Preceding anti-spike IgG levels predicted risk and severity of COVID-19 during the Omicron-dominant wave in Santa Fe city, Argentina.

The SARS-CoV-2 Omicron variant has increased infectivity and immune escape compared with previous variants, and caused the surge of massive COVID-19 waves globally. Despite a vast majority (~90%) of the population of Santa Fe city, Argentina had been vaccinated and/or had been infected by SARS-CoV-2 when Omicron emerged, the epidemic wave that followed its arrival was by far the largest one experienced in the city. A serosurvey conducted prior to the arrival of Omicron allowed to assess the acquired humoral defences preceding the wave and to conduct a longitudinal study to provide individual-level real-world data linking antibody levels and protection against COVID-19 during the wave. A very large proportion of 1455 sampled individuals had immunological memory against COVID-19 at the arrival of Omicron (almost 90%), and about half (48.9%) had high anti-spike immunoglobulin G levels (>200 UI/ml). However, the antibody titres varied greatly among the participants, and such variability depended mainly on the vaccine platform received, on having had COVID-19 previously and on the number of days elapsed since last antigen exposure (vaccine shot or natural infection). A follow-up of 514 participants provided real-world evidence of antibody-mediated protection against COVID-19 during a period of high risk of exposure to an immune-escaping highly transmissible variant. Pre-wave antibody titres were strongly negatively associated with COVID-19 incidence and severity of symptoms during the wave. Also, receiving a vaccine shot during the follow-up period reduced the COVID-19 risk drastically (15-fold). These results highlight the importance of maintaining high defences through vaccination at times of high risk of exposure to immune-escaping variants.

Adaptive metabolic rewiring to chronic SFK inhibition.

Src family kinases (SFK) are key regulators of cellular proliferation, differentiation, survival, motility and angiogenesis. As such, SFK inhibitors are being tested in clinical trials to prevent metastasis as an alternative to current treatment regimens for a variety of cancers including breast cancer. To contribute to the development of molecular tools improving SFK-targeted therapies, we used the SFK inhibitor dasatinib and a well characterized triple negative breast cancer cell line (BT20). Comparison of the response of BT20 cells with acquired resistance to dasatinib and its' parental counterpart suggest that chronic exposure to SFK inhibition results in increased dependency on TGFß signaling for proliferation, both in the absence or the presence of dasatinib. In addition, we found that acquired (but not de novo) resistance to dasatinib was reduced by non-cytotoxic concentrations compounds hindering on PI3K, mTORC1 signaling, endoplasmic reticulum stress or autophagy.

Sensitization to oxaliplatin in HCT116 and HT29 cell lines by metformin and ribavirin and differences in response to mitochondrial glutaminase inhibition.

AIM OF STUDY: In the present study, we evaluated the effect of ribavirin and metformin on the sensitivity of oxaliplatin and 5-fluorouracil (5-FU) on colon cancer. MATERIALS AND METHODS: Cell viability of two commercially available colon cancer cell lines (HT29 and HCT116) were analyzed by sulforhodamine B (SRB) assay. RESULTS: A clinically achievable and nontoxic concentration of ribavirin and metformin showed a significant synergistic effect on oxaliplatin in HT29 and HCT116 cell lines. Ribavirin showed a synergistic effect on oxaliplatin in HT29 (R = 2.93, P < 0.001) and HCT116 (R = 1.71, P < 0.001), while only in HT29 metformin synergized with oxaliplatin by 2.66 (± 0.28, P < 0.01). In addition, both cell lines showed significant differences in response to Compound 968, inhibitor of mitochondrial glutaminase activity. CONCLUSION: The data suggested that these cell lines not only turn to metabolic different sustainability process after oxaliplatin treatment but that they also have different basal metabolic requirements of glutamine in vitro which can be exploits in the future for colorectal cancer (CRC) treatment and further studies are required.

Resistance to Dasatinib in primary chronic lymphocytic leukemia lymphocytes involves AMPK-mediated energetic re-programming.

Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults in the western world. Although promising new therapies for this incurable disease are being tested in clinical trials, the therapeutic relevance of metabolic rewiring in chronic lymphocytic leukemia (CLL) is poorly understood. The aim of this study was to identify targetable metabolic differences in primary CLL lymphocytes by the use of Dasatinib. Dasatinib is a multi-tyrosine kinase inhibitor used to treat chronic myelogenous leukemia (CML) and is being tested in clinical trials for several cancers including CLL. This drug has been shown to be beneficial to CML patients suffering from diabetes by reducing their glucose plasma levels. In keeping with this previous observation, we report that Dasatinib induced glucose use while reducing lactate production, suggesting that this tyrosine kinase inhibitor decreases aerobic glycolysis and shifts glucose use in primary CLL lymphocytes. Our results suggest that primary CLL lymphocytes (independently of traditional prognostic factors) can be stratified in two subsets by their sensitivity to Dasatinib in vitro. Increased glucose use induced by Dasatinib or by inhibition of mitochondrial respiration was not sufficient to sustain survival and ATP levels in CLL samples sensitive to Dasatinib. The two subsets of primary CLL lymphocytes are characterized as well by a differential dependency on mitochondrial respiration and the use of anabolic or catabolic processes to cope with induced metabolic/energetic stress. Differential metabolic reprogramming between subsets is supported by the contrasting effect on the survival of Dasatinib treated CLL lymphocytes with pharmacological inhibition of two master metabolic regulators (mTorc1 and AMPK) as well as induced autophagy. Alternative metabolic organization between subsets is further supported by the differential basal expression (freshly purified lymphocytes) of active AMPK, regulators of glucose metabolism and modulators of AKT signaling. The contrasting metabolic features revealed by our strategy could be used to metabolically target CLL lymphocyte subsets creating new therapeutic windows for this disease for mTORC1 or AMPK inhibitors. Indeed, we report that Metformin, a drug used to treat diabetes was selectively cytotoxic to Dasatinib sensitive samples. Ultimately, we suggest that a similar strategy could be applied to other cancer types by using Dasatinib and/or relevant tyrosine kinase inhibitors.

Telomerase contributes to fludarabine resistance in primary human leukemic lymphocytes.

We report that Imetelstat, a telomerase inhibitor that binds to the RNA component of telomerase (hTR), can sensitize primary CLL lymphocytes to fludarabine in vitro. This effect was observed in lymphocytes from clinically resistant cases and with cytogenetic abnormalities associated with bad prognosis. Imetelstat mediated-sensitization to fludarabine was not associated with telomerase activity, but with the basal expression of Ku80. Since both Imetelstat and Ku80 bind hTR, we assessed 1) if Ku80 and Imetelstat alter each other's binding to hTR in vitro and 2) the effect of an oligonucleotide complementary to the Ku binding site in hTR (Ku oligo) on the survival of primary CLL lymphocytes exposed to fludarabine. We show that Imetelstat interferes with the binding of Ku70/80 (Ku) to hTR and that the Ku oligo can sensitize CLL lymphocytes to FLU. Our results suggest that Ku binding to hTR may contribute to fludarabine resistance in CLL lmphocytes. This is the first report highlighting the potentially broad effectiveness of Imetelstat in CLL, and the potential biological and clinical implications of a functional interaction between Ku and hTR in primary human cancer cells.

Irinotecan and DNA-PKcs inhibitors synergize in killing of colon cancer cells.

This study sought to measure the degree of synergy induced by specific small molecule inhibitors of DNA-PK [NU7026 and IC486241 (ICC)], a major component of the non-homologous end-joining (NHEJ) pathway, with SN38 or oxaliplatin. Synergy between the DNA damaging drugs and the DNA-PK inhibitors was assessed using the sulforhodamine-B assay (SRB). Effects of drug combinations on cell cycle and DNA-PK activity were determined using flow cytometry and western blot analysis. DNA damage was assessed via comet assay and quantification of γH2AX. The role of homologous recombination repair (HRR) was determined by nuclear Rad51 protein levels and a GFP reporter recombination assay. Significant reductions in the IC(50) values of SN38 were observed at 5 and 10 µM of DNA-PK inhibitors. Moreover, at 1-2 µM (attainable concentrations with ICC in mice) these DNA-PKcs inhibitors demonstrated synergistic reductions in the IC(50) of SN38. Flow cytometric data indicated that SN38 and SN38 in combination with DNA-PKcs inhibitors showed dramatic G2/M arrest at 24 h. Furthermore, reduced phosphorylation of DNA-PKcs and increased DNA damage were observed at this time point with SN38 in combination with DNA-PKcs inhibitors as compared to cells treated with SN38 alone. SN38 alone and in the presence of ICC increased nuclear Rad51 protein levels. Furthermore, inhibition of DNA-PKcs increased HRR suggesting that NHEJ is a negative regulator of HRR. These data indicate that small molecule inhibitors of DNA-PKcs dramatically enhance the efficacy of SN38 in colon cancer cell lines.

