Dr. Hepp-Castro, Matías

Se define cáncer como un conjunto de enfermedades con características comunes, las cuales se originan por la acumulación de modificaciones genéticas en una célula que pierde los mecanismos de control de la proliferación y sobrevivencia. Se han descrito más de 200 tipos diferentes de cáncer, los que afectan a cualquier órgano del cuerpo, con síntomas y tratamientos distintos 1 . Estas desregulaciones se deben a factores genéticos heredados, o factores externos como: tabaco, alcohol, obesidad, exposición a radiación, etc. Estos factores, pueden generar procesos celulares como: 1) hiperproliferación celular; 2) evasión de factores supresores de crecimiento; 3) activación de invasión y metástasis; 4) inmortalidad replicativa; 5) inducción de angiogénesis y 6) resistencia a apoptosis, conocidos como los sellos distintivos del cáncer ( hallmarks ) 2,3 , vinculados con el control del ciclo celular. Aunque el ciclo celular esté regulado adecuadamente, el ADN puede sufrir alteraciones genéticas, en genes reparadores, genes supresores de tumores y oncogenes. Esto conlleva desregulación de varios procesos, generando activación de protooncogenes, inactivación de genes supresores de tumores o ambos 3. Cuando no hay equilibrio entre la proliferación y la apoptosis, aumenta el riesgo de tumorigénesis, y las células neoplásicas crecen sin control. Aquí aparecen otras propiedades de las células cancerosas, como desplazarse y colonizar sectores del organismo, generando metástasis o invasión celular. Estas células cancerígenas, además, podrían activar a células endoteliales y generar angiogénesis para poder nutrirse, lo que produciría crecimiento tumoral 3,4 . A los sellos distintivos del cáncer ya mencionados, se incorporan 4 procesos emergentes en la formación y mantención del cáncer 2 , generando procesos que modifican el contexto celular. Dos de estos apuntan a facilitar la aparición de los marcadores principales. Uno es la inestabilidad genómica, lo que confiere a las células cancerosas alteraciones genéticas que conducen a progresión tumoral. El otro, producir inflamación por células inmunes innatas diseñadas para combatir infecciones o sanar heridas, dando lugar a procesos que son promotores de respuesta inflamatoria tumoral. Los otros dos mecanismos son focos emergentes directos. Siendo uno, el modificar o reprogramar el metabolismo celular para favorecer la proliferación neoplásica. El otro es la evasión de células cancerosas a destrucción inmunológica, por linfocitos T y B, macrófagos y natural killers (NK) 2 , generándose blancos terapéuticos diferentes a los convencionales. Existen muchos tratamientos para el cáncer, pero los convencionales (cirugía, radiación y quimioterapia) siguen siendo más utilizados, y aún efectivos, pero invasivos y tóxicos. El tratamiento se selecciona según el tipo de tumor, estadio de enfermedad y condiciones del paciente 4 . La tasa de éxito de los tratamientos convencionales está limitada por la toxicidad y la no especificidad para el tipo de tumor 3 . En el mundo científico se abre espacio para terapias emergentes, como utilizar la respuesta inmune como mecanismo terapéutico 5,6 . Pero ¿cómo se podría abordar la respuesta inmunológica? Para esto, primero debemos entender en qué consiste.

Radiata pine bark is a widely available organic waste, requiring alternative uses due to its environmental impact on soil, fauna, and forest fires. Pine bark waxes could be used as cosmetic substitutes, but their toxicity requires evaluation since pine bark may contain toxic substances or xenobiotics, depending on the extraction process. This study evaluates the toxicity of radiata pine bark waxes obtained through various extraction methods on human skin cells grown in vitro. The assessment includes using XTT to evaluate mitochondrial activity, violet crystal dye to assess cell membrane integrity, and ApoTox-Glo triple assay to measure cytotoxicity, viability, and apoptosis signals. Pine bark waxes extracted via T3 (acid hydrolysis and petroleum ether incubation) and T9 (saturated steam cycle, alkaline hydrolysis, and petroleum ether incubation) exhibit non-toxicity up to 2% concentration, making them a potential substitute for petroleum-based cosmetic materials. Integrating the forestry and cosmetic industries through pine bark wax production under circular economy principles could promote development while replacing petroleum-based materials. Extraction methodology affects pine bark wax toxicity in human skin cells due to the retention of xenobiotic compounds including methyl 4-ketohex-5-enoate; 1-naphthalenol; dioctyl adipate; eicosanebioic acid dimethyl ester; among others. Future research will investigate whether the extraction methodology alters the molecular structure of the bark, affecting the release of toxic compounds in the wax mixture.

