DYRK1A in cancer: good or evil? : Defining properties of DYRK1A kinase as a novel tumor driver

DYRK (dual-specificity tyrosine-regulated kinases) is an evolutionary conserved family of protein kinases involved in the regulation of cellular processes, such as proliferation and survival, which play pivotal role in tumor development. In this Thesis work, the potential role of DYRK genes as tumor drivers has been explored through an extensive analysis of The Cancer Genome Atlas data. DYRKs were found altered in tumor samples at different levels. In particular, the dosage sensitive member DYRK1A emerged as a potential tumor driver. Functional screens on DYRK1A cancer somatic mutations showed that most of the mutants analyzed have a strong impact on the catalytic activity and/or stability of the protein, suggesting that DYRK1A loss-of-function is positively selected in cancer. Reversion, by a CRISPR/Cas9 strategy, of a DYRK1A mutant allele to wt in an endometrial cancer cell line strongly impaired tumor cell growth. The DYRK1A tumor suppressive role was confirmed in vivo using mouse tumor xenografts. Finally, an integrated transcriptomic, proteomic and phospho-proteomic analysis has uncovered potential molecular mechanisms underlying the DYRK1A-mediated tumor driver function.Las proteínas quinasas DYRK (dual-specificity tyrosine-regulated kinases) son una familia evolutivamente conservada que participan en la regulación de procesos celulares con funciones fundamentales en la transformación maligna. En este trabajo de tesis, el papel de los genes DYRK como conductores (drivers) de tumores se ha explorado mediante un extenso análisis de los datos de The Cancer Genome Atlas. El análisis encontró alterados los genes DYRK en muestras de tumores a diferentes niveles, e identificó al miembro de la familia DYRK1A como un potencial gen conductor en un grupo de tumores. Mediante ensayos funcionales se ha demostrado que las mutaciones somáticas de DYRK1A en cáncer investigadas tienen un fuerte impacto en la actividad catalítica y/o la estabilidad de la proteína, lo que sugiere que la perdida de función de DYRK1A se selecciona positivamente en la célula tumoral. La corrección, mediante la técnica CRISPR/Cas9, de una línea celular de cáncer de endometrio mutada en DYRK1A tiene un fuerte impacto en el crecimiento de estas células tumorales, sugiriendo un papel para DYRK1A como supresor de tumores en cáncer de endometrio, que se ha confirmado in vivo en modelos tumorales de xenoinjerto en ratón. Finalmente, un análisis transcriptómico, proteómico y fosfo-proteómico integrado ha desvelado posibles mecanismos moleculares que participan en la función de DYRK1A como supresor tumoral

Oxidative stress and inflammation cause auditory system damage via glial cell activation and dysregulated expression of gap junction proteins in an experimental model of styrene-induced oto/neurotoxicity

BackgroundRedox imbalance and inflammation have been proposed as the principal mechanisms of damage in the auditory system, resulting in functional alterations and hearing loss. Microglia and astrocytes play a crucial role in mediating oxidative/inflammatory injury in the central nervous system; however, the role of glial cells in the auditory damage is still elusive.ObjectivesHere we investigated glial-mediated responses to toxic injury in peripheral and central structures of the auditory pathway, i.e., the cochlea and the auditory cortex (ACx), in rats exposed to styrene, a volatile compound with well-known oto/neurotoxic properties.MethodsMale adult Wistar rats were treated with styrene (400 mg/kg daily for 3 weeks, 5/days a week). Electrophysiological, morphological, immunofluorescence and molecular analyses were performed in both the cochlea and the ACx to evaluate the mechanisms underlying styrene-induced oto/neurotoxicity in the auditory system.ResultsWe showed that the oto/neurotoxic insult induced by styrene increases oxidative stress in both cochlea and ACx. This was associated with macrophages and glial cell activation, increased expression of inflammatory markers (i.e., pro-inflammatory cytokines and chemokine receptors) and alterations in connexin (Cxs) and pannexin (Panx) expression, likely responsible for dysregulation of the microglia/astrocyte network. Specifically, we found downregulation of Cx26 and Cx30 in the cochlea, and high level of Cx43 and Panx1 in the ACx.ConclusionsCollectively, our results provide novel evidence on the role of immune and glial cell activation in the oxidative/inflammatory damage induced by styrene in the auditory system at both peripheral and central levels, also involving alterations of gap junction networks. Our data suggest that targeting glial cells and connexin/pannexin expression might be useful to attenuate oxidative/inflammatory damage in the auditory system

Study of breast cancer incidence in patients of lymphangioleiomyomatosis

The online version of this article (doi:10.1007/s10549-016-3737-8) contains supplementary material, which is available to authorized user