Effects of the novel DNA dependent protein kinase inhibitor, IC486241, on the DNA damage response to doxorubicin and cisplatin in breast cancer cells.

The purpose of this study was to determine the degree to which the novel DNA-PKcs inhibitor, IC486241 (ICC), synergizes the cytotoxicity of DNA damaging agents in 3 genetically diverse breast cancer cell lines. The sulforhodamine B (SRB) assay was employed as a primary screening method to determine the in-vitro cytotoxicity and the degree of synergy of ICC in combination with the topoisomerase II inhibitor, doxorubicin, or the DNA cross linking agent, cisplatin. Molecular mechanisms underlying drug toxicity were probed using immunostaining and flow cytometry, as well as, the alkaline comet assay to detect DNA damage. In this study, improved cytotoxicity and significant synergy were observed with both anticancer agents in the presence of nontoxic concentrations of ICC. Moreover, ICC decreased doxorubicin-induced DNA-PKcs autophosphorylation on Ser2056 and increased doxorubicin-induced DNA fragmentation. In conclusion, the novel DNA-PKcs inhibitor, ICC, synergistically sensitized 3 breast cancer cell lines to doxorubicin and cisplatin. Enhanced efficacy of doxorubicin was achieved by inhibiting non-homologous end joining resulting in increased accumulation of DNA damage.

The effect of a DNA repair gene on cellular invasiveness: XRCC3 over-expression in breast cancer cells.

Over-expression of DNA repair genes has been associated with resistance to radiation and DNA-damage induced by chemotherapeutic agents such as cisplatin. More recently, based on the analysis of genome expression profiling, it was proposed that over-expression of DNA repair genes enhances the invasive behaviour of tumour cells. In this study we present experimental evidence utilizing functional assays to test this hypothesis. We assessed the effect of the DNA repair proteins known as X-ray complementing protein 3 (XRCC3) and RAD51, to the invasive behavior of the MCF-7 luminal epithelial-like and BT20 basal-like triple negative human breast cancer cell lines. We report that stable or transient over-expression of XRCC3 but not RAD51 increased invasiveness in both cell lines in vitro. Moreover, XRCC3 over-expressing MCF-7 cells also showed a higher tumorigenesis in vivo and this phenotype was associated with increased activity of the metalloproteinase MMP-9 and the expression of known modulators of cell-cell adhesion and metastasis such as CD44, ID-1, DDR1 and TFF1. Our results suggest that in addition to its' role in facilitating repair of DNA damage, XRCC3 affects invasiveness of breast cancer cell lines and the expression of genes associated with cell adhesion and invasion.

Therapeutic implications of Src independent calcium mobilization in diffuse large B-cell lymphoma.

We report that 38% of primary large B-cell lymphoma (DLBCL) tested expressed active Src family kinases, which are targeted by dasatinib. The expression of active Src family of kinases (SFK) in primary DLBCL tumors correlated with unfavorable prognostic markers such as Ki67 and Mum1. Using four DLBCL cell lines we found that: (1) sensitivity to dasatinib (but not imatinib) varied 400-fold; (2) dasatinib resistance was associated with distinct signaling profiles downstream of BCR activation. In particular, although Src family kinase phosphorylation was inhibited by 100-150 nM dasatinib in all cell lines, this failed to inhibit BCR-mediated Blnk phosphorylation, calcium signaling and proliferation in a dasatinib resistant cell line.

Admixture in Mexico City: implications for admixture mapping of type 2 diabetes genetic risk factors.

Admixture mapping is a recently developed method for identifying genetic risk factors involved in complex traits or diseases showing prevalence differences between major continental groups. Type 2 diabetes (T2D) is at least twice as prevalent in Native American populations as in populations of European ancestry, so admixture mapping is well suited to study the genetic basis of this complex disease. We have characterized the admixture proportions in a sample of 286 unrelated T2D patients and 275 controls from Mexico City and we discuss the implications of the results for admixture mapping studies. Admixture proportions were estimated using 69 autosomal ancestry-informative markers (AIMs). Maternal and paternal contributions were estimated from geographically informative mtDNA and Y-specific polymorphisms. The average proportions of Native American, European and, West African admixture were estimated as 65, 30, and 5%, respectively. The contributions of Native American ancestors to maternal and paternal lineages were estimated as 90 and 40%, respectively. In a logistic model with higher educational status as dependent variable, the odds ratio for higher educational status associated with an increase from 0 to 1 in European admixture proportions was 9.4 (95%, credible interval 3.8-22.6). This association of socioeconomic status with individual admixture proportion shows that genetic stratification in this population is paralleled, and possibly maintained, by socioeconomic stratification. The effective number of generations back to unadmixed ancestors was 6.7 (95% CI 5.7-8.0), from which we can estimate that genome-wide admixture mapping will require typing about 1,400 evenly distributed AIMs to localize genes underlying disease risk between populations of European and Native American ancestry. Sample sizes of about 2,000 cases will be required to detect any locus that contributes an ancestry risk ratio of at least 1.5.

