Polyglutamic acid-based Crosslinked Doxorubicin Nanogels as an Anti-Metastatic Treatment for Triple Negative Breast Cancer

PreprintTreatment of triple negative breast cancer (TNBC)-associated metastasis represents an unmet clinical need, and we lack effective therapeutics for a disease that exhibits high relapse rates and associates with poor patient outcomes. Advanced nanosized drug delivery systems may enhance the efficacy of first-line chemotherapeutics by altering drug pharmacokinetics and enhancing tumor/metastasis targeting to signif-icantly improve efficacy and safety. Herein, we propose the application of injectable poly-amino acid-based nanogels (NGs) as a versatile hydrophilic drug delivery platform for the treatment of TNBC lung metastasis. We prepared biocompatible and biodegradable cross-linked NGs from polyglutamic acid (PGA) loaded with the chemotherapeutic agent doxorubicin (DOX). Our optimized synthetic procedures generated NGs of ~100 nm in size and 25 wt% drug loading content that became rapidly internalized in TNBC cell lines and displayed IC50 values comparable to the free form of DOX. Importantly, PGA-DOX NGs significantly inhibited lung metastases and almost completely suppressed lymph node metastases in a spontaneously metastatic orthotopic mouse TNBC model. Overall, our newly developed PGA-DOX NGs represent a potentially effective therapeutic strategy for the treatment of TNBC metastases.A.S-H thanks MC IEF actions (Project 302717). We thank Dr. M. A. Molina for AFM experiments, Ser-vicio SEM Cordoba (Argentina), and Mario Soriano Navarro from the electron microscopy service for Cryo-TEM pictures at CIPF. The authors would also like to thank Dr. Stuart P. Atkinson for his collabo-ration in the revision of the manuscript. This work has been supported by the European Research Council (grant ERC-CoG-2014-648831 “MyNano”), by the Spanish Ministry of Science and Innovation (SAF2013-44848-R, SAF2016-80427-R, RTI2018-099227-B-I00), by a Marie Curie IEF (Project 302717), and the Bundesministerium für Bildung und Forschung (BMBF) through the NanoMatFutur award (13N12561). Part of the equipment employed in this work has been funded by Generalitat Valen-ciana and co-financed with FEDER funds (PO FEDER of Comunitat Valenciana 2014–2020

Tumor microenvironment-targeted poly-L-glutamic acid-based combination conjugate for enhanced triple negative breast cancer treatment

[EN] The intrinsic characteristics of the tumor microenvironment (TME), including acidic pH and overexpression of hydrolytic enzymes, offer an exciting opportunity for the rational design of TME-drug delivery systems (DDS). We developed and characterized a pH-responsive biodegradable poly-L-glutamic acid (PGA)-based combination conjugate family with the aim of optimizing anticancer effects. We obtained combination conjugates bearing Doxorubicin (Dox) and aminoglutethimide (AGM) with two Dox loadings and two different hydrazone pH sensitive linkers that promote the specific release of Dox from the polymeric backbone within the TME. Low Dox loading coupled with a short hydrazone linker yielded optimal effects on primary tumor growth, lung metastasis (-90% reduction), and toxicological profile in a preclinical metastatic triple-negative breast cancer (TNBC) murine model. The use of transcriptomic analysis helped us to identify the molecular mechanisms responsible for such results including a differential immunomodulation and cell death pathways among the conjugates. This data highlights the advantages of targeting the TME, the therapeutic value of polymer-based combination approaches, and the utility of -omits-based analysis to accelerate anticancer DDS.The authors would like to thank Dr. Stuart P. Atkinson for his collaboration in manuscript preparation and English revision, and Irene Borreda for essential immunohistological support. This work has been supported by the European Research Council (grant ERC-CoG-2014-648831 "MyNano") and the Spanish Ministry of Science and Innovation (CTQ2010-18195, SAF2013-44848-R, BES-2008-006801, IPT-2012-0712-010000, Programa I3, and BIO2015-71658-R). LBN is funded through a University of South Florida-Helmsley Foundation award. FHL is funded through NIH grant. Part of the equipment employed in this work has been funded by Generalitat Valenciana and co-financed with FEDER funds (PO FEDER of Comunitat Valenciana 2014-2020).Arroyo-Crespo, JJ.; Armiñán, A.; Charbonnier, D.; Balzano-Nogueira, L.; Huertas-López, F.; Martí, C.; Tarazona Campos, S.... (2018). Tumor microenvironment-targeted poly-L-glutamic acid-based combination conjugate for enhanced triple negative breast cancer treatment. Biomaterials. 186:8-21. https://doi.org/10.1016/j.biomaterials.2018.09.02382118

Characterization of triple‐negative breast cancer preclinical models provides functional evidence of metastatic progression

Triple‐negative breast cancer (TNBC), an aggressive, metastatic and recurrent breast cancer (BC) subtype, currently suffers from a lack of adequately described spontaneously metastatic preclinical models that faithfully reproduce the clinical scenario. We describe two preclinical spontaneously metastatic TNBC orthotopic murine models for the development of advanced therapeutics: an immunodeficient human MDA‐MB‐231‐Luc model and an immunocompetent mouse 4T1 model. Furthermore, we provide a broad range of multifactorial analysis for both models that could provide relevant information for the development of new therapies and diagnostic tools. Our comparisons uncovered differential growth rates, stromal arrangements and metabolic profiles in primary tumors, and the presence of cancer‐associated adipocyte infiltration in the MDA‐MB‐231‐Luc model. Histopathological studies highlighted the more rapid metastatic spread to the lungs in the 4T1 model following a lymphatic route, while we observed both homogeneous (MDA‐MB‐231‐Luc) and heterogeneous (4T1) metastatic spread to axillary lymph nodes. We encountered unique metabolomic signatures in each model, including crucial amino acids and cell membrane components. Hematological analysis demonstrated severe leukemoid and lymphoid reactions in the 4T1 model with the partial reestablishment of immune responses in the immunocompromised MDA‐MB‐231‐Luc model. Additionally, we discovered β‐immunoglobulinemia and increased basal levels of G‐CSF correlating with a metastatic switch, with G‐CSF also promoting extramedullary hematopoiesis (both models) and causing hepatosplenomegaly (4T1 model). Overall, we believe that the characterization of these preclinical models will foster the development of advanced therapeutic strategies for TNBC treatment, especially for the treatment of patients presenting both, primary tumors and metastatic spread

Higher small pulmonary artery and vein volume on computed tomography is associated with mortality in current and former smokers

Background: In chronic obstructive pulmonary disease (COPD), vascular alterations have been shown to contribute to hypoxia and pulmonary hypertension, but the independent contribution of small vessel abnormalities to mortality remains unclear. Methods: We quantified artery and vein dimensions on computed tomography (CT) down to 0.2 mm. Small vessel volumes (&lt;1 mmᴓ) were normalized by body surface area. In 7903 current and former smokers of the COPDGene study (53.2% male) the independent contribution of small artery and small vein volume to all-cause mortality was tested in multivariable Cox models. Additionally, we calculated the 95th percentile of small arteries and veins in 374 never smokers to create two groups: normal and high small artery or vein volume. We describe clinical, physiological and imaging characteristics of subjects with a high small artery and high small vein volume. Findings: Both high small artery and high small vein volumes were independently associated with mortality with an adjusted hazard ratio of 1.07 [1.01, 1.14] and 1.34 [1.21, 1.49] per mL/m2 increase, respectively. In COPDGene, 447 (5.7%) had high small artery volume and 519 (9.1%) subjects had high small vein volume and both had more emphysema, more air trapping and more severe coronary calcium.Interpretation:In smokers, abnormally high volumes in small arteries and veins are both relevant for mortality, which urges investigations into the aetiology of small pulmonary vessels and cardiac function in smokers. </p

Reactivity of neodymium carriers in deep sea sediments: Implications for boundary exchange and paleoceanography

The dissolved neodymium (Nd) isotopic distribution in the deep oceans is determined by continental weathering inputs, water mass advection, and boundary exchange between particulate and dissolved fractions. Reconstructions of past Nd isotopic variability may therefore provide evidence on temporal changes in continental weathering inputs and/or ocean circulation patterns over a range of timescales. However, such an approach is limited by uncertainty in the mechanisms and importance of the boundary exchange process, and the challenge in reliably recovering past seawater Nd isotopic composition (εNd) from deep sea sediments. This study addresses these questions by investigating the processes involved in particulate–solution interactions and their impact on Nd isotopes. A better understanding of boundary exchange also has wider implications for the oceanic cycling and budgets of other particle-reactive elements. Sequential acid-reductive leaching experiments at pH ∼2–5 on deep sea sediments from the western Indian Ocean enable us to investigate natural boundary exchange processes over a timescale appropriate to laboratory experiments. We provide evidence that both the dissolution of solid phases and exchange processes influence the εNd of leachates, which suggests that both processes may contribute to boundary exchange. We use major element and rare earth element (REE) data to investigate the pools of Nd that are accessed and demonstrate that sediment leachate εNd values cannot always be explained by admixture between an authigenic component and the bulk detrital component. For example, in core WIND 24B, acid-reductive leaching generates εNd values between −11 and −6 as a function of solution/solid ratios and leaching times, whereas the authigenic components have εNd ≈ −11 and the bulk detrital component has εNd ≈ −15. We infer that leaching in the Mascarene Basin accesses authigenic components and a minor radiogenic volcanic component that is more reactive than Madagascan-derived clays. The preferential mobilisation of such a minor component demonstrates that the Nd released by boundary exchange could often have a significantly different εNd composition than the bulk detrital sediment. These experiments further demonstrate certain limitations on the use of acid-reductive leaching to extract the εNd composition of the authigenic fraction of bulk deep sea sediments. For example, the detrital component may contain a reactive fraction which is also acid-extractible, while the incongruent nature of this dissolution suggests that it is often inappropriate to use the bulk detrital sediment elemental chemistry and/or εNd composition when assessing possible detrital contamination of leachates. Based on the highly systematic controls observed, and evidence from REE patterns on the phases extracted, we suggest two approaches that lead to the most reliable extraction of the authigenic εNd component and good agreement with foraminiferal-based approaches; either (i) leaching of sediments without a prior decarbonation step, or (ii) the use of short leaching times and low solution/solid ratios throughout

Characteristics of Severe Asthma Patients and Predictors of Asthma Control in the Swiss Severe Asthma Registry

Background: Asthma is a chronic airway disease, affecting over 300 million people worldwide. 5–10% of patients suffer from severe asthma and account for 50% of asthma-related financial burden. Availability of real-life data about the clinical course of severe asthma is insufficient. Objectives: The aims of this study were to characterize patients with severe asthma in Switzerland, enrolled in the Swiss Severe Asthma Registry (SSAR), and evaluate predictors for asthma control. Method: A descriptive characterisation of 278 patients was performed, who were prospectively enrolled in the registry until January 2022. Socio-demographic variables, comorbidities, diagnostic values, asthma treatment, and healthcare utilisation were evaluated. Groups of controlled and uncontrolled asthma according to the asthma control test were compared. Results: Forty-eight percent of patients were female and the mean age was 55.8 years (range 13–87). The mean body mass index (BMI) was 27.4 kg/m2 (±6). 10.8% of patients were current smokers. Allergic comorbidities occurred in 54.3% of patients, followed by chronic rhinosinusitis (46.4%) and nasal polyps (34.1%). According to the ACT score, 54.7% had well controlled, 16.2% partly controlled and 25.9% uncontrolled asthma. The most common inhalation therapy was combined inhaled corticosteroids/long-acting β2-agonists (78.8%). Biologics were administered to 81.7% of patients and 19.1% received oral steroids. The multivariable analysis indicated that treatment with biologics was positively associated with asthma control whereas higher BMI, oral steroids, exacerbations, and COPD were negative predictors for asthma control. Conclusion: Biologics are associated with improved control in severe asthma. Further studies are required to complete the picture of severe asthma in order to provide improved care for those patients

Anticancer activity driven by drug linker modification in a polyglutamic acid-based combination-drug conjugate

Combination nanotherapies for the treatment of breast cancer permits synergistic drug targeting of multiple pathways. However, poor carrier degradability, poor synergism of the combined drugs, low drug release regulation, and a lack of control on final macromolecule solution conformation (which drives the biological fate) limit the application of this strategy. The present study describes the development of a family of drug delivery systems composed of chemotherapeutic (doxorubicin) and endocrine therapy (aromatase inhibitor aminoglutethimide) agents conjugated to a biodegradable poly‐l‐glutamic acid backbone via various linking moieties. Data from in vitro cytotoxicity and drug release assessments and animal model validation select a conjugate family member with optimal biological performance. Exhaustive physicochemical characterization in relevant media (including the study of secondary structure, size measurements, and detailed small‐angle neutron scattering analysis) correlates biological data with the intrinsic supramolecular characteristics of the conjugate. Overall, this study demonstrates how a small flexible Gly linker can modify the spatial conformation of the entire polymer–drug conjugate, promote the synergistic release of both drugs, and significantly improve biological activity. These findings highlight the need for a deeper understanding of polymer–drug conjugates at supramolecular level to allow the design of more effective polymer–drug conjugates

Partial loss of heterozygosity events at the mutated gene in tumors from MLH1/MSH2 large genomic rearrangement carriers

<p>Abstract</p> <p>Background</p> <p>Depending on the population studied, large genomic rearrangements (LGRs) of the mismatch repair (<it>MMR</it>) genes constitute various proportions of the germline mutations that predispose to hereditary non-polyposis colorectal cancer (HNPCC). It has been reported that loss of heterozygosity (LOH) at the LGR region occurs through a gene conversion mechanism in tumors from <it>MLH1</it>/<it>MSH2 </it>deletion carriers; however, the converted tracts were delineated only by extragenic microsatellite markers. We sought to determine the frequency of LGRs in Slovak HNPCC patients and to study LOH in tumors from LGR carriers at the LGR region, as well as at other heterozygous markers within the gene to more precisely define conversion tracts.</p> <p>Methods</p> <p>The main <it>MMR </it>genes responsible for HNPCC, <it>MLH1</it>, <it>MSH2</it>, <it>MSH6</it>, and <it>PMS2</it>, were analyzed by MLPA (multiplex ligation-dependent probe amplification) in a total of 37 unrelated HNPCC-suspected patients whose <it>MLH1/MSH2 </it>genes gave negative results in previous sequencing experiments. An LOH study was performed on six tumors from LGR carriers by combining MLPA to assess LOH at LGR regions and sequencing to examine LOH at 28 SNP markers from the <it>MLH1 </it>and <it>MSH2 </it>genes.</p> <p>Results</p> <p>We found six rearrangements in the <it>MSH2 </it>gene (five deletions and dup5-6), and one aberration in the <it>MLH1 </it>gene (del5-6). The <it>MSH2 </it>deletions were of three types (del1, del1-3, del1-7). We detected LOH at the LGR region in the single <it>MLH1 </it>case, which was determined in a previous study to be LOH-negative in the intragenic D3S1611 marker. Three tumors displayed LOH of at least one SNP marker, including two cases that were LOH-negative at the LGR region.</p> <p>Conclusion</p> <p>LGRs accounted for 25% of germline <it>MMR </it>mutations identified in 28 Slovakian HNPCC families. A high frequency of LGRs among the <it>MSH2 </it>mutations provides a rationale for a MLPA screening of the Slovakian HNPCC families prior scanning by DNA sequencing. LOH at part of the informative loci confined to the <it>MLH1 </it>or <it>MSH2 </it>gene (heterozygous LGR region, SNP, or microsatellite) is a novel finding and can be regarded as a partial LOH. The conversion begins within the gene, and the details of conversion tracts are discussed for each case.</p

Correction to: Macromolecular interactions in vitro, comparing classical and novel approaches

Correction to: European Biophysics Journal https://doi.org/10.1007/s00249-021-01517-5Peer reviewe

Regional and scale-specific effects of land use on amphibian diversity [poster]

Background/Question/Methods Habitat loss and degradation influence amphibian distributions and are important drivers of population declines. Our previous research demonstrated that road disturbance, development and wetland area consistently influence amphibian richness across regions of the U.S. Here, we examined the relative importance of these factors in different regions and at multiple spatial scales. Understanding the scales at which habitat disturbance may be affecting amphibian distributions is important for conservation planning. Specifically, we asked: 1) Over what spatial scales do distinct landscape features affect amphibian richness? and 2) Do road types (non-rural and rural) have similar effects on amphibian richness? This is the second year of a collaborative, nationwide project involving 11 U.S. colleges integrated within undergraduate biology curricula. We summarized North American Amphibian Monitoring Program data in 13 Eastern and Central U.S states and used geographic information systems to extract landscape data for 471 survey locations. We developed models to quantify the influence of landscape variables on amphibian species richness and site occupancy across five concentric buffers ranging from 300m to 10,000m. Results/Conclusions Across spatial scales, development, road density and agriculture were the best predictors of amphibian richness and site occupancy by individual species. Across regions, we found that scale did not exert a large influence on how landscape features influenced amphibian richness as effects were largely comparable across buffers. However, development and percent impervious surface had stronger influence on richness at smaller spatial scales. Richness was lower at survey locations with higher densities of non-rural and rural roads, and non-rural road density had a larger negative effect at smaller scales. Within regions, landscape features driving patterns of species richness varied. The scales at which these factors were associated with richness were highly variable within regions, suggesting the scale effects may be region specific. Our project demonstrates that networks of undergraduate students can collaborate to compile and analyze large ecological data sets, while engaging students in authentic and inquiry-based learning in landscape-scale ecology