The bark of Pinus radiata D. Don is a bioresource of great worldwide abundance. While various forms of use have been studied, it is still a little-used bioresource. Due to its great accumulation, significant solid emissions of this residue generate environmental problems such as changes in soil chemistry, ecological problems such as the alteration of arthropod communities and fire risk. The opportunity to take advantage of this bioresource could be in its wax content, which could be a replacement for the main raw materials used in the production of cosmetics. These currently correspond to petroleum-derived substances, such as petroleum jelly, paraffin, or mineral oil. The importance of replacing these raw materials is that several studies report that they are the main causes of human skin diseases, such as chemical hypersensitivity syndrome and allergic contact dermatitis. This study seeks to prove that lipophilic extracts (waxes) from pine bark can replace petroleum-derived raw materials used in cosmetics. To achieve this, pine bark at drying conditions was characterized, and the performance of wax extraction by various treatments was studied. The density, viscosity, melting point, and solubility in culture media of the obtained waxes were determined. The waxes were chemically characterized by FT-IR and GC–MS analysis. The results reveal that the maximum moisture of the bark is 14.54 %, the best extraction yields are obtained by using water at 120 ◦C and 1.2 atm, and petroleum ether (3.12 %), alkaline hydrolysis 1 mol L− 1 (NaOH) and petroleum ether (3.53 %) ethyl acetate (3.23 %). Values were close to the reference study using the rapid lipid extraction method. The density of the wax is 0.845 g mL− 1 and its viscosity of 530 cP (24 ◦C), and the melting point varied according to the extractive treatment at between 25 and 40 ◦C. Solubility tests made it possible to determine that the 10/50/1000 μL ratio of modified Eagle Dulbecco wax/dimethylsulfoxide/medium allows homogeneous solubilization of the wax without the presence of precipitates. Chemical characterization identified typical functional groups of plant-based waxes such as long-chain alkanes, alkyls, methyl groups, esters, and carbonyls, with the most abundant fatty acids being C:22 and C:24.

The SALL2 transcription factor, an evolutionarily conserved gene through vertebrates, is involved in normal development and neuronal differentiation. In disease, SALL2 is associated with eye, kidney, and brain disorders, but mainly is related to cancer. Some studies support a tumor suppressor role and others an oncogenic role for SALL2, which seems to depend on the cancer type. An additional consideration is tissue-dependent expression of different SALL2 isoforms. Human and mouse SALL2 gene loci contain two promoters, each controlling the expression of a different protein isoform (E1 and E1A). Also, several improvements on the human genome assembly and gene annotation through next-generation sequencing technologies reveal correction and annotation of additional isoforms, obscuring dissection of SALL2 isoform-specific transcriptional targets and functions. We here integrated current data of normal/tumor gene expression databases along with ChIP-seq binding profiles to analyze SALL2 isoforms expression distribution and infer isoform-specific SALL2 targets. We found that the canonical SALL2 E1 isoform is one of the lowest expressed, while the E1A isoform is highly predominant across cell types. To dissect SALL2 isoform-specific targets, we analyzed publicly available ChIP-seq data from Glioblastoma tumor-propagating cells and in-house ChIP-seq datasets performed in SALL2 wild-type and E1A isoform knockout HEK293 cells. Another available ChIP-seq data in HEK293 cells (ENCODE Consortium Phase III) overexpressing a non-canonical SALL2 isoform (short_E1A) was also analyzed. Regardless of cell type, our analysis indicates that the SALL2 long E1 and E1A isoforms, but not short_E1A, are mostly contributing to transcriptional control, and reveals a highly conserved network of brain-specific transcription factors (i.e., SALL3, POU3F2, and NPAS3). Our data integration identified a conserved molecular network in which SALL2 regulates genes associated with neural function, cell differentiation, development, and cell adhesion between others. Also, we identified PODXL as a gene that is likely regulated by SALL2 across tissues. Our study encourages the validation of publicly available ChIP-seq datasets to assess a specific gene/isoform’s transcriptional targets. The knowledge of SALL2 isoforms expression and function in different tissue contexts is relevant to understanding its role in disease.

