A novel program of infiltrative control in astrocytomas: ADAM23 depletion promotes cell invasion by activating γ-secretase complex.

Background: Infiltration is a life-threatening growth pattern in malignant astrocytomas and a significant cause of therapy resistance. It results in the tumor cell spreading deeply into the surrounding brain tissue, fostering tumor recurrence and making complete surgical resection impossible. We need to thoroughly understand the mechanisms underlying diffuse infiltration to develop effective therapies. Methods: We integrated in vitro and in vivo functional assays, RNA sequencing, clinical, and expression information from public data sets to investigate the role of ADAM23 expression coupling astrocytoma's growth and motility. Results: ADAM23 downregulation resulted in increased infiltration, reduced tumor growth, and improved overall survival in astrocytomas. Additionally, we show that ADAM23 deficiency induces γ-secretase (GS) complex activity, contributing to the production and deposition of the Amyloid-ß and release of NICD. Finally, GS ablation in ADAM23-low astrocytomas induced a significant inhibitory effect on the invasive programs. Conclusions: Our findings reveal a role for ADAM23 in regulating the balance between cell proliferation and invasiveness in astrocytoma cells, proposing GS inhibition as a therapeutic option in ADAM23 low-expressing astrocytomas.

l-carvone decreases breast cancer cells adhesion, migration, and invasion by suppressing FAK activation.

Breast cancer is one of the most common types of cancer in the world and current therapeutic strategies present severe drawbacks. l-carvone (CRV), a monoterpene found in Mentha spicata (spearmint), has been reported to have potent anti-inflammatory activity. Here, we examined the role of CRV in breast cancer cell adhesion, migration and invasion in vitro and how this component could suppress the growth of Ehrlich carcinoma-bearing mice. In vivo, treatment with CRV significantly decreased tumor growth, increased tumor necrosis area, and reduced the expression of VEGF and HIF-1α in Ehrlich carcinoma-bearing mice. Furthermore, the anticancer efficacy of CRV was similar to currently used chemotherapy (Methotrexate), and the combination of CRV with MTX potentiated the chemotherapy effects. Further mechanistic investigation in vitro revealed that CRV modulates the interaction of breast cancer cells with the extracellular matrix (ECM) by disrupting focal adhesion, which was shown by scanning electron microscopy (SEM) and immunofluorescence. Moreover, CRV caused a decrease in ß1-integrin expression and inhibited focal adhesion kinase (FAK) activation. FAK is one of the most important downstream activators of several metastatic processes, including MMP-2 mediated invasion and HIF-1α/VEGF angiogenesis stimulus, both of which were found to be reduced in MDA-MB-231 cells exposed to CRV. Our results provide new insight about targeting ß1-integrin/FAK signaling pathway with CRV, which could be a new potential agent in the treatment of breast cancer.

Review of anticancer activity of monoterpenoids: Geraniol, nerol, geranial and neral.

In recent years head-to-tail monoterpene geraniol has been widely explored as a potential anticancer agent. Natural analogs like alcohol nerol, aldehydes geranial, and neral have been investigated. We explored the synergism of these terpenes with clinically and non-clinically used compounds as potential candidates for treating different types of cancer. Promising activity for these compounds has also inspired new analog syntheses. The anticancer potential of these compounds is described in this review.

A Key Pathway to Cancer Resilience: The Role of Autophagy in Glioblastomas.

