Cellular and humoral immune response to SARS-CoV-2 mRNA vaccines in patients treated with either Ibrutinib or Rituximab.

Patients treated with B-cell-targeting therapies like Rituximab or Ibrutinib have decreased serological response to various vaccines. In this study, we tested serological and cellular response to SARS-CoV-2 mRNA vaccines in 16 patients treated with Ibrutinib, 16 treated with maintenance Rituximab, 18 patients with chronic lymphocytic leukaemia (CLL) with watch and wait status and 21 healthy volunteers. In comparison with the healthy volunteers, where serological response was achieved by 100% subjects, patients on B-cell-targeting therapy (Ibrutinib and Rituximab) had their response dramatically impaired. The serological response was achieved in 0% of Rituximab treated, 18% of Ibrutinib treated and 50% of untreated CLL patients. Cell-mediated immunity analysed by the whole blood Interferon-γ Release immune Assay developed in 80% of healthy controls, 62% of Rituximab treated, 75% of Ibrutinib treated and 55% of untreated CLL patients. The probability of cell-mediated immune response development negatively correlates with disease burden mainly in CLL patients. Our study shows that even though the serological response to SARS-CoV-2 vaccine is severely impaired in patients treated with B-cell-targeting therapy, the majority of these patients develop sufficient cell-mediated immunity. The vaccination of these patients therefore might be meaningful in terms of protection against SARS-CoV-2 infection.

Fluorinated diselenide nanoparticles for radiosensitizing therapy of cancer.

Radiation resistance of cancer cells represents one of the major challenges in cancer treatment. The novel self-assembled fluoralkylated diselenide nanoparticles (fluorosomes) based on seleno-l-cystine (17FSe2) possess redox-active properties that autocatalytically decompose hydrogen peroxide (H2O2) and oxidize the intracellular glutathione (GSH) that results in regulation of cellular oxidative stress. Alkylfluorinated diselenide nanoparticles showed a significant cytotoxic and radiosensitizing effect on cancer cells. The EL-4 tumor-bearing C56BL/6 mice treated with 17FSe2 followed by fractionated radiation treatment (4 × 2Gy) completely suppressed tumor growth. Our results suggest that described diselenide system behaves as a potent radiosensitizer agent targeting tumor growth and preventing tumor recurrence.

Photoacoustic Properties of Polypyrrole Nanoparticles.

Photoacoustic imaging, an emerging modality, provides supplemental information to ultrasound imaging. We investigated the properties of polypyrrole nanoparticles, which considerably enhance contrast in photoacoustic images, in relation to the synthesis procedure and to their size. We prepared polypyrrole nanoparticles by water-based redox precipitation polymerization in the presence of ammonium persulphate (ratio nPy:nOxi 1:0.5, 1:1, 1:2, 1:3, 1:5) or iron(III) chloride (nPy:nOxi 1:2.3) acting as an oxidant. To stabilize growing nanoparticles, non-ionic polyvinylpyrrolidone was used. The nanoparticles were characterized and tested as a photoacoustic contrast agent in vitro on an imaging platform combining ultrasound and photoacoustic imaging. High photoacoustic signals were obtained with lower ratios of the oxidant (nPy:nAPS ≥ 1:2), which corresponded to higher number of conjugated bonds in the polymer. The increasing portion of oxidized structures probably shifted the absorption spectra towards shorter wavelengths. A strong photoacoustic signal dependence on the nanoparticle size was revealed; the signal linearly increased with particle surface. Coated nanoparticles were also tested in vivo on a mouse model. To conclude, polypyrrole nanoparticles represent a promising contrast agent for photoacoustic imaging. Variations in the preparation result in varying photoacoustic properties related to their structure and allow to optimize the nanoparticles for in vivo imaging.

Hematopoiesis Remains Permissive to Bone Marrow Transplantation After Expansion of Progenitors and Resumption of Blood Cell Production.