SALL2/Sall2 is a transcription factor associated with development, neuronal differentiation, and cancer. Interestingly, SALL2/Sall2 deficiency leads to failure of the optic fissure closure and neurite outgrowth, suggesting a positive role for SALL2/Sall2 in cell migration. However, in some cancer cells, SALL2 deficiency is associated with increased cell migration. To further investigate the role of Sall2 in the cell migration process, we used immortalized Sall2 knockout (Sall2−/−) and Sall2 wild-type (Sall2+/+) mouse embryonic fibroblasts (iMEFs). Our results indicated that Sall2 positively regulates cell migration, promoting cell detachment and focal adhesions turnover. Sall2 deficiency decreased cell motility and altered focal adhesion dynamics. Accordingly, restoring Sall2 expression in the Sall2−/− iMEFs by using a doxycycline-inducible Tet-On system recovered cell migratory capabilities and focal adhesion dynamics. In addition, Sall2 promoted the autophosphorylation of Focal Adhesion Kinase (FAK) at Y397 and increased integrin β1 mRNA and its protein expression at the cell surface. We demonstrated that SALL2 increases ITGB1 promoter activity and binds to conserved SALL2-binding sites at the proximal region of the ITGB1 promoter, validated by ChIP experiments. Furthermore, the overexpression of integrin β1 or its blockade generates a cell migration phenotype similar to that of Sall2+/+ or Sall2−/− cells, respectively. Altogether, our data showed that Sall2 promotes cell migration by modulating focal adhesion dynamics, and this phenotype is associated with SALL2/Sall2-transcriptional regulation of integrin β1 expression and FAK autophosphorylation. Since deregulation of cell migration promotes congenital abnormalities, tumor formation, and spread to other tissues, our findings suggest that the SALL2/Sall2-integrin β1 axis could be relevant for those processes.

Spalt-like proteins are Zinc finger transcription factors from Caenorhabditis elegans to vertebrates, with critical roles in development. In vertebrates, four paralogues have been identified (SALL1-4), and SALL2 is the family’s most dissimilar member. SALL2 is required during brain and eye development. It is downregulated in cancer and acts as a tumor suppressor, promoting cell cycle arrest and cell death. Despite its critical functions, information about SALL2 regulation is scarce. Public data indicate that SALL2 is ubiquitinated and phosphorylated in several residues along the protein, but the mechanisms, biological consequences, and enzymes responsible for these modifications remain unknown. Bioinformatic analyses identified several putative phosphorylation sites for Casein Kinase II (CK2) located within a highly conserved C-terminal PEST degradation motif of SALL2. CK2 is a serine/threonine kinase that promotes cell proliferation and survival and is often hyperactivated in cancer. We demonstrated that CK2 phosphorylates SALL2 residues S763, T778, S802, and S806 and promotes SALL2 degradation by the proteasome. Accordingly, pharmacological inhibition of CK2 with Silmitasertib (CX-4945) restored endogenous SALL2 protein levels in SALL2-deficient breast MDA-MB-231, lung H1299, and colon SW480 cancer cells. Silmitasertib induced a methuosis-like phenotype and cell death in SW480 cells. However, the phenotype was significantly attenuated in CRISPr/Cas9-mediated SALL2 knockout SW480 cells. Similarly, Sall2-deficient tumor organoids were more resistant to Silmitasertib-induced cell death, confirming that SALL2 sensitizes cancer cells to CK2 inhibition. We identified a novel CK2-dependent mechanism for SALL2 regulation and provided new insights into the interplay between these two proteins and their role in cell survival and proliferation.