There are no effective strategies for the successful treatment of glioblastomas (GBM). Current therapeutic modalities effectively target bulk tumor cells but leave behind marginal GBM cells that escape from the surgical margins and radiotherapy field, exhibiting high migratory phenotype and resistance to all available anti-glioma therapies. Drug resistance is mostly driven by tumor cell plasticity: a concept associated with reactivating transcriptional programs in response to adverse and dynamic conditions from the tumor microenvironment. Autophagy, or "self-eating", pathway is an emerging target for cancer therapy and has been regarded as one of the key drivers of cell plasticity in response to energy demanding stress conditions. Many studies shed light on the importance of autophagy as an adaptive mechanism, protecting GBM cells from unfavorable conditions, while others recognize that autophagy can kill those cells by triggering a non-apoptotic cell death program, called 'autophagy cell death' (ACD). In this review, we carefully analyzed literature data and conclude that there is no clear evidence indicating the presence of ACD under pathophysiological settings in GBM disease. It seems to be exclusively induced by excessive (supra-physiological) stress signals, mostly from in vitro cell culture studies. Instead, pre-clinical and clinical data indicate that autophagy is an emblematic example of the 'dark-side' of a rescue pathway that contributes profoundly to a pro-tumoral adaptive response. From a standpoint of treating the real human disease, only combinatorial therapy targeting autophagy with cytotoxic drugs in the adjuvant setting for GBM patients, associated with the development of less toxic and more specific autophagy inhibitors, may inhibit adaptive response and enhance the sensibility of glioma cells to conventional therapies.

FRC-QE: a robust and comparable 3D microscopy image quality metric for cleared organoids.

SUMMARY: Here, we propose Fourier ring correlation-based quality estimation (FRC-QE) as a new metric for automated image quality estimation in 3D fluorescence microscopy acquisitions of cleared organoids that yields comparable measurements across experimental replicates, clearing protocols and works for different microscopy modalities. AVAILABILITY AND IMPLEMENTATION: FRC-QE is written in ImgLib2/Java and provided as an easy-to-use and macro-scriptable plugin for Fiji. Code, documentation, sample images and further information can be found under https://github.com/PreibischLab/FRC-QE. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Metformin as an Alternative Radiosensitizing Agent to 5-Fluorouracil During Neoadjuvant Treatment for Rectal Cancer.

