripsy (after inserting artificial stones into the collecting system) and two pigs for submucosal injection of metal particles (provided by the manufacturer of Trilogy) was conducted. Porcine kidney histology analysis for metal leftovers was conducted immediately or 2 weeks after the surgery. A prospective observational study design included 10 consecutive patients treated with conventional 30F PCNL or with 22F mini-PCNL technique. Only the patients with the confirmed metal particles in the PCS during the initial PCNL and the need for additional retrograde intrarenal surgery over a period of 2-4 weeks were selected. The presence of metal particles was evaluated during the second endoscopic surgery. Results: The generated metal particles during PCNL and the submucosally injected particles were not found macroscopically 2 weeks postoperatively in porcine models. No pathologic changes such as foreign body granuloma or inflammation were found. Similarly, no metal particles were observed during the second look endoscopy (n = 10). Conclusion: Metal particles observed endoscopically using the Trilogy lithotripter are cleared with no pathologic evidence of tissue damage from the metal particles 2 weeks after the procedure. Thus, the intraoperative release of any particle by the Trilogy lithotripter should not raise any safety concerns.

Cdt1 overexpression drives colorectal carcinogenesis through origin overlicensing and DNA damage.

Chromatin licensing and DNA replication factor 1 (CDT1), a protein of the pre-replicative complex, is essential for loading the minichromosome maintenance complex (MCM) helicases onto the origins of DNA replication. While several studies have shown that dysregulation of CDT1 expression causes re-replication and DNA damage in cell lines, and CDT1 is highly expressed in several human cancers, whether CDT1 deregulation is sufficient to enhance tumorigenesis in vivo is currently unclear. To delineate its role in vivo, we overexpressed Cdt1 in the mouse colon and induced carcinogenesis using azoxymethane/dextran sodium sulfate (AOM/DSS). Here, we show that mice overexpressing Cdt1 develop a significantly higher number of tumors with increased tumor size, and more severe dysplastic changes (high-grade dysplasia), compared with control mice under the same treatment. These tumors exhibited an increased growth rate, while cells overexpressing Cdt1 loaded greater amounts of Mcm2 onto chromatin, demonstrating origin overlicensing. Adenomas overexpressing Cdt1 showed activation of the DNA damage response (DDR), apoptosis, formation of micronuclei, and chromosome segregation errors, indicating that aberrant expression of Cdt1 results in increased genomic and chromosomal instability in vivo, favoring cancer development. In line with these results, high-level expression of CDT1 in human colorectal cancer tissue specimens and colorectal cancer cell lines correlated significantly with increased origin licensing, activation of the DDR, and microsatellite instability (MSI). © 2022 The Pathological Society of Great Britain and Ireland.

The Role of Hippo Signaling Pathway and ILK in the Pathophysiology of Human Hypertrophic Scars and Keloids: An Immunohistochemical Investigation.

BACKGROUND: Keloids and hypertrophic scars are characterized by abnormal fibroblast activation and proliferation. While their molecular pathogenesis remains unclear, myofibroblasts have been associated with their development. Hippo pathway effectors YAP/TAZ promote cell proliferation and matrix stiffening. Integrin-linked kinase (ILK), a central component of focal adhesions that mediates cell-matrix interactions, has been linked to tissue repair and fibrosis. The aim of this study was to investigate the expression of key Hippo pathway molecules and ILK in hypertrophic scars and keloids. METHODS: YAP/TAZ, TEAD4, ILK and a-SMA expression were evaluated by immunohistochemistry in keloids (n = 55), hypertrophic scars (n = 38) and normal skin (n = 14). RESULTS: The expression of YAP/TAZ, TEAD4, ILK and a-SMA was higher in fibroblasts of keloids compared to hypertrophic scars while negative in normal skin. There was a significant positive correlation between the expression of ILK and Hippo pathway effectors. CONCLUSIONS: Our results suggest that the deregulation of Hippo signaling and ILK are implicated in keloid and hypertrophic scar formation.

A Molecular Lateral Flow Assay for SARS-CoV-2 Quantitative Detection.

