A Small Step, a Giant Leap: Somatic Hypermutation of a Single Amino Acid Leads to Anti-La Autoreactivity.

The anti-La mab 312B, which was established by hybridoma technology from human-La transgenic mice after adoptive transfer of anti-human La T cells, immunoprecipitates both native eukaryotic human and murine La protein. Therefore, it represents a true anti-La autoantibody. During maturation, the anti-La mab 312B acquired somatic hypermutations (SHMs) which resulted in the replacement of four aa in the complementarity determining regions (CDR) and seven aa in the framework regions. The recombinant derivative of the anti-La mab 312B in which all the SHMs were corrected to the germline sequence failed to recognize the La antigen. We therefore wanted to learn which SHM(s) is (are) responsible for anti-La autoreactivity. Humanization of the 312B ab by grafting its CDR regions to a human Ig backbone confirms that the CDR sequences are mainly responsible for anti-La autoreactivity. Finally, we identified that a single amino acid replacement (D > Y) in the germline sequence of the CDR3 region of the heavy chain of the anti-La mab 312B is sufficient for anti-La autoreactivity.

And Yet It Moves: Oxidation of the Nuclear Autoantigen La/SS-B Is the Driving Force for Nucleo-Cytoplasmic Shuttling.

Decades ago, we and many other groups showed a nucleo-cytoplasmic translocation of La protein in cultured cells. This shuttling of La protein was seen after UV irradiation, virus infections, hydrogen peroxide exposure and the Fenton reaction based on iron or copper ions. All of these conditions are somehow related to oxidative stress. Unfortunately, these harsh conditions could also cause an artificial release of La protein. Even until today, the shuttling and the cytoplasmic function of La/SS-B is controversially discussed. Moreover, the driving mechanism for the shuttling of La protein remains unclear. Recently, we showed that La protein undergoes redox-dependent conformational changes. Moreover, we developed anti-La monoclonal antibodies (anti-La mAbs), which are specific for either the reduced form of La protein or the oxidized form. Using these tools, here we show that redox-dependent conformational changes are the driving force for the shuttling of La protein. Moreover, we show that translocation of La protein to the cytoplasm can be triggered in a ligand/receptor-dependent manner under physiological conditions. We show that ligands of toll-like receptors lead to a redox-dependent shuttling of La protein. The shuttling of La protein depends on the redox status of the respective cell type. Endothelial cells are usually resistant to the shuttling of La protein, while dendritic cells are highly sensitive. However, the deprivation of intracellular reducing agents in endothelial cells makes endothelial cells sensitive to a redox-dependent shuttling of La protein.

Biological Activity of Sacha Inchi (Plukenetia volubilis Linneo) and Potential Uses in Human Health: A Review.

Sacha inchi (Plukenetia volubilis Linneo) is an ancestral plant originating in the Amazon jungle that has been adopted as a food source due to its high nutritional value, which has gradually been recognized to have potential benefits for human health. Diverse prospective studies have evaluated the effect of consuming components from the plant, derivatives from its seeds, leaves and shell on preventing the risk of cardiovascular disease, chronic inflammatory disease, dermatitis and controlling tumor proliferation, especially given its recognized high content of essential fatty acids, phenolic compounds and vitamin E, showing antioxidant, hypolipidemic, immunomodulation and emollient activity, as well as the capacity to remove heavy metals from aqueous solutions. This review offers a complete description of the existing information on the use and biological activity of P. volubilis L., based on its essential lipid components and evidenced on its use in the field of human health, in prevention, therapeutic and nutritional contexts, along with industrial uses, making it a promising bioresource.

Nanosensors in clinical development of CAR-T cell immunotherapy.