Prevalence of dopamine and 5HT2C receptor polymorphisms in Amerindians and in an urban population from Argentina.

We report on the frequency of DRD2A1, DRD3A1, DRD4/2R-10R, and 5HT2CA1 variants in the population of the city of La Plata (Argentina) and in Amerindians from Argentina, Paraguay, and Chile. In the Amerindian sample, the prevalence of DRD2A1 and DRD4/4R variants were, respectively, significantly lower and significantly higher than frequencies reported in other Native Americans. Comparison of average allele and genotype frequencies between La Plata and Amerindians showed significant differences for 5HT2CA1 and DRD4. As La Plata is a population with predominant European and Amerindian components, we used mtDNA and Y-specific markers to subdivide the La Plata sample into two strata: Amerindian La Plata and non-Amerindian La Plata. Significant variations between the two strata were detected for DRD2A1, DRD3A1, and DRD4/4R allele frequencies, and for the homozygous DRD4/4R/4R genotype. Several controversial reports suggest a possible association between a variant of DRD and/or 5HT2C receptor genes and the clinical expression of several psychiatric disorders. We suggest that ethnic variations in the prevalence of the allelic forms of these genes may be a confounding factor to be taken into consideration in studies of association between dopaminergic and serotonergic receptor genotypes and neuropsychiatric and mood disorders.

Characterization of admixture in an urban sample from Buenos Aires, Argentina, using uniparentally and biparentally inherited genetic markers.

In this study we analyzed a sample of the urban population of La Plata, Argentina, using 17 mtDNA haplogroups, the DYS 199 Y-chromosome polymorphism, and 5 autosomal population-associated alleles (PAAs). The contribution of native American maternal lineages to the population of La Plata was estimated as 45.6%, whereas the paternal contribution was much lower (10.6%), clearly indicating directional mating. Regarding autosomal evidence of admixture, the relative European, native American, and West African genetic contributions to the gene pool of La Plata were estimated to be 67.55% (+/-2.7), 25.9% (+/-4.3), and 6.5% (+/-6.4), respectively. When admixture was calculated at the individual level, we found a low correlation between the ancestral contribution estimated with uniparental lineages and autosomal markers. Most of the individuals from La Plata with a native American mtDNA haplogroup or the DYS199*T native American allele show a genetic contribution at the autosomal level that can be traced primarily to Europe. The results of this study emphasize the need to use both uniparentally and biparentally inherited genetic markers to understand the history of admixed populations.

Identificación de amerindios por medio del análisis de ADN: su aplicación en los litigios por posesión de tierras, y otros temas legales y éticos / Identification of native amerindians through ADN analysis: its use in land possesion litigation and other legal and ethical matters

La mayoría de los países americanos reconocen los derechos fundamentales de las comunidades indígenas, tales como su existencia dentro de la sociedad, la aceptación de su cultura, identidad y lenguaje, la protección de las tierras que ocupan y la recuperación de las que reclaman como propias. Sin embargo los parámetros tradicionales empleados para definir a una comunidad o a un individuo como Americano Nativo suele ser ambiguos y generar numerosos litigios con respectos a la propiedad de las tierras. Durante la última década el hallazgo de marcadores de ADN específicos de los Americanos Nativos significó un gran aporte a los elementos utilizados para definir al indígena. El empleo de estos marcadores en los pleitos iniciados por los indígenas para obtener la propiedad de las tierras que habitan está produciendo el desarrollo de una nueva área de la biomedicina forense: la antropología molecular forense; pero está también generando nuevos conflictos de índole legal y ética. El objetivo de este trabajo es presentar algunas de las principales áreas de controversia y actuar como catalizador para iniciar un activo intercambio de ideas entre genetistas, bioeticistas, juristas, filósofos y fundamentalmente, representantes de los pueblos indígenas con el fin de encontrar respuesta a los problemas éticos-legales vinculados con las investigaciones y estudios en los cuales participan las comunidades indígenas