Since the onset of the SARS-CoV-2 pandemic, several COVID-19 detection methods, both commercially available and in the lab, have been developed using different biomolecules as analytes and different detection and sampling methods with high analytical performance. Developing novel COVID-19 detection assays is an exciting research field, as rapid accurate diagnosis is a valuable tool to control the current pandemic, and also because the acquired knowledge can be deployed for facing future infectious outbreaks. We here developed a novel gold-nanoparticle-based nucleic acid lateral flow assay for the rapid, visual, and quantitative detection of SARS-CoV-2. Our method was based on the use of a DNA internal standard (competitor) for quantification and involved RT-PCR, the hybridization of biotinylated PCR products to specific oligonucleotide probes, and detection with a dual lateral flow assay using gold nanoparticles conjugated to an anti-biotin antibody as reporters. The developed test allowed for rapid detection by the naked eye and the simultaneous quantification of SARS-CoV-2 in nasopharyngeal swabs with high specificity, detectability, and repeatability. This novel molecular strip test for COVID-19 detection represents a simple, cost-effective, and accurate rapid test that is very promising to be used as a future diagnostic tool.

Focal adhesion proteins in hepatocellular carcinoma: RSU1 a novel tumour suppressor with prognostic significance.

AIM: Hepatocellular carcinoma (HCC) is a common cause a cancer-related death. Focal adhesions (FAs) represent multiprotein complexes at integrin-mediated cell-extracellular matrix adhesion sites that orchestrate vital cellular functions. The heterotrimeric ILK-PINCH-PARVB (IPP) complex, RSU1, a PINCH binding protein and CTEN, a member of the tensin family of proteins exert a critical role in FAs, where they regulate important cancer related functions such as cell adhesion, migration, proliferation and survival. Previous studies implicate these FA proteins in liver pathophysiology but their detailed role in human HCC is not fully understood. Here in we investigated expression and function of IPP, RSU1 and CTEN in human HCC. METHODS: The expression of focal adhesion proteins was studied in human HCC by immunohistochemistry in relation to clinicopathological parameters, previous studied genomic instability markers and patient's survival. Effects on cell proliferation and FA proteins expression upon ILK inhibition and RSU1 silencing were also investigated in HCC in vitro. RESULTS: IPP complex and CTEN proteins are overexpressed while RSU1 expression is decreased in human HCC. CTEN expression correlates with reduced patients' survival while RSU1 represents an independent favorable prognostic indicator in human HCC. Nuclear ILK expression correlates with markers of genomic instability. Pharmacological targeting of ILK suppresses, while RSU1 silencing promotes cell growth of HCC cells in vitro, while in both experimental conditions expression and/or localization of focal adhesion proteins is deregulated. CONCLUSION: Our results suggest that FA signaling is implicated in hepatocellular carcinogenesis with prognostic significance. RSU1 seems to exert tumor suppressive functions in HCC and represents a novel favorable prognostic indicator.

Intrinsic neural stem cell properties define brain hypersensitivity to genotoxic stress.

Impaired replication has been previously linked to growth retardation and microcephaly; however, why the brain is critically affected compared with other organs remains elusive. Here, we report the differential response between early neural progenitors (neuroepithelial cells [NECs]) and fate-committed neural progenitors (NPs) to replication licensing defects. Our results show that, while NPs can tolerate altered expression of licensing factors, NECs undergo excessive replication stress, identified by impaired replication, increased DNA damage, and defective cell-cycle progression, leading eventually to NEC attrition and microcephaly. NECs that possess a short G1 phase license and activate more origins than NPs, by acquiring higher levels of DNA-bound MCMs. In vivo G1 shortening in NPs induces DNA damage upon impaired licensing, suggesting that G1 length correlates with replication stress hypersensitivity. Our findings propose that NECs possess distinct cell-cycle characteristics to ensure fast proliferation, although these inherent features render them susceptible to genotoxic stress.

The deadly cross-talk between Hippo pathway and epithelial-mesenchymal transition (EMT) in cancer.

