Crosslinking of CD38 Receptors Triggers Apoptosis of Malignant B Cells

Recently, we designed an inventive paradigm in nanomedicine—drug-free macromolecular therapeutics (DFMT). The ability of DFMT to induce apoptosis is based on biorecognition at cell surface, and crosslinking of receptors without the participation of low molecular weight drugs. The system is composed of two nanoconjugates: a bispecific engager, antibody or Fab’ fragment—morpholino oligonucleotide (MORF1) conjugate; the second nanoconjugate is a multivalent effector, human serum albumin (HSA) decorated with multiple copies of complementary MORF2. Here, we intend to demonstrate that DFMT is a platform that will be effective on other receptors than previously validated CD20. We appraised the impact of daratumumab (DARA)- and isatuximab (ISA)-based DFMT to crosslink CD38 receptors on CD38+ lymphoma (Raji, Daudi) and multiple myeloma cells (RPMI 8226, ANBL-6). The biological properties of DFMTs were determined by flow cytometry, confocal fluorescence microscopy, reactive oxygen species determination, lysosomal enlargement, homotypic cell adhesion, and the hybridization of nanoconjugates. The data revealed that the level of apoptosis induction correlated with CD38 expression, the nanoconjugates meet at the cell surface, mitochondrial signaling pathway is strongly involved, insertion of a flexible spacer in the structure of the macromolecular effector enhances apoptosis, and simultaneous crosslinking of CD38 and CD20 receptors increases apoptosis.

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Functional Multi-Omics Reveals Genetic and Pharmacologic Regulation of Surface CD38 in Multiple Myeloma

Blood ◽

10.1182/blood-2021-147007 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 2648-2648

Author(s):

Priya Choudhry ◽

Olivia Gugliemini ◽

Huimin Geng ◽

Vishesh Sarin ◽

Letitia Sarah ◽

...

Keyword(s):

Transcription Factor ◽

Immune System ◽

Cell Surface ◽

Surface Protein ◽

Surface Expression ◽

Surface Proteins ◽

Cd38 Expression ◽

Genome Wide ◽

Rpmi 8226 ◽

Pharmacologic Regulation

Abstract Background: CD38 is a surface ectoenzyme expressed at high levels on myeloma plasma cells and is the target for the monoclonal antibodies (mAbs) daratumumab and isatuximab. These antibodies have multiple mechanisms of action, primarily involving recruiting and modulating components of the immune system, but they may also carry direct anti-tumor effects. CD38 density on tumor cells is an important determinant of mAb efficacy while CD38 is lost after mAb treatment. Several small molecules have been found to increase tumor surface CD38, with the goal of boosting mAb efficacy in a co-treatment strategy. However, we do not yet have a broad global sense of the transcriptional or post-transcriptional networks that most strongly impact CD38 expression. There may be alternate strategies to even more potently increase CD38 expression that have not yet been identified. Furthermore, prior clinical studies showed that CD38 downregulation after daratumumab treatment was accompanied by increases in the complement inhibitors CD55 and CD59. Are there other features of myeloma surface remodeling driven by CD38 downregulation? Here we sought to extend our currently limited insight into CD38 surface expression by using a multi-omics approach. Methods: Genome-wide CRISPR interference screening was performed in RPMI-8226 cells stably expressing the dCas9-KRAB fusion protein. Cells were grown for 14 days after library transduction, flow-sorted on the top and bottom 25% of CD38 surface expression, and sgRNA's deep sequenced. Antibody-dependent cellular cytoxicity assays were performed with NK92-CD16 cells. Cell surface proteomics was performed using N-glycoprotein cell surface capture in triplicate. Phosphoproteomics was performed used immobilized metal affinity chromatography in triplicate. Murine studies were performed in NSG mice under approved IACUC-approved institutional protocols. Results: A genome-wide CRISPR-interference screen in RPMI-8226 cells demonstrated that transcriptional and epigenetic factors played the most prominent role in surface CD38 regulation (Fig. 1A). One of the genes that when knocked down led to greatest surface CD38 increase was RARA. This finding supports the promise of all-trans retinoic acid (ATRA), which leads to RARA degradation, as a potent agent to induce CD38 upregulation. Validation of additional screen hits TLE3 and HEXIM1 also illustrated that these negative regulatory transcription factors suppress CD38 expression at baseline (not shown). We found the transcription factor SPI1 to be a prominent positive regulator of CD38. SPI1 knockdown led to daratumumab resistance both in vitro and in vivo, similar to the resistance observed after CD38 knockdown (not shown). Analysis of myeloma patient ATAC-seq data, assessing transcription factor motifs present at the CD38 locus, combined with a predictive machine learning model, further identified XBP1 as one of the most potent transcriptional regulators of CD38 (Fig. 1B). We next used "antigen escape profiling" - knockdown of CD38 followed by unbiased cell surface proteomics - to mimic surface alterations in the context of CD38 loss. We found minimal changes in other cell surface proteins beyond CD38 (Fig. 1C), indicating the CD38 loss alone is not sufficient to remodel the myeloma surfaceome. This finding also supports the hypothesis that complement or other immune system interactions are necessary to lead to other myeloma surface protein alterations. In a parallel analysis of pharmacologic regulation, we also used cell surface proteomics integrated with RNA-seq to demonstrate that ATRA leads to few other surface protein changes beyond CD38 (not shown). In contrast, other molecules, such as azacytidine and panobinostat, led to broader changes across many more surface proteins, showing a lack of specificity when driving CD38 upregulation. Finally, unbiased phosphoproteomics revealed partial inhibition of the MAP kinase pathway after daratumumab binding (Fig. 1D). This result may comprise a direct anti-proliferative effect of anti-CD38 therapeutic antibody engagement in myeloma. Conclusions: Our work provides a resource to design strategies to enhance efficacy of CD38-targeting immunotherapies in myeloma. Our approach also outlines a broad multi-omic strategy to evaluate surface and transcriptional regulation of other key immunotherapeutic targets in hematologic malignancies. Figure 1 Figure 1. Disclosures Choudhry: Genentech: Current Employment, Current equity holder in publicly-traded company. Ramkumar: Senti Biosciences: Current Employment, Current holder of individual stocks in a privately-held company.