Immunotherapy using CAR-T cells is a new technological paradigm for cancer treatment. To avoid severe side effects and tumor escape variants observed for conventional CAR-T cells approach, adaptor CAR technologies are under development, where intermediate target modules redirect immune cells against cancer. In this work, silicon nanowire field-effect transistors are used to develop target modules for an optimized CAR-T cell operation. Focusing on a library of seven variants of E5B9 peptide that is used as CAR targeting epitope, we performed multiplexed binding tests using nanosensor chips. These peptides had been immobilized onto the sensor to compare the transistor signals upon titration with anti-La 5B9 antibodies. The correlation of binding affinities and sensor sensitivities enabled a selection of candidates for the interaction between CAR and target modules. An extremely low detection limit was observed for the sensor, down to femtomolar concentration, outperforming the current assay of the same purpose. Finally, the CAR T-cells redirection capability of selected peptides in target modules was proven successful in an in-vitro cytotoxicity assay. Our results open the perspective for the nanosensors to go beyond the early diagnostics in clinical cancer research towards developing and monitoring immunotherapeutic treatment, where the quantitative analysis with the standard techniques is limited.

Transcriptional epigenetic regulation of Fkbp1/Pax9 genes is associated with impaired sensitivity to platinum treatment in ovarian cancer.

BACKGROUND: In an effort to contribute to overcoming the platinum resistance exhibited by most solid tumors, we performed an array of epigenetic approaches, integrating next-generation methodologies and public clinical data to identify new potential epi-biomarkers in ovarian cancer, which is considered the most devastating of gynecological malignancies. METHODS: We cross-analyzed data from methylome assessments and restoration of gene expression through microarray expression in a panel of four paired cisplatin-sensitive/cisplatin-resistant ovarian cancer cell lines, along with publicly available clinical data from selected individuals representing the state of chemoresistance. We validated the methylation state and expression levels of candidate genes in each cellular phenotype through Sanger sequencing and reverse transcription polymerase chain reaction, respectively. We tested the biological role of selected targets using an ectopic expression plasmid assay in the sensitive/resistant tumor cell lines, assessing the cell viability in the transfected groups. Epigenetic features were also assessed in 189 primary samples obtained from ovarian tumors and controls. RESULTS: We identified PAX9 and FKBP1B as potential candidate genes, which exhibited epigenetic patterns of expression regulation in the experimental approach. Re-establishment of FKBP1B expression in the resistant OVCAR3 phenotype in which this gene is hypermethylated and inhibited allowed it to achieve a degree of platinum sensitivity similar to the sensitive phenotype. The evaluation of these genes at a translational level revealed that PAX9 hypermethylation leads to a poorer prognosis in terms of overall survival. We also set a precedent for establishing a common epigenetic signature in which the validation of a single candidate, MEST, proved the accuracy of our computational pipelines. CONCLUSIONS: Epigenetic regulation of PAX9 and FKBP1B genes shows that methylation in non-promoter areas has the potential to control gene expression and thus biological consequences, such as the loss of platinum sensitivity. At the translational level, PAX9 behaves as a predictor of chemotherapy response to platinum in patients with ovarian cancer. This study revealed the importance of the transcript-specific study of each gene under potential epigenetic regulation, which would favor the identification of new markers capable of predicting each patient's progression and therapeutic response.

An oligo-His-tag of a targeting module does not influence its biodistribution and the retargeting capabilities of UniCAR T cells.

Recently, we established the controllable modular UniCAR platform technology to advance the efficacy and safety of CAR T cell therapy. The UniCAR system is composed of (i) target modules (TMs) and (ii) UniCAR armed T cells. TMs are bispecific molecules that are able to bind to the tumor cell surface and simultaneously to UniCAR T cells. For interaction with UniCAR T cells, TMs contain a peptide epitope sequence which is recognised by UniCAR T cells. So far, a series of TMs against a variety of tumor targets including against the prostate stem cell antigen (PSCA) were constructed and functionally characterised. In order to facilitate their purification all these TMs are expressed as recombinant proteins equipped with an oligo-His-tag. The aim of the here presented manuscript was to learn whether or not the oligo-His-tag of the TM influences the UniCAR system. For this purpose, we constructed TMs against PSCA equipped with or lacking an oligo-His-tag. Both TMs were compared side by side including for functionality and biodistribution. According to our data, an oligo-His-tag of a UniCAR TM has only little if any effect on its binding affinity, in vitro and in vivo killing capability and in vivo biodistribution.