Hippo signaling pathway is an evolutionarily conserved network that regulates organ size growth and tissue regeneration. Hippo signaling dysfunction results in uncontrolled cell proliferation and influences cell differentiation. Aberrant Hippo pathway signaling is implicated in cancer progression, by promoting cell proliferation, cancer stem cell properties, chemoresistance and metastatic capacity. Epithelial-mesenchymal transition (EMT) is also well known to be implicated in carcinogenesis. Loss of cell polarity, disruption of cell-cell junctions and cytoskeletal remodeling are essential during EMT. At the same time, signals related to intercellular contact, cell-extracellular matrix contact, polarity and mechanotransduction are included in the list of regulatory inputs into Hippo pathway. Therefore, the emerging association between Hippo pathway and EMT in cancer is not surprising. Recent studies have begun to unravel the mechanisms of interaction between Hippo signaling pathway and EMT. In this review, we describe the existing evidence of cross talk between Hippo signaling pathway key molecules and the process of EMT, with emphasis on the role of Hippo-EMT interplay in cancer.

Rapid Multiplex Strip Test for the Detection of Circulating Tumor DNA Mutations for Liquid Biopsy Applications.

In the era of personalized medicine, molecular profiling of patient tumors has become the standard practice, especially for patients with advanced disease. Activating point mutations of the KRAS proto-oncogene are clinically relevant for many types of cancer, including colorectal cancer (CRC). While several approaches have been developed for tumor genotyping, liquid biopsy has been gaining much attention in the clinical setting. Analysis of circulating tumor DNA for genetic alterations has been challenging, and many methodologies with both advantages and disadvantages have been developed. We here developed a gold nanoparticle-based rapid strip test that has been applied for the first time for the multiplex detection of KRAS mutations in circulating tumor DNA (ctDNA) of CRC patients. The method involved ctDNA isolation, PCR-amplification of the KRAS gene, multiplex primer extension (PEXT) reaction, and detection with a multiplex strip test. We have optimized the efficiency and specificity of the multiplex strip test in synthetic DNA targets, in colorectal cancer cell lines, in tissue samples, and in blood-derived ctDNA from patients with advanced colorectal cancer. The proposed strip test achieved rapid and easy multiplex detection (normal allele and three major single-point mutations) of the clinically relevant KRAS mutations in ctDNA in blood samples of CRC patients with high specificity and repeatability. This multiplex strip test represents a minimally invasive, rapid, low-cost, and promising diagnostic tool for the detection of clinically relevant mutations in cancer patients.

The protein arginine methyltransferases (PRMTs) PRMT1 and CARM1 as candidate epigenetic drivers in prostate cancer progression.

ABSTRACT: Epigenetic changes are implicated in prostate cancer (PCa) progression and resistance to therapy. Arginine residue methylation is an understudied histone post-translational modification that is increasingly associated with cancer progression and is catalyzed by enzymes called protein arginine methyltransferases (PRMTs). The molecular consequences of aberrant expression of PRMTs in PCa and the relationship between PRMTs and PCa progression are largely unknown. Using immunohistochemistry, we examined the expression of PRMT1 and CARM1, two of the best-studied PRMTs, in 288 patients across the spectrum of PCa and correlated them with markers of androgen receptor (AR) signaling, and milestones of carcinogenesis. Our findings indicate that PRMT1 and CARM1 are upregulated early in PCa progression, and that CARM1 is further upregulated after therapy. In addition, a correlation of CARM1 with AR post-translational modifications was noted in the setting of therapy resistance, highlighting CARM1 as one of the adaptation mechanisms of PCa cells in an androgen-depleted environment. Finally, CARM1 correlated with markers of cell cycle regulation, and both CARM1 and PRMT1 correlated with markers of epithelial-to-mesenchymal transition signaling. Taken together these findings indicate that an epigenetic network drives PCa progression through enhancement of milestone pathways including AR signaling, the cell cycle, and epithelial-to-mesenchymal transition.

Liquid biopsy genotyping by a simple lateral flow strip assay with visual detection.