Colorectal cancer (CRC) is one of the most common malignant neoplasms and the second leading cause of death from tumors worldwide. Therefore, there is a great need to study new therapeutical strategies, such as effective immunotherapies against these malignancies. Unfortunately, many CRC patients do not respond to current standard immunotherapies, making it necessary to search for adjuvant treatments. Histone deacetylase 6 (HDAC6) is involved in several processes, including immune response and tumor progression. Specifically, it has been observed that HDAC6 is required to activate the Signal Transducer and Activator of Transcription 3 (STAT3), a transcription factor involved in immunogenicity, by activating different genes in these pathways, such as PD-L1. Over-expression of immunosuppressive pathways in cancer cells deregulates T-cell activation. Therefore, we focused on the pharmacological inhibition of HDAC6 in CRC cells because of its potential as an adjuvant to avoid immunotolerance in immunotherapy. We investigated whether HDAC6 inhibitors (HDAC6is), such as Nexturastat A (NextA), affected STAT3 activation in CRC cells. First, we found that NextA is less cytotoxic than the non-selective HDACis panobinostat. Then, NextA modified STAT3 and decreased the mRNA and protein expression levels of PD-L1. Importantly, transcriptomic analysis showed that NextA treatment affected the expression of critical genes involved in immunomodulatory pathways in CRC malignancies. These results suggest that treatments with NextA reduce the functionality of STAT3 in CRC cells, impacting the expression of immunomodulatory genes involved in the inflammatory and immune responses. Therefore, targeting HDAC6 may represent an interesting adjuvant strategy in combination with immunotherapy.

Introducción: La cuantificación de SARS-CoV-2 en aguas residuales es una herramienta que permite determinar la tendencia de la circulación viral en un área geográfica determinada. Objetivo: Cuantificar el virus SARS-CoV-2 en 15 plantas de tratamiento de aguas residuales en diferentes ciudades de Chile para establecer una comparación con las variables de: i) casos activos por cada 100.000 habs.; ii) positividad diaria (casos nuevos); y iii) fases del plan de confinamiento. Metodología: SARS-CoV-2 se concentró a partir de muestras de aguas residuales. Para obtener el número de genomas del virus por litro se realizó una cuantificación absoluta utilizando qRT-PCR. Resultados: Entre enero y junio de 2021 se procesaron 253 muestras, siendo todas positivas para la presencia del virus. Asimismo, se logró determinar que la tasa de casos activos por cada 100.000 habs. es la variable que mejor se ajusta a las tendencias obtenidas con la cuantificación de la carga viral en las aguas residuales. Conclusiones: La cuantificación de SARS-CoV-2 en las aguas residuales de manera permanente es una herramienta eficiente para determinar la tendencia del virus en un área geográfica determinada y, en conjunto con una vigilancia genómica, puede constituirse en una vigilancia centinela ideal generando alertas sobre futuros brotes.

Diverse factors play roles in chromatin dynamics, including linker proteins. Among them are high mobility group (HMG) box family proteins and linker histones. In the yeast Saccharomyces cerevisiae, Hmo1 has been identified as an HMG-box protein. This protein displays properties that are in agreement with this allocation. However, a number of studies have postulated that Hmo1 functions as a linker histone in yeast. On the other hand, when discovered, the Hho1 protein was identified as a linker histone. While multiple studies support this classification, some findings point to characteristics of Hho1 that are dissimilar to those commonly assigned to linker histones. In order to better understand the roles played by Hmo1 and Hho1 in chromatin dynamics and transcriptional regulation, we performed several analyses directly comparing these two proteins. Our analyses of genome-wide binding profiles support the belonging of Hmo1 to the HMGB family and Hho1 to the linker histones family. Interestingly, by performing protein-protein interaction analyses we found that both Hmo1 and Hho1 display physical interaction with the ATP-dependent chromatin remodeling complexes RSC, ISW1a and SWI/SNF. Moreover, by carrying out nucleosome remodeling assays, we found that both proteins stimulate the activity of the ISW1a complex. However, in the case of RSC, Hmo1 and Hho1 displayed differential properties, with Hho1 mainly showing an inhibitory effect. Our results are in agreement with the opposite roles played by RSC and ISW1a in chromatin dynamics and transcriptional regulation, and expand the view for the roles played by Hho1 and linker histones.