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Histological, Biomechanical, and Biological Properties of Genipin-Crosslinked Decellularized Peripheral Nerves

International Journal of Molecular Sciences ◽

10.3390/ijms22020674 ◽

2021 ◽

Vol 22 (2) ◽

pp. 674

Author(s):

Óscar Darío García-García ◽

Marwa El Soury ◽

David González-Quevedo ◽

David Sánchez-Porras ◽

Jesús Chato-Astrain ◽

...

Keyword(s):

Ex Vivo ◽

Nerve Repair ◽

Wistar Rat ◽

Biomechanical Properties ◽

Biological Properties ◽

Suitable Alternative ◽

Promising Alternative ◽

Histological Pattern ◽

The Impact

Acellular nerve allografts (ANGs) represent a promising alternative in nerve repair. Our aim is to improve the structural and biomechanical properties of biocompatible Sondell (SD) and Roosens (RS) based ANGs using genipin (GP) as a crosslinker agent ex vivo. The impact of two concentrations of GP (0.10% and 0.25%) on Wistar rat sciatic nerve-derived ANGs was assessed at the histological, biomechanical, and biocompatibility levels. Histology confirmed the differences between SD and RS procedures, but not remarkable changes were induced by GP, which helped to preserve the nerve histological pattern. Tensile test revealed that GP enhanced the biomechanical properties of SD and RS ANGs, being the crosslinked RS ANGs more comparable to the native nerves used as control. The evaluation of the ANGs biocompatibility conducted with adipose-derived mesenchymal stem cells cultured within the ANGs confirmed a high degree of biocompatibility in all ANGs, especially in RS and RS-GP 0.10% ANGs. Finally, this study demonstrates that the use of GP could be an efficient alternative to improve the biomechanical properties of ANGs with a slight impact on the biocompatibility and histological pattern. For these reasons, we hypothesize that our novel crosslinked ANGs could be a suitable alternative for future in vivo preclinical studies.

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Earthworms as an Ecological Indicator of Soil Recovery after Mechanized Logging Operations in Mixed Beech Forests

Forests ◽

10.3390/f12010018 ◽

2020 ◽

Vol 12 (1) ◽

pp. 18

Author(s):

Hadi Sohrabi ◽

Meghdad Jourgholami ◽

Mohammad Jafari ◽

Farzam Tavankar ◽

Rachele Venanzi ◽

...