The immense regenerative power of hematopoietic tissue stems from the activation of the immature stem cells and the progenitor cells. After partial damage, hematopoiesis is reconstituted through a period of intense regeneration when blood cell production originates from erythro-myeloid progenitors in the virtual absence of stem cells. Since the damaged hematopoiesis can also be reconstituted from transplanted hematopoietic cells, we asked whether this also leads to the transient state when activated progenitors initially execute blood cell production. We first showed that the early reconstitution of hematopoiesis from transplanted cells gives rise to extended populations of developmentally advanced but altered progenitor cells, similar to those previously identified in the bone marrow regenerating from endogenous cells. We then identified the cells that give rise to these progenitors after transplantation as LSK CD48- cells. In the submyeloablative irradiated host mice, the transplanted LSK CD48- cells preferably colonized the spleen. Unlike the endogenous hematopoiesis reconstituting cells, the transplanted whole bone marrow cells and sorted LSK CD48- cells had greater potential to differentiate to B-lymphopoiesis. Separate transplantation of the CD150- and CD150+ subsets of LSK CD48- cells suggested that CD150- cells had a greater preference to B-lymphopoiesis than CD150+ cells. In the intensively regenerating hematopoiesis, the CD71/Sca-1 plot of immature murine hematopoietic cells revealed that the expanded populations of altered myeloid progenitors were highly variable in the different places of hematopoietic tissues. This high variability is likely caused by the heterogeneity of the hematopoiesis supporting stroma. Lastly, we demonstrate that during the period when active hematopoiesis resumes from transplanted cells, the hematopoietic tissues still remain highly permissive for further engraftment of transplanted cells, particularly the stem cells. Thus, these results provide a rationale for the transplantation of the hematopoietic stem cells in successive doses that could be used to boost the transplantation outcome.

Enhanced Antitumor Efficacy through an "AND gate" Reactive Oxygen-Species-Dependent pH-Responsive Nanomedicine Approach.

Anticancer drug delivery strategies are designed to take advantage of the differential chemical environment in solid tumors independently, or to high levels of reactive oxygen species (ROS) or to low pH, compared to healthy tissue. Here, the design and thorough characterization of two functionalizable "AND gate" multiresponsive (MR) block amphiphilic copolymers are reported, aimed to take full advantage of the coexistence of two chemical cues-ROS and low pH-present in the tumor microenvironment. The hydrophobic blocks contain masked pH-responsive side chains, which are exposed exclusively in response to ROS. Hence, the hydrophobic polymer side chains will undergo a charge shift in a very relevant pH window present in the extracellular milieu in most solid tumors (pH 5.6-7.2) after demasking by ROS. Doxorubicin (DOX)-loaded nanosized "AND gate" MR polymersomes (MRPs) are fabricated via microfluidic self-assembly. Chemical characterization reveals ROS-dependent pH sensitivity and accelerated DOX release under influence of both ROS and low pH. Treatment of tumor-bearing mice with DOX-loaded nonresponsive and "AND gate" MRPs dramatically decreases cardiac toxicity. The most optimal "AND gate" MRPs outperform free DOX in terms of tumor growth inhibition and survival, shedding light on chemical requirements for successful cancer nanomedicine.

pH-responsive polymersome-mediated delivery of doxorubicin into tumor sites enhances the therapeutic efficacy and reduces cardiotoxic effects.

The delivery of therapeutics into sites of action by using cargo-delivery platforms potentially minimizes their premature degradation and fast clearance from the bloodstream. Additionally, drug-loaded stimuli-responsive supramolecular assemblies can be produced to respond to the inherent features of tumor microenvironments, such as extracellular acidosis. We report in this framework the use of pH-responsive polymersomes (PSs) manufactured using poly([N-(2-hydroxypropyl)] methacrylamide)35-b-poly[2-(diisopropylamino)ethyl methacrylate]75 as the building unit (PHPMA35-b-PDPA75). The self-assemblies were produced with desired size towards long circulation time and tumor accumulation (hydrodynamic diameter - DH ~ 100 nm), and they could be successfully loaded with 10% w/w DOX (doxorubicin), while maintaining colloidal stability. The DOX loaded amount is presumably mainly burst-released at the acidic microenvironment of tumors thanks to the pH-switchable property of PDPA (pKa ~ 6.8), while reduced drug leakage has been monitored in pH 7.4. Compared to the administration of free DOX, the drug-loaded supramolecular structures greatly enhanced the therapeutic efficacy with effective growth inhibition of EL4 lymphoma tumor model and 100% survival rate in female C57BL/6 black mice over 40 days. The approach also led to reduced cardiotoxic effect. These features highlight the potential application of such nanotechnology-based treatment in a variety of cancer therapies where low local pH is commonly found, and emphasize PHPMA-based nanomedicines as an alternative to PEGylated formulations.