BACKGROUND: Neoadjuvant chemoradiation for locally advanced rectal cancer combining 5-fluorouracil with radiation increases tumor regression compared with radiation alone. However, it occurs at the cost of significant treatment-related toxicity. Patients with rectal cancer using metformin have been associated with improved response to radiotherapy. OBJECTIVE: The purpose of this study was to evaluate the radiosensitizing effects of metformin in vitro and in vivo and compare it with a standard combination of radiation/5-fluorouracil. DESIGN: Colorectal cancer cell lines SW480, HT29, and HCT116 were used as models. Cell viability was compared under treatments with radiation, radiation/5-fluorouracil, metformin, radiation/metformin, and radiation/5-fluorouracil/metformin. Nude mice were injected subcutaneously with SW480 cells and treated for 1 week with radiation/5-fluorouracil, metformin, radiation/metformin, or radiation/5-fluorouracil/metformin. Tumor volume was evaluated for 4 weeks after treatment completion. The phosphorylation status of key proteins of the PI3K/Akt/mTOR pathway was determined by immunoblots. SETTINGS: This was an experimental study conducted in vitro and in vivo. PATIENTS: Animal models/cell lines were used. MAIN OUTCOME MEASURES: The end point was to investigate how metformin compares with 5-fluorouracil as a radiosensitizer. RESULTS: All cell lines significantly decreased cell viability after treatment with radiation/metformin when compared with radiation alone. Radiation/metformin was superior to radiation/5-fluorouracil in SW480 (37% vs 74%; p < 0.001). In HT29 and in HCT116, radiation/metformin was inferior to radiation/5-fluorouracil (40.0% vs 13.8%, p < 0.001 and 40.0% vs 7.0%, p < 0.001), mainly because of increased 5-fluorouracil toxicity (≤20% of cell viability). In vivo assays indicated that radiation/metformin treatment was comparable with radiation/5-fluorouracil (557 vs 398 mm; p > 0.05) and that the addition of metformin to the standard radiation/5-fluorouracil did not improve tumor response (349 mm; p > 0.05). Metformin exerted strong PI3K/Akt/mTOR pathway inactivation effects after 24-hour exposure (increasing pAMPK, p < 0.01; decreasing pAkt, p < 0.01; and pS6, p <0.05). LIMITATIONS: In vitro and in vivo chemoradiation regimens cannot be directly translated to human delivery methods. CONCLUSIONS: Metformin enhances tumor response to radiation in vitro and in vivo. Metformin is an attractive alternative radiosensitizing agent to be considered in future studies/trials. See Video Abstract at http://links.lww.com/DCR/B219. LA METFORMINA COMO AGENTE RADIOSENSIBILIZADOR ALTERNATIVO A 5FU DURANTE EL TRATAMIENTO NEOADYUVANTE PARA CÁNCER DE RECTO: La quimiorradiación neoadyuvante para el cáncer de recto localmente avanzado que combina 5FU con radiación aumenta la regresión tumoral en comparación con la radiación sola. Sin embargo, se produce a costa de una toxicidad significativa relacionada con el tratamiento. Los pacientes con cáncer de recto que usan metformina se han asociado con una mejor respuesta a la radioterapia.Evaluar los efectos radiosensibilizantes de metformina in vitro e in vivo y compararlo con la combinación estándar de radiación / 5FU.Se usaron como modelos las líneas celulares de cáncer colorrectal SW480, HT29 y HCT116. La viabilidad celular se comparó en tratamientos con radiación, radiación / 5FU, metformina, radiación / metformina y radiación / 5FU / metformina. A los ratones desnudos se les inyectó por vía subcutánea células SW480 y fueron tratados durante una semana con radiación / 5FU, metformina, radiación / metformina o radiación / 5FU / metformina. El volumen tumoral se evaluó durante 4 semanas después de la finalización del tratamiento. El estado de fosforilación de las proteínas clave de la vía PI3K / Akt / mTOR se determinó mediante inmunotransferencias.Estudio experimental in vitro e in vivo.Modelo animal / líneas celulares.El punto final fue investigar cómo la metformina se compara con 5FU como un radiosensibilizador.Todas las líneas celulares disminuyeron significativamente la viabilidad celular después del tratamiento con radiación / metformina en comparación con la radiación sola. La radiación / metformina fue superior a la radiación / 5FU en SW480 (37% frente a 74%; p <0,001). En el HT29 y el HCT116 la radiación / metformina fue inferior a la radiación / 5FU (40% vs 13.8%, p <0.001 y 40% vs 7%, p <0.001; respectivamente), debido principalmente al aumento de la toxicidad de 5FU (≤20% de la célula viabilidad). Los ensayos in vivo indicaron que el tratamiento con radiación / metformina era comparable a la radiación / 5FU (557 vs 398 mm, p > 0.05), y que la adición de metformina a la radiación estándar / 5FU no mejoró la respuesta tumoral (349 mm, p > 0.05). La metformina ejerció fuertes efectos de inactivación de la vía PI3K / Akt / mTOR después de 24 horas de exposición (aumentando pAMPK p < 0.01, disminuyendo pAkt, p < 0.01; y pS6, p < 0.05).Los regímenes de CRT in vitro e in vivo no se pueden traducir directamente a los métodos de entrega en humanos.La metformina mejora la respuesta tumoral a la radiación in vitro e in vivo. La metformina es un agente alternativo de radiosensibilización atractivo para ser considerado en futuros estudios / ensayos. Consulte Video Resumen en http://links.lww.com/DCR/B219. (Traducción-Dr Gonzalo Hagerman).

Quantification of Tumor Cell Adhesion in Lymph Node Cryosections.

Tumor-draining lymph nodes (LNs) are not merely filters of tumor-produced waste. They are one of the most common regional sites of provisional residence of disseminated tumor cells in patients with different types of cancer. The detection of these LN-residing tumor cells is an important biomarker associated with poor prognosis and adjuvant therapy decisions. Recent mouse models have indicated that LN-residing tumor cells could be a substantial source of malignant cells for distant metastases. The ability to quantify the adhesivity of tumor cells to LN parenchyma is a critical gauge in experimental research that focuses on the identification of genes or signaling pathways relevant for lymphatic/metastatic dissemination. Because LNs are complex 3D structures with a variety of appearances and compositions in tissue sections depending on the plane of section, their matrices are difficult to replicate experimentally in vitro in a fully controlled way. Here, we describe a simple and inexpensive method that allows the quantification of adhesive tumor cells to LN cryosections. Using serial sections of the same LN, we adapt the classic method developed by Brodt to use nonradioactive labels and directly count the number of adhering tumor cells per LN surface area. LN-adherent tumor cells are readily identified by light microscopy and confirmed by a fluorescence-based method, giving an adhesion index that reveals the cell-binding affinity to LN parenchyma, which is suggestive evidence of molecular alterations in the affinity binding of integrins to their correlate LN-ligands.