Keyword(s):

Soil Properties ◽

Soil Depth ◽

Biological Properties ◽

Economic Benefits ◽

Ecological Indicator ◽

Traffic Intensity ◽

Soil Biological Properties ◽

Density And Biomass ◽

Soil Recovery ◽

The Impact

Soil damage caused by logging operations conducted to obtain and maximize economic benefits has been established as having long-term effects on forest soil quality and productivity. However, a comprehensive study of the impact of logging operations on earthworms as a criterion for soil recovery has never been conducted in the Hyrcanian forests of Iran. The aim of this study was to determine the changes in soil biological properties (earthworm density and biomass) and its recovery process under the influence of traffic intensity, slope and soil depth in various intervals according to age after logging operations. Soil properties were compared among abandoned skid trails with different ages (i.e., 3, 10, 20, and 25 years) and an undisturbed area. The results showed that earthworm density and biomass in the high traffic intensity and slope class of 20–30% at the 10–20 cm depth of the soil had the lowest value compared to the other treatments. Twenty-five years after the logging operations, the earthworm density at soil depth of 0–10 and 10–20 cm was 28.4% (0.48 ind. m−2) and 38.6% (0.35 ind. m−2), which were less than those of the undisturbed area, respectively. Meanwhile, the earthworm biomass at a soil depth of 0–10 and 10–20 cm was 30.5% (2.05 mg m−2) and 40.5% (1.54 mg m−2) less than the values of the undisturbed area, respectively. The earthworm density and biomass were positively correlated with total porosity, organic carbon and nitrogen content, while negatively correlated with soil bulk density and C/N ratio. According to the results, 25 years after logging operations, the earthworm density and biomass on the skid trails were recovered, but they were significantly different with the undisturbed area. Therefore, full recovery of soil biological properties (i.e., earthworm density and biomass) takes more than 25 years. The conclusions of our study reveal that the effects of logging operations on soil properties are of great significance, and our understanding of the mechanism of soil change and recovery demand that harvesting operations be extensively and properly implemented.

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Constitutive and Regulated Shedding of Soluble FGF Receptors Releases Biologically Active Inhibitors of FGF-2

International Journal of Molecular Sciences ◽

10.3390/ijms22052712 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2712

Author(s):

Anne Hanneken ◽

Maluz Mercado ◽

Pamela Maher

Keyword(s):

Cell Surface ◽

Ligand Binding ◽

Cellular Proliferation ◽

Biological Activities ◽

Biological Properties ◽

Matrix Metalloproteases ◽

Biologically Active ◽

Ectodomain Shedding ◽

High Affinity

The identification of soluble fibroblast growth factor (FGF) receptors in blood and the extracellular matrix has led to the prediction that these proteins modulate the diverse biological activities of the FGF family of ligands in vivo. A recent structural characterization of the soluble FGF receptors revealed that they are primarily generated by proteolytic cleavage of the FGFR-1 ectodomain. Efforts to examine their biological properties are now focused on understanding the functional consequences of FGFR-1 ectodomain shedding and how the shedding event is regulated. We have purified an FGFR-1 ectodomain that is constitutively cleaved from the full-length FGFR-1(IIIc) receptor and released into conditioned media. This shed receptor binds FGF-2; inhibits FGF-2-induced cellular proliferation; and competes with high affinity, cell surface FGF receptors for ligand binding. FGFR-1 ectodomain shedding downregulates the number of high affinity receptors from the cell surface. The shedding mechanism is regulated by ligand binding and by activators of PKC, and the two signaling pathways appear to be independent of each other. Deletions and substitutions at the proposed cleavage site of FGFR-1 do not prevent ectodomain shedding. Broad spectrum inhibitors of matrix metalloproteases decrease FGFR-1 ectodomain shedding, suggesting that the enzyme responsible for constitutive, ligand-activated, and protein kinase C-activated shedding is a matrix metalloprotease. In summary, shedding of the FGFR-1 ectodomain is a highly regulated event, sharing many features with a common system that governs the release of diverse membrane proteins from the cell surface. Most importantly, the FGFR ectodomains are biologically active after shedding and are capable of functioning as inhibitors of FGF-2.

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Liver X Receptor Nuclear Receptors Are Transcriptional Regulators of Dendritic Cell Chemotaxis

Molecular and Cellular Biology ◽

10.1128/mcb.00534-17 ◽

2018 ◽

Vol 38 (10) ◽

Cited By ~ 13

Author(s):

Susana Beceiro ◽

Attila Pap ◽

Zsolt Czimmerer ◽

Tamer Sallam ◽

Jose A. Guillén ◽

...