Liquid biopsy, as a minimally invasive method that allows real-time monitoring of the tumor genome, represents a competing approach for cancer diagnosis, prognosis and therapy decision making. Liquid biopsy in cancer patients mainly includes analysis of circulating tumor cells (CTC) and cell-free circulating tumor DNA (ctDNA). ctDNA is the tumor-derived fraction of the cell-free DNA present in the blood. ctDNA is detected based on cancer-specific genomic aberrations (mainly mutations) and represents a challenging analyte due to high fragmentation and low concentration. Several methodologies have been developed for ctDNA analysis in cancer patients but many of these technologies are too time-intensive, complicated and expensive for implementation in diagnostic testing. Herein, we developed a novel lateral flow strip assay for mutational analysis of ctDNA in blood samples and visual detection that is based on gold nanoparticles as reporters. As a model, common single-point mutations of the KRAS gene, related to colorectal cancer (CRC), have been selected for method development. The proposed DNA biosensor has been successfully applied for the detection of three KRAS mutations (KRAS G12D/A/V), along with the wild-type KRAS gene in synthetic DNA targets, cancer cell lines and cfDNA from blood samples of healthy individuals and CRC patients. The main advantages of the proposed lateral flow assay are simplicity, rapid analysis time (∼10 min) and visual detection without the requirement of special instrumentation. The assay is also cost-effective with high detectability, specificity and reproducibility and has the potential to be used as a portable and universal device. In conclusion, the proposed assay offers a rapid diagnostic strip test for visual genotyping, as an alternative approach for liquid biopsy applications.

Integrin-Linked-Kinase Overexpression Is Implicated in Mechanisms of Genomic Instability in Human Colorectal Cancer.

BACKGROUND: Genomic instability is a hallmark of cancer cells contributing to tumor development and progression. Integrin-linked kinase (ILK) is a focal adhesion protein with well-established role in carcinogenesis. We have previously shown that ILK overexpression is critically implicated in human colorectal cancer (CRC) progression. In light of the recent findings that ILK regulates centrosomes and mitotic spindle formation, we aimed to determine its implication in mechanisms of genomic instability in human CRC. METHODS: Association of ILK expression with markers of genomic instability (micronuclei formation, nucleus size, and intensity) was investigated in diploid human colon cancer cells HCT116 upon ectopic ILK overexpression, by immunofluorescence and in human CRC samples by Feulgen staining. We also evaluated the role of ILK in mitotic spindle formation, by immunofluorescence, in HCT116 cells upon inhibition and overexpression of ILK. Finally, we evaluated association of ILK overexpression with markers of DNA damage (p-H2AX, p-ATM/ATR) in human CRC tissue samples by immunohistochemistry and in ILK-overexpressing cells by immunofluorescence. RESULTS: We showed that ILK overexpression is associated with genomic instability markers in human colon cancer cells and tissues samples. Aberrant mitotic spindles were observed in cells treated with specific ILK inhibitor (QLT0267), while ILK-overexpressing cells failed to undergo nocodazole-induced mitotic arrest. ILK overexpression was also associated with markers of DNA damage in HCT116 cells and human CRC tissue samples. CONCLUSIONS: The above findings indicate that overexpression of ILK is implicated in mechanisms of genomic instability in CRC suggesting a novel role of this protein in cancer.

H3K4 Methylation Status and Lysine Specific Methyltransferase KMT2C Expression Correlate with Prognosis in Lung Adenocarcinoma.

BACKGROUND: Genetic events cannot account for the complexity of human carcinogenesis alone. Mutations of epigenetic regulators and aberrations of their expression patterns have been detected in various human malignancies. Methylation of histone H3 at lysine 4 (H3K4me), is an evolutionarily conserved histone modification that marks regions of active transcription and regulates cell growth, migration, and invasion. The MLL/KMT2 family of histone methyltransferases specifically methylate histone H3 at lysine 4. OBJECTIVE: The aim of this study was to explore the role of KMT2C/MLL3 as well as key histone modification activating markers, such as H3K4me2 and H3K4me3 in a cohort of surgically resected human lung adenocarcinomas in an effort to reveal possible biomarkers for lung adenocarcinoma diagnosis and prognosis and potential therapeutic targets. METHOD: The immunohistochemical expression of KMT2C/MLL3, H3K4me2 and H3K4me3 was analyzed in formalin fixed paraffin embedded tissue from 96 patients with lung adenocarcinoma. Results were associated with clinicopathologic parameters and patient's prognosis. RESULTS: Nuclear expression of KMT2C/MLL3 in epithelial cells was independently associated with shorter overall survival. Cytoplasmic H3K4me2 expression was associated withT stage and nuclear H3K4me2 expression was associated with female gender and patients' prognosis. The latter association persisted after multivariate analysis. No association was found between H3K4me3 expression and clinicopathological data or disease outcome in our cohort of patients. CONCLUSION: These results suggest that the pattern of histone modifications and KMT2C/MLL3 expression can be used as an independent prognostic factor in lung adenocarcinoma, revealing that chromatin remodeling is criticallyinvolved in cancer progression.