Lymphangioleiomyomatosis biomarkers linked to lung metastatic potential and cell stemness

Lymphangioleiomyomatosis (LAM) is a rare lung-metastasizing neoplasm caused by the proliferation of smooth muscle-like cells that commonly carry loss-of-function mutations in either the tuberous sclerosis complex 1 or 2 (TSC1 or TSC2) genes. While allosteric inhibition of the mechanistic target of rapamycin (mTOR) has shown substantial clinical benefit, complementary therapies are required to improve response and/or to treat specific patients. However, there is a lack of LAM biomarkers that could potentially be used to monitor the disease and to develop other targeted therapies. We hypothesized that the mediators of cancer metastasis to lung, particularly in breast cancer, also play a relevant role in LAM. Analyses across independent breast cancer datasets revealed associations between low TSC1/2 expression, altered mTOR complex 1 (mTORC1) pathway signaling, and metastasis to lung. Subsequently, immunohistochemical analyses of 23 LAM lesions revealed positivity in all cases for the lung metastasis mediators fascin 1 (FSCN1) and inhibitor of DNA binding 1 (ID1). Moreover, assessment of breast cancer stem or luminal progenitor cell biomarkers showed positivity in most LAM tissue for the aldehyde dehydrogenase 1 (ALDH1), integrin-ß3 (ITGB3/CD61), and/or the sex-determining region Y-box 9 (SOX9) proteins. The immunohistochemical analyses also provided evidence of heterogeneity between and within LAM cases. The analysis of Tsc2-deficient cells revealed relative over-expression of FSCN1 and ID1; however, Tsc2-deficient cells did not show higher sensitivity to ID1-based cancer inhibitors. Collectively, the results of this study reveal novel LAM biomarkers linked to breast cancer metastasis to lung and to cell stemness, which in turn might guide the assessment of additional or complementary therapeutic opportunities for LAM

Colorectal Cancer Stage at Diagnosis Before vs During the COVID-19 Pandemic in Italy

IMPORTANCE Delays in screening programs and the reluctance of patients to seek medical attention because of the outbreak of SARS-CoV-2 could be associated with the risk of more advanced colorectal cancers at diagnosis. OBJECTIVE To evaluate whether the SARS-CoV-2 pandemic was associated with more advanced oncologic stage and change in clinical presentation for patients with colorectal cancer. DESIGN, SETTING, AND PARTICIPANTS This retrospective, multicenter cohort study included all 17 938 adult patients who underwent surgery for colorectal cancer from March 1, 2020, to December 31, 2021 (pandemic period), and from January 1, 2018, to February 29, 2020 (prepandemic period), in 81 participating centers in Italy, including tertiary centers and community hospitals. Follow-up was 30 days from surgery. EXPOSURES Any type of surgical procedure for colorectal cancer, including explorative surgery, palliative procedures, and atypical or segmental resections. MAIN OUTCOMES AND MEASURES The primary outcome was advanced stage of colorectal cancer at diagnosis. Secondary outcomes were distant metastasis, T4 stage, aggressive biology (defined as cancer with at least 1 of the following characteristics: signet ring cells, mucinous tumor, budding, lymphovascular invasion, perineural invasion, and lymphangitis), stenotic lesion, emergency surgery, and palliative surgery. The independent association between the pandemic period and the outcomes was assessed using multivariate random-effects logistic regression, with hospital as the cluster variable. RESULTS A total of 17 938 patients (10 007 men [55.8%]; mean [SD] age, 70.6 [12.2] years) underwent surgery for colorectal cancer: 7796 (43.5%) during the pandemic period and 10 142 (56.5%) during the prepandemic period. Logistic regression indicated that the pandemic period was significantly associated with an increased rate of advanced-stage colorectal cancer (odds ratio [OR], 1.07; 95%CI, 1.01-1.13; P = .03), aggressive biology (OR, 1.32; 95%CI, 1.15-1.53; P < .001), and stenotic lesions (OR, 1.15; 95%CI, 1.01-1.31; P = .03). CONCLUSIONS AND RELEVANCE This cohort study suggests a significant association between the SARS-CoV-2 pandemic and the risk of a more advanced oncologic stage at diagnosis among patients undergoing surgery for colorectal cancer and might indicate a potential reduction of survival for these patients