Chelating Polymers for Hereditary Hemochromatosis Treatment.

Hemochromatosis (iron overload) encompasses a group of diseases that are characterized by a toxic hyperaccumulation of iron in parenchymal organs. Currently, only few treatments for this disease have been approved; however, all these treatments possess severe side effects. In this study, a paradigm for hemochromatosis maintenance/preventive therapy is investigated: polymers with negligible systemic biological availability form stable complexes with iron ions in the gastrointestinal tract, which reduces the biological availability of iron. Macroporous polymer beads are synthesized with three different iron-chelating moieties (benzene-1,2-diol, benzene-1,2,3-triol, and 1,10-phenanthroline). The polymers rapidly chelate iron ions from aqueous solutions in vitro in the course of minutes, and are noncytotoxic and nonprooxidant. Moreover, the in vivo biodistribution and pharmacokinetics show a negligible uptake from the gastrointestinal tract (using 125 I-labeled polymer and single photon emission computed tomography/computed tomography), which generally prevents them from having systemic side effects. The therapeutic efficacy of the prepared polymers is successfully tested in vivo, and exhibits a significant inhibition of iron uptake from the gastrointestinal tract without any noticeable signs of toxicity. Furthermore, an in silico method is developed for the prediction of chelator selectivity. Therefore, this paradigm can be applied to the next-generation maintenance/preventive treatment for hemochromatosis and/or other diseases of similar pathophysiology.

Altered Erythro-Myeloid Progenitor Cells Are Highly Expanded in Intensively Regenerating Hematopoiesis.

Regeneration of severely damaged adult tissues is currently only partially understood. Hematopoietic tissue provides a unique opportunity to study tissue regeneration due to its well established steady-state structure and function, easy accessibility, well established research methods, and the well-defined embryonic, fetal, and adult stages of development. Embryonic/fetal liver hematopoiesis and adult hematopoiesis recovering from damage share the need to expand populations of progenitors and stem cells in parallel with increasing production of mature blood cells. In the present study, we analyzed adult hematopoiesis in mice subjected to a submyeloablative dose (6 Gy) of gamma radiation and targeted the period of regeneration characterized by massive production of mature blood cells along with ongoing expansion of immature hematopoietic cells. We uncovered significantly expanded populations of developmentally advanced erythroid and myeloid progenitors with significantly altered immunophenotype. Their population expansion does not require erythropoietin stimulation but requires the SCF/c-Kit receptor signaling. Regenerating hematopoiesis significantly differs from the expanding hematopoiesis in the fetal liver but we find some similarities between the regenerating hematopoiesis and the early embryonic definitive hematopoiesis. These are in (1) the concomitant population expansion of myeloid progenitors and increasing production of myeloid blood cells (2) performing these tasks despite the severely reduced transplantation capacity of the hematopoietic tissues, and (3) the expression of CD16/32 in most progenitors. Our data thus provide a novel insight into tissue regeneration by suggesting that cells other than stem cells and multipotent progenitors can be of fundamental importance for the rapid recovery of tissue function.

Reactive Oxygen Species (ROS)-Responsive Polymersomes with Site-Specific Chemotherapeutic Delivery into Tumors via Spacer Design Chemistry.