A theranostic PSMA ligand for PET imaging and retargeting of T cells expressing the universal chimeric antigen receptor UniCAR.

Chimeric antigen receptor (CAR) T cells have shown impressive therapeutic potential. Due to the lack of direct control mechanisms, therapy-related adverse reactions including cytokine release- and tumor lysis syndrome can even become life-threatening. In case of target antigen expression on non-malignant cells, CAR T cells can also attack healthy tissues. To overcome such side effects, we have established a modular CAR platform termed UniCAR: UniCAR T cells per se are inert as they recognize a peptide epitope (UniCAR epitope) that is not accessible on the surface of living cells. Bifunctional adapter molecules termed target modules (TM) can cross-link UniCAR T cells with target cells. In the absence of TMs, UniCAR T cells automatically turn off. Until now, all UniCAR TMs were constructed by fusion of the UniCAR epitope to an antibody domain. To open up the wide field of low-molecular-weight compounds for retargeting of UniCAR T cells to tumor cells, and to follow in parallel the progress of UniCAR T cell therapy by PET imaging we challenged the idea to convert a PET tracer into a UniCAR-TM. For proof of concept, we selected the clinically used PET tracer PSMA-11, which binds to the prostate-specific membrane antigen overexpressed in prostate carcinoma. Here we show that fusion of the UniCAR epitope to PSMA-11 results in a low-molecular-weight theranostic compound that can be used for both retargeting of UniCAR T cells to tumor cells, and for non-invasive PET imaging and thus represents a member of a novel class of theranostics.

Seroprevalencia y factores de riesgo asociados a toxoplasmosis gestacional en el Nororiente Colombiano / Seroprevalence and risk factors for congenital toxoplasmosis in the northeastern region of Colombia / Soroprevalência e fatores de risco associados à toxoplasmose gestacional no nordeste da Colômbia

Introduction: Toxoplasmosis is a zoonosis prevalent in one-third of the world's population, adversely affecting maternal-fetal health by causing varying degrees of damage to the fetus. Objective: To assess the seroprevalence of Toxoplasma gondii IgG and IgM antibodies and associated risk factors in first-trimester pregnant women in Cúcuta, Colombia, in 2018. Materials and Methods: Cross-sectional and correlational study in 111 women who voluntarily participated. Specific IgM and IgG antibodies were detected by luminescence immunoassay (LIA). Results: Total seropositivity for IgM and IgG was 19.8% and 35.1%, respectively, 11.7% was for IgM only, and 53.2% corresponded to the overall seronegativity rate for T. gondii. Risk factors (CI = 95%) such as undercooked meat consumption (54.1% of cases, OR = 1.8, p = 0.120), tap water consumption (48.6%, OR = 1.4, p = 0.421), and goat or cow raw milk consumption (39.6%, OR = 0.78, p = 0.553) were identified; in addition, living with cats (23.4%) was identified as a risk factor significantly associated with parasite seropositivity (OR = 2.8, p = 0.025). Discussion and Conclusions: Our findings showed a possible risk of primary infection in more than half of the pregnant population, given the seronegativity against the parasite. A considerable frequency of suspected cases of very recent infection was also found. In addition to being associated with a previously recognized risk factor, this fact suggests the presence of other dietary risks that should be addressed through prevention strategies during prenatal care and the need to strengthen event surveillance.