Integrin-linked kinase (ILK) regulates KRAS, IPP complex and Ras suppressor-1 (RSU1) promoting lung adenocarcinoma progression and poor survival.

Integrin-linked kinase (ILK) forms a heterotrimeric protein complex with PINCH and PARVIN (IPP) in Focal Adhesions (FAs) that acts as a signaling platform between the cell and its microenvironment regulating important cancer-related functions. We aimed to elucidate the role of ILK in lung adenocarcinoma (LUADC) focusing on a possible link with KRAS oncogene. We used immunohistochemistry on human tissue samples and KRAS-driven LUADC in mice, analysis of large scale publicly available RNA sequencing data, ILK overexpression and pharmacological inhibition as well as knockdown of KRAS in lung cancer cells. ILK, PINCH1 and PARVB (IPP) proteins are overexpressed in human LUADC and KRAS-driven LUADC in mice representing poor prognostic indicators. Genes implicated in ILK signaling are significantly enriched in KRAS-driven LUADC. Silencing of KRAS, as well as, overexpression and pharmacological inhibition of ILK in lung cancer cells provide evidence of a two-way association between ILK and KRAS. Upregulation of PINCH, PARVB and Ras suppressor-1 (RSU1) expression was demonstrated in ILK overexpressing lung cancer cells in addition to a significant positive correlation between these factors in tissue samples, while KRAS silencing downregulates IPP and RSU1. Pharmacological inhibition of ILK in KRAS mutant lung cancer cells suppresses cell growth, migration, EMT and increases sensitivity to platinum-based chemotherapy. ILK promotes an aggressive lung cancer phenotype with prognostic and therapeutic value through functions that involve KRAS, IPP complex and RSU1, rendering ILK a promising biomarker and therapeutic target in lung adenocarcinoma.

Expression and prognostic significance of YAP, TAZ, TEAD4 and p73 in human laryngeal cancer.

OBJECTIVES: The Hippo signaling pathway plays a critical role in organ size control and tissue homeostasis and its perturbation is associated with tumorigenesis. YAP (Yes associated protein) and TAZ (transcriptional co-activator with PDZ- binding motif) are the major nuclear effectors of the Hippo pathway interacting with TEADs (TEA domain) and p73 transcriptional factors to regulate gene expression. Altered expression of the above proteins promotes tumor initiation, progression and metastasis in a variety of cancer types. This study addresses their expression and prognostic significance in human laryngeal carcinoma. METHODS: Protein expression of YAP, TAZ, TEAD4 and p73 was examined by immunohistochemistry in 121 human laryngeal squamous cell carcinomas. Correlations with clinicopathological data and survival were evaluated. RESULTS: All proteins were overexpressed in human laryngeal carcinomas compared to non-neoplastic adjacent epithelium. High expression of YAP, TAZ, TEAD4 and p73 correlated significantly with high grade, advanced stage, supraglottic location of tumor, nodal metastases and recurrence. Furthermore, high expression of all proteins was significantly associated with poor overall and disease- free survival. p73 expression proved to be an independent predictive factor of survival and YAP expression proved to be an independent predictive factor of disease recurrence. CONCLUSIONS: Deregulation of the expression of the Hippo pathway proteins is implicated in human laryngeal carcinogenesis and YAP and p73 have prognostic significance in the outcome of the disease.