Study of breast cancer incidence in patients of lymphangioleiomyomatosis

Molecular evidence has linked the pathophysiology of lymphangioleiomyomatosis (LAM) to that of metastatic breast cancer. Following on this observation, we assessed the association between LAM and subsequent breast cancer. An epidemiological study was carried out using three LAM country cohorts, from Japan, Spain, and the United Kingdom. The number of incident breast cancer cases observed in these cohorts was compared with the number expected on the basis of the country-specific incidence rates for the period 2000–2014. Immunohistochemical studies and exome sequence analysis were performed in two and one tumors, respectively. All cohorts revealed breast cancer standardized incidence ratios (SIRs) ≥ 2.25. The combined analysis of all cases or restricted to pre-menopausal age groups revealed significantly higher incidence of breast cancer: SIR = 2.81, 95 % confidence interval (CI) = 1.32–5.57, P = 0.009; and SIR = 4.88, 95 % CI = 2.29–9.99, P = 0.0007, respectively. Immunohistochemical analyses showed positivity for known markers of lung metastatic potential. This study suggests the existence of increased breast cancer risk among LAM patients. Prospective studies may be warranted to corroborate this result, which may be particularly relevant for pre-menopausal women with LAM. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10549-016-3737-8) contains supplementary material, which is available to authorized users

DYRK1A in cancer: good or evil? : Defining properties of DYRK1A kinase as a novel tumor driver

DYRK (dual-specificity tyrosine-regulated kinases) is an evolutionary conserved family of protein kinases involved in the regulation of cellular processes, such as proliferation and survival, which play pivotal role in tumor development. In this Thesis work, the potential role of DYRK genes as tumor drivers has been explored through an extensive analysis of The Cancer Genome Atlas data. DYRKs were found altered in tumor samples at different levels. In particular, the dosage sensitive member DYRK1A emerged as a potential tumor driver. Functional screens on DYRK1A cancer somatic mutations showed that most of the mutants analyzed have a strong impact on the catalytic activity and/or stability of the protein, suggesting that DYRK1A loss-of-function is positively selected in cancer. Reversion, by a CRISPR/Cas9 strategy, of a DYRK1A mutant allele to wt in an endometrial cancer cell line strongly impaired tumor cell growth. The DYRK1A tumor suppressive role was confirmed in vivo using mouse tumor xenografts. Finally, an integrated transcriptomic, proteomic and phospho-proteomic analysis has uncovered potential molecular mechanisms underlying the DYRK1A-mediated tumor driver function.Las proteínas quinasas DYRK (dual-specificity tyrosine-regulated kinases) son una familia evolutivamente conservada que participan en la regulación de procesos celulares con funciones fundamentales en la transformación maligna. En este trabajo de tesis, el papel de los genes DYRK como conductores (drivers) de tumores se ha explorado mediante un extenso análisis de los datos de The Cancer Genome Atlas. El análisis encontró alterados los genes DYRK en muestras de tumores a diferentes niveles, e identificó al miembro de la familia DYRK1A como un potencial gen conductor en un grupo de tumores. Mediante ensayos funcionales se ha demostrado que las mutaciones somáticas de DYRK1A en cáncer investigadas tienen un fuerte impacto en la actividad catalítica y/o la estabilidad de la proteína, lo que sugiere que la perdida de función de DYRK1A se selecciona positivamente en la célula tumoral. La corrección, mediante la técnica CRISPR/Cas9, de una línea celular de cáncer de endometrio mutada en DYRK1A tiene un fuerte impacto en el crecimiento de estas células tumorales, sugiriendo un papel para DYRK1A como supresor de tumores en cáncer de endometrio, que se ha confirmado in vivo en modelos tumorales de xenoinjerto en ratón. Finalmente, un análisis transcriptómico, proteómico y fosfo-proteómico integrado ha desvelado posibles mecanismos moleculares que participan en la función de DYRK1A como supresor tumoral

Morphological analysis of neurons: Automatic identification of elongations

Our study is focused on the development of a new method for the automatic analysis of cell images. We focused on neurons (cells line SH-SY5Y) treated/untreated with ultrasound and stained with Haematoxylin-Eosin. The aim of the algorithm is the automatic detection of the cell body as well as the determination of the number and the length of neuron elongations. Starting point of the algorithm was the convolution of an image with a bank of rotating Gaussian kernels and the construction of a module map. Then several strategies were implemented to detect cell bodies and to detect and extract data about cell elongations. We have also realized a graphical user interface allowing the loading, saving and processing of images. Results show that this method is able to properly and efficiently detect cell contours and elongations. The automated evaluation is in strong agreement with manual evaluation performed by an expert operator, with an average error of 11% with most parameter combinations. This tool constitutes an important support in biological research activities, where operators need to analyze a large number of images to investigate about cell morphology before and after a treatment