Beyond the Proteolytic Activity: Examining the Functional Relevance of the Ancillary Domains Using Tri-Dimensional (3D) Spheroid Invasion Assay.

In this chapter, we describe a straightforward protocol to generate multicellular tumor spheroids (MTSs) and evaluate the role of specific genes in regulating cell invasiveness in real-time and tridimensional (3D) matrices. This approach provides advantages over other conventional invasion assays by offering intimate cell-cell and cell-ECM contacts and by mimicking the pathophysiological characteristics observed in tumor microenvironments (e.g., microregional gradients in glucose and O2 concentrations and metabolic and proliferative tumor heterogeneity). We also provide an original and semiautomated approach to quantify MTS invasion using the freely available ImageJ software and plugins.

Inhibition of the RhoA GTPase Activity Increases Sensitivity of Melanoma Cells to UV Radiation Effects.

Ultraviolet radiation is the main cause of DNA damage to melanocytes and development of melanoma, one of the most lethal human cancers, which leads to metastasis due to uncontrolled cell proliferation and migration. These phenotypes are mediated by RhoA, a GTPase overexpressed or overactivated in highly aggressive metastatic tumors that plays regulatory roles in cell cycle progression and cytoskeleton remodeling. This work explores whether the effects of UV on DNA damage, motility, proliferation, and survival of human metastatic melanoma cells are mediated by the RhoA pathway. Mutant cells expressing dominant-negative (MeWo-RhoA-N19) or constitutively active RhoA (MeWo-RhoA-V14) were generated and subjected to UV radiation. A slight reduction in migration and invasion was observed in MeWo and MeWo-RhoA-V14 cells but not in MeWo-RhoA-N19 cells, which presented inefficient motility and invasiveness associated with stress fibers fragmentation. Proliferation and survival of RhoA-deficient cells were drastically reduced by UV compared to cells displaying normal or high RhoA activity, suggesting increased sensitivity to UV. Loss of RhoA activity also caused less efficient DNA repair, with elevated levels of DNA lesions such as strand breaks and cyclobutane pyrimidine dimers (CPDs). Thus, RhoA mediates genomic stability and represents a potential target for sensitizing metastatic tumors to genotoxic agents.

Ações antagônicas de FGF2 em células tumorais de camundongo: disparo de mitogênese e de morte celular / Antagonistic actions of FGF2 in mouse cells tumors: shot of mitogênese and of cell death

O objetivo desta tese é estudar o papel de FGF2 no controle do ciclo celular em células de mamíferos. Nosso principal modelo, a linhagem Y1, é derivada de um tumor funcional de córtex de camundongo que possui o proto-oncogene c-ki-ras amplificado, tendo consequentemente a super-expressão da proteína c-Ki-Ras na forma ativa (c-Ki-Ras-GTP). Células Y1 sincronizadas na interface GO G1 do ciclo celular são prontamente responsivas a tratamentos de FGF2 (fator de crescimento de fibroblasto-2) sendo este capaz de ativar toda a progressão GO G1 S do ciclo celular, mas surpreendentemente , sob estas mesmas condições, FGF2 induz em cultura e in vivo morte celular nesta linhagem, bloqueando o progresso no ciclo após a entrada na fase S. Sob condições induzidas de abaixo c-Ki-Ras-GTP, células Y1 respondem à FGF2 com um aumento na proliferação, mostrando que a indução de morte nesta linhagem está diretamente relacionado com os níveis de Ki-Ras-GTP...