Keyword(s):

Nuclear Receptors ◽

Low Density Lipoprotein ◽

Myeloid Cells ◽

Density Lipoprotein ◽

Loss Of Function ◽

Transcriptional Responses ◽

Cd38 Expression ◽

The Impact

ABSTRACTThe liver X receptors (LXRs) are ligand-activated nuclear receptors with established roles in the maintenance of lipid homeostasis in multiple tissues. LXRs exert additional biological functions as negative regulators of inflammation, particularly in macrophages. However, the transcriptional responses controlled by LXRs in other myeloid cells, such as dendritic cells (DCs), are still poorly understood. Here we used gain- and loss-of-function models to characterize the impact of LXR deficiency on DC activation programs. Our results identified an LXR-dependent pathway that is important for DC chemotaxis. LXR-deficient mature DCs are defective in stimulus-induced migrationin vitroandin vivo. Mechanistically, we show that LXRs facilitate DC chemotactic signaling by regulating the expression of CD38, an ectoenzyme important for leukocyte trafficking. Pharmacological or genetic inactivation of CD38 activity abolished the LXR-dependent induction of DC chemotaxis. Using the low-density lipoprotein receptor-deficient (LDLR−/−) LDLR−/−mouse model of atherosclerosis, we also demonstrated that hematopoietic CD38 expression is important for the accumulation of lipid-laden myeloid cells in lesions, suggesting that CD38 is a key factor in leukocyte migration during atherogenesis. Collectively, our results demonstrate that LXRs are required for the efficient emigration of DCs in response to chemotactic signals during inflammation.

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Assessing the Potential of Nontraditional Water Sources for Landscape Irrigation

HortTechnology ◽

10.21273/horttech04023-18 ◽

2018 ◽

Vol 28 (4) ◽

pp. 436-444 ◽

Cited By ~ 2

Author(s):

Raul I. Cabrera ◽

James E. Altland ◽

Genhua Niu

Keyword(s):

Best Management Practices ◽

Urban Areas ◽

Management Practices ◽

Urban Landscape ◽

Biological Properties ◽

Water Sources ◽

Long Term Effects ◽

Natural Ecosystems ◽

Aesthetic Quality ◽

The Impact

Scarcity and competition for good quality and potable water resources are limiting their use for urban landscape irrigation, with several nontraditional sources being potentially available for these activities. Some of these alternative sources include rainwater, stormwater, brackish aquifer water, municipal reclaimed water (MRW), air-conditioning (A/C) condensates, and residential graywater. Knowledge on their inherent chemical profile and properties, and associated regional and temporal variability, is needed to assess their irrigation quality and potential short- and long-term effects on landscape plants and soils and to implement best management practices that successfully deal with their quality issues. The primary challenges with the use of these sources are largely associated with high concentrations of total salts and undesirable specific ions [sodium (Na), chloride (Cl), boron (B), and bicarbonate (HCO3−) alkalinity]. Although the impact of these alternative water sources has been largely devoted to human health, plant growth and aesthetic quality, and soil physicochemical properties, there is emergent interest in evaluating their effects on soil biological properties and in natural ecosystems neighboring the urban areas where they are applied.

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Impact of sewage water irrigation on agricultural soil

10.37281/drcsf/2.2.7 ◽

2021 ◽

pp. 141-148

Keyword(s):

Industrial Development ◽

Municipal Wastewater ◽

Microbial Biomass Carbon ◽

Biological Properties ◽

Sewage Water ◽

Viable Count ◽

Steady Increase ◽

Available Nitrogen ◽

The Impact ◽

Flooded Field

The rapidly increasing population growth and the steady increase in water requirements for agricultural and industrial development have placed severe stress on the water resources available and the long term use of sewage water for irrigation highly affects soil properties. In this study soil samples were collected from a cauliflower field prior and after sewage water irrigation, and the impact of sewage water irrigation on physical, chemical, and biological properties of soil was compared. For this, tested were pH, Electrical Conductivity (EC), Organic Carbon, available Nitrogen, Phosphorus, Potassium, Calcium, Magnesium, Zinc, Iron, Copper, Manganese, and microbial activity. Soil microbial biomass carbon, basal soil respiration, total viable count of bacteria, coliform population, Pseudomonas species, and Azotobacter significantly increased after sewage water irrigation as compared to prior to irrigation. Nevertheless, the bulk density and Rhizobium species of the soil flooded with sewage water was decreased relative to the same characteristics prior to irrigation. Cauliflower yield was significantly increased when sewage water flooded field as compared to the tube well water flooded field (i.e., water delivered via an iron pipe). Escherichia coli contamination was greater in sewage water and groundwater that can pose health risks for the nearby communities, to farmers and consumer of farm products. Hence, the efficient use of sewage and municipal wastewater successfully increase water resource for irrigation and may help in expanding agricultural production. But excessive use of sewage water may also affect the soil flora and fertility.

