Hsp70-Targeting and Size-Tunable Nanoparticles Combine with PD-1 Checkpoint Blockade to Treat Glioma.

Invasive glioma usually disrupts the integrity of the blood-brain barrier (BBB), making the delivery of nanodrugs across the BBB possible, but sufficient targeting ability is still avidly needed to improve drug accumulation in glioma. Membrane-bound heat shock protein 70 (Hsp70) is expressed on the membrane of glioma cells rather than adjacent normal cells, therefore it can serve as a specific glioma target. Meanwhile, prolonging the retention in tumors is important for active-targeting nanoparticles to overcome receptor-binding barriers. Herein, the Hsp70-targeting and acid-triggered self-assembled gold nanoparticles (D-A-DA/TPP) are proposed to realize selective delivery of doxorubicin (DOX) to glioma. In the weakly acidic glioma matrix, D-A-DA/TPP formed aggregates to prolong retention, improve receptor-binding efficiency and facilitate acid-responsive DOX release. DOX accumulation in glioma induced immunogenic cell death (ICD) to promote antigen presentation. Meanwhile, combination with the PD-1 checkpoint blockade further activate T cells and provokes robust anti-tumor immunity. The results showed that D-A-DA/TPP can induce more glioma apoptosis. Furthermore, in vivo studies indicated D-A-DA/TPP plus PD-1 checkpoint blockade significantly improved median survival time. This study offeres a potential nanocarrier combining size-tunable strategy with active targeting ability to increase drug enrichment in glioma and synergizes with PD-1 checkpoint blockade to achieve chemo-immunotherapy.

Hepatitis B virus envelope proteins can serve as therapeutic targets embedded in the host cell plasma membrane.

Hepatitis B virus (HBV) infection is a major health threat causing 880,000 deaths each year. Available therapies control viral replication but do not cure HBV, leaving patients at risk to develop hepatocellular carcinoma. Here, we show that HBV envelope proteins (HBs)-besides their integration into endosomal membranes-become embedded in the plasma membrane where they can be targeted by redirected T-cells. HBs was detected on the surface of HBV-infected cells, in livers of mice replicating HBV and in HBV-induced hepatocellular carcinoma. Staining with HBs-specific recombinant antibody MoMab recognising a conformational epitope indicated that membrane-associated HBs remains correctly folded in HBV-replicating cells in cell culture and in livers of HBV-transgenic mice in vivo. MoMab coated onto superparamagnetic iron oxide nanoparticles allowed to detect membrane-associated HBs after HBV infection by electron microscopy in distinct stretches of the hepatocyte plasma membrane. Last but not least, we demonstrate that HBs located on the cell surface allow therapeutic targeting of HBV-positive cells by T-cells either engrafted with a chimeric antigen receptor or redirected by bispecific, T-cell engager antibodies. TAKE AWAYS: HBs become translocated to the plasma membrane. Novel, recombinant antibody confirmed proper conformation of HBs on the membrane. HBs provide an interesting target by T-cell-based, potentially curative therapies.

Recent Advances in Copper-Based Organic Complexes and Nanoparticles for Tumor Theranostics.

Treatment of drug-resistant forms of cancer requires consideration of their hallmark features, such as abnormal cell death mechanisms or mutations in drug-responding molecular pathways. Malignant cells differ from their normal counterparts in numerous aspects, including copper metabolism. Intracellular copper levels are elevated in various cancer types, and this phenomenon could be employed for the development of novel oncotherapeutic approaches. Copper maintains the cell oxidation levels, regulates the protein activity and metabolism, and is involved in inflammation. Various copper-based compounds, such as nanoparticles or metal-based organic complexes, show specific activity against cancer cells according to preclinical studies. Herein, we summarize the major principles of copper metabolism in cancer cells and its potential in cancer theranostics.

Potential of Magnetic Hyperthermia to Stimulate Localized Immune Activation.

Magnetic hyperthermia (MH) harnesses the heat-releasing properties of superparamagnetic iron oxide nanoparticles (SPIONs) and has potential to stimulate immune activation in the tumor microenvironment whilst sparing surrounding normal tissues. To assess feasibility of localized MH in vivo, SPIONs are injected intratumorally and their fate tracked by Zirconium-89-positron emission tomography, histological analysis, and electron microscopy. Experiments show that an average of 49% (21-87%, n = 9) of SPIONs are retained within the tumor or immediately surrounding tissue. In situ heating is subsequently generated by exposure to an externally applied alternating magnetic field and monitored by thermal imaging. Tissue response to hyperthermia, measured by immunohistochemical image analysis, reveals specific and localized heat-shock protein expression following treatment. Tumor growth inhibition is also observed. To evaluate the potential effects of MH on the immune landscape, flow cytometry is used to characterize immune cells from excised tumors and draining lymph nodes. Results show an influx of activated cytotoxic T cells, alongside an increase in proliferating regulatory T cells, following treatment. Complementary changes are found in draining lymph nodes. In conclusion, results indicate that biologically reactive MH is achievable in vivo and can generate localized changes consistent with an anti-tumor immune response.

Pleiotropic roles of cold shock proteins with special emphasis on unexplored cold shock protein member of Plasmodium falciparum.

The cold shock domain (CSD) forms the hallmark of the cold shock protein family that provides the characteristic feature of binding with nucleic acids. While much of the information is available on bacterial, plants and human cold shock proteins, their existence and functions in the malaria parasite remains undefined. In the present review, the available information on functions of well-characterized cold shock protein members in different organisms has been collected and an attempt was made to identify the presence and role of cold shock proteins in malaria parasite. A single Plasmodium falciparum cold shock protein (PfCoSP) was found in P. falciparum which is reported to be essential for parasite survival. Essentiality of PfCoSP underscores its importance in malaria parasite life cycle. In silico tools were used to predict the features of PfCoSP and to identify its homologues in bacteria, plants, humans, and other Plasmodium species. Modelled structures of PfCoSP and its homologues in Plasmodium species were compared with human cold shock protein 'YBOX-1' (Y-box binding protein 1) that provide important insights into their functioning. PfCoSP model was subjected to docking with B-form DNA and RNA to reveal a number of residues crucial for their interaction. Transcriptome analysis and motifs identified in PfCoSP implicate its role in controlling gene expression at gametocyte, ookinete and asexual blood stages of malaria parasite. Overall, this review emphasizes the functional diversity of the cold shock protein family by discussing their known roles in gene expression regulation, cold acclimation, developmental processes like flowering transition, and flower and