The lack of cellular and tissue specificities in conventional chemotherapies along with the generation of a complex tumor microenvironment (TME) limits the dosage of active agents that reaches tumor sites, thereby resulting in ineffective responses and side effects. Therefore, the development of selective TME-responsive nanomedicines is of due relevance toward successful chemotherapies, albeit challenging. In this framework, we have synthesized novel, ready-to-use ROS-responsive amphiphilic block copolymers (BCs) with two different spacer chemistry designs to connect a hydrophobic boronic ester-based ROS sensor to the polymer backbone. Hydrodynamic flow focusing nanoprecipitation microfluidics (MF) was used in the preparation of well-defined ROS-responsive PSs; these were further characterized by a combination of techniques [1H NMR, dynamic light scattering (DLS), static light scattering (SLS), transmission electron microscopy (TEM), and cryogenic TEM (cryo-TEM)]. The reaction with hydrogen peroxide releases an amphiphilic phenol or a hydrophilic carboxylic acid, which affects polymersome (PS) stability and cargo release. Therefore, the importance of the spacer chemistry in BC deprotection and PS stability and cargo release is herein highlighted. We have also evaluated the impact of spacer chemistry on the PS-specific release of the chemotherapeutic drug doxorubicin (DOX) into tumors in vitro and in vivo. We demonstrate that by spacer chemistry design one can enhance the efficacy of DOX treatments (decrease in tumor growth and prolonged animal survival) in mice bearing EL4 T cell lymphoma. Side effects (weight loss and cardiotoxicity) were also reduced compared to free DOX administration, highlighting the potential of the well-defined ROS-responsive PSs as TME-selective nanomedicines. The PSs could also find applications in other environments with high ROS levels, such as chronic inflammations, aging, diabetes, cardiovascular diseases, and obesity.

Synthesis and modification of uniform PEG-neridronate-modified magnetic nanoparticles determines prolonged blood circulation and biodistribution in a mouse preclinical model.

Magnetite (Fe3O4) nanoparticles with uniform sizes of 10, 20, and 31 nm were prepared by thermal decomposition of Fe(III) oleate or mandelate in a high-boiling point solvent (>320 °C). To render the particles with hydrophilic and antifouling properties, their surface was coated with a PEG-containing bisphosphonate anchoring group. The PEGylated particles were characterized by a range of physicochemical methods, including dynamic light scattering, transmission electron microscopy, thermogravimetric analysis, Fourier transform infrared spectroscopy, and magnetization measurements. As the particle size increased from 10 to 31 nm, the amount of PEG coating decreased from 28.5 to 9 wt.%. The PEG formed a dense brush-like shell on the particle surface, which prevented particles from aggregating in water and PBS (pH 7.4) and maximized the circulation time in vivo. Magnetic resonance relaxometry confirmed that the PEG-modified Fe3O4 nanoparticles had high relaxivity, which increased with increasing particle size. In the in vivo experiments in a mouse model, the particles provided visible contrast enhancement in the magnetic resonance images. Almost 70% of administrated 20-nm magnetic nanoparticles still circulated in the blood stream after four hours; however, their retention in the tumor was rather low, which was likely due to the antifouling properties of PEG.

Thermoresponsive ß-glucan-based polymers for bimodal immunoradiotherapy - Are they able to promote the immune system?

A conceptually new bimodal immunoradiotherapy treatment was demonstrated using thermoresponsive polymer ß-glucan-graft-poly(2-isopropyl-2-oxazoline-co-2-butyl-2-oxazoline) bearing complexes of 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid with yttrium-90(III) at the graft ends. The behavior of this thermoresponsive polymer in aqueous solutions was studied, and it showed the appropriate cloud point temperature for brachytherapy applications. The polymer was tested in vitro, and it exhibited nontoxicity and active uptake into cancer cells and macrophages with colocalization in the lysosomes and macrophagosomes. Moreover, the observed oxidative burst response of the leukocytes established the immunostimulatory properties of the polymer, which were also studied in vivo after injection into the thigh muscles of healthy mice. The subsequent histological evaluation revealed the extensive immune activation reactions at the site of injection. Furthermore, the production of tumor necrosis factor α induced by the prepared polymer was observed in vitro, denoting the optimistic prognosis of the treatment. The biodistribution study in vivo indicated the formation of the polymer depot, which was gradually degraded and excluded from the body. The radiolabeled polymer was used during in vivo antitumor efficiency experiments on mice with EL4 lymphoma. The immunoradiotherapy group (treated with the radiolabeled polymer) demonstrated the complete inhibition of tumor growth during the beginning of the treatment. Moreover, 7 of the 15 mice were completely cured in this group, while the others exhibited significantly prolonged survival time compared to the control group. The in vivo experiments indicated the considerable synergistic effect of using immunoradiotherapy compared to separately using immunotherapy or radiotherapy.