Introdução: A toxoplasmose é uma zoonose prevalente em um terço da população mundial, que afeta negativamente a saúde materno-fetal, causando diversos graus de danos ao feto. Objetivo: Propôs-se avaliar o estado sorológico IgG e IgM anti - Toxoplasma gondii e fatores de risco relacionados, em gestantes no primeiro trimestre em Cúcuta, Colômbia, no ano de 2018. Material e Métodos: Estudo transversal e correlacional em Participaram 111 mulheres voluntariamente, que foram testadas para IgM e IgG específicos pelo imunoensaio LIA. Resultados: 19,8% e 35,1% do total de soropositividade foram encontrados para IgM e IgG, respectivamente, 11,7% foi apenas para IgM e 53,2% corresponde à frequência de soronegatividade geral para T.gondii; Foram identificados fatores de risco (IC=95%) , como consumo de carne mal cozida (54,1% dos casos, OR= 1,8, p=0,120 ), água da torneira (48,6%, OR=1,4, p=0,421), cru leite de cabra ou vaca (39,6%, OR=0,78, p=0,553 ), além de conviver com gatos (23,4%), sendo este último significativamente associado à soropositividade ao parasita ( OR=2,8 , p =0,025 ). Discussão e conclusões: Nossos achados revelam um possível risco de infecção primária em mais da metade da população gestante dada a sua soronegatividade contra o parasita, mas também uma frequência considerável de casos com suspeita de infecção muito recente, que, além de estar associada a um fator de risco previamente reconhecido , sugere outros aspectos de risco em torno da alimentação que devem ser impactados por meio de estratégias de prevenção durante o pré-natal, sugerindo a necessidade de fortalecer a vigilância em torno do evento.

Effect of the addition of Moringa Oleifera to fruit drinks on clinical parameters associated with iron deficiency anaemia in schoolchildren

Worldwide, iron deficiency anaemia (IDA) is one the most significant nutritional maladies, especially in low and middle-income countries. This is mainly due to the associated morbidity, economic implications on the health system and the welfare state. One of the most prevalent causes of IDA in children is insufficient food intake, which has an obvious connection with the social environment. The main strategy in the management of this condition is counteracting such deficiency through the improvement of nutrition and family upbringing. Aiming to explore affordable, novel and straightforward approaches to treat this nutritional condition, we developed a descriptive study associated with the incorporation of Moringa Oleifera in homemade fruit beverages delivered to 32 children from low-income families, in order to assess the evolution of IDA, evaluating specific clinical parameters such as red blood cells volume, as well as hemoglobin, ferritin and serum iron levels, within a two months lapse. At the end of this period, we observed a significant statistical rise in the levels of all the assessed parameters, finding about a 1.3-fold increase in hemoglobin and ferritin levels and in red blood cells volume; serum iron was 1.1 times higher. These findings are demonstrative of the wide spectrum of the medicinal features of Moringa and establish a promising and accessible strategy easy to incorporate into the diet of families of children suffering this pathology(AU)

La anemia ferropénica (AF) es una de las carencias nutricionales más significativas a nivel mundial, especialmente en los países de bajos y medios ingresos, debido principalmente a la morbilidad asociada, las implicaciones en el sistema de salud y el estado de bienestar. Una de las causas más frecuentes de esta deficiencia en los niños es la ingesta insuficiente, que tiene una evidente conexión con el entorno social. La principal estrategia en el manejo de esta afección es contrarrestar esta deficiencia a través de la mejora de la nutrición y la educación familiar. Con el objetivo de explorar enfoques asequibles, novedosos y directos para el tratamiento de esta enfermedad, se desarrolló un estudio descriptivo asociado a la incorporación de la Moringa Oleifera en bebidas de frutas caseras que se suministraron a 32 niños de familias de bajos ingresos para hacerle seguimiento a la evolución de la anemia por deficiencia de hierro, evaluando parámetros clínicos específicos como el volumen de células rojas y los niveles de hemoglobina, ferritina y hierro sérico en un lapso de dos meses. Se observó un aumento estadísticamente significativo en los niveles de todos los parámetros evaluados una vez finalizado el período de intervención. Al final del ensayo, observamos un aumento estadístico significativo en los niveles de todos los parámetros evaluados, encontrando un aumento de aproximadamente 1.3 veces en los niveles de hemoglobina y ferritina y en el volumen de los glóbulos rojos; el hierro sérico fue 1.1 veces mayor. Estos hallazgos son una demostración del amplio espectro de las características medicinales de la Moringa y establecen una estrategia prometedora y accesible, fácil de incorporar a la dieta de las familias de los niños que sufren esta patología(AU)