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Using bio-manipulation to optimise nutrient management within intensive farm systems

10.31219/osf.io/esdp2 ◽

2021 ◽

Author(s):

Vito Abbruzzese

Keyword(s):

Nutrient Availability ◽

Nutrient Management ◽

Biological Properties ◽

Retention Capacity ◽

Know How ◽

System P ◽

Livestock Farms ◽

Farm System ◽

The Impact

The research project aims to enhance organic nutrient management in livestock farms using microbial and enzyme inoculations, with a particular focus on the phosphorus biogeochemical cycle. In order to do this the first approach consists of characterising the chemical and biological properties of farm slurries as a baseline to evaluate possible amendments of the intrinsic properties of the slurry. Consequently, it is pivotal to consider properties such as plant nutrients, i.e., phosphorus, nitrogen and potassium, as well as the microbial community within the slurry. Likewise, attention needs to be paid to soil chemical and biological properties, e.g. pH, salinity and organic matter, as well as to the variety of organisms inhabiting the soil, in order to determine the impact of inoculation on phosphorus cycling and nutrient availability for plant use. Furthermore, it is important to know how soil and its productivity may be influenced by the addition of the inoculated slurry. Of particular interest are also the soil properties which have an effect on plant growth. The pH of soil and, notably, nutrient availability and retention capacity are some of the features on which to direct the research in order to assess the quality of soil and, as a result, the production of a grass crop in livestock farms. The characterisation of these properties will be performed using a variety of approaches, beginning with analysis at laboratory- and mesocosm-scales and progressing to a fieldwork approach in order to evaluate the results directly in a farm system.

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Combination of in vivo phage therapy data with in silico model highlights key parameters for treatment efficacy

10.1101/2021.03.04.433924 ◽

2021 ◽

Author(s):

Raphaelle Delattre ◽

Jeremy Seurat ◽

Feyrouz Haddad ◽

Thu-Thuy Nguyen ◽

Baptiste Gaborieau ◽

...

Keyword(s):

Phage Therapy ◽

Biological Properties ◽

General Strategy ◽

In Vivo Experiments ◽

Bacterial Dynamics ◽

Optimal Dosing ◽

Dosing Regimens ◽

The Impact

The clinical (re)development of phage therapy to treat antibiotic resistant infections requires grasping specific biological properties of bacteriophages (phages) as antibacterial. However, identification of optimal dosing regimens is hampered by the poor understanding of phage-bacteria interactions in vivo. Here we developed a general strategy coupling in vitro and in vivo experiments with a mathematical model to characterize the interplay between phage and bacterial dynamics during pneumonia induced by a pathogenic strain of Escherichia coli. The model estimates some key parameters for phage therapeutic efficacy, in particular the impact of dose and route of administration on phage dynamics and the synergism of phage and the innate immune response on the bacterial clearance rate. Simulations predict a low impact of the intrinsic phage characteristics in agreement with the current semi-empirical choices of phages for compassionate treatments. Model-based approaches will foster the deployment of future phage therapy clinical trials.

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The importance of sanitary microbiological indices in the evaluation of epidemiological safety of water use in conditions of chemical contamination of water

Hygiene and Sanitation ◽

10.18821/0016-9900-2016-95-10-934-938 ◽

2019 ◽

Vol 95 (10) ◽

pp. 934-938 ◽

Cited By ~ 2

Author(s):

Yu. A. Rakhmanin ◽

L. V. Ivanova ◽

T. Z. Artemova ◽

E. K. Gipp ◽

Anzhelika V. Zagainova ◽

...

Keyword(s):

Water Use ◽

Inhibitory Effect ◽

Biological Properties ◽

Chemical Substance ◽

Water Bodies ◽

Chemical Pollution ◽

Chemical Contamination ◽

Natural Conditions ◽

The Impact ◽

The Given

The increasing chemicalization of production and life leads to the pollution of water bodies by chemicals, the effect of which on the micro - and macro - organisms is poorly understood. This section of the study in sanitary bacteriology is becoming ever more topical and is an important task of modern hygienic science. One of complicacies of the study of the problem is related with the fact that the presence of only experimental data fails to be sufficient, as the impact of any given chemical substance on different bacteria in the experiment does not mean that under natural conditions, similar results will be obtained. One reason for this may be the inhibitory effect of the given chemical on biological properties of bacteria, while in field conditions in the water several chemicals interacting with each other can exist. In this regard, the aim of the work was to assess the indicator value of sanitary and microbiological indices of epidemic hazard of water use in conditions of chemical pollution of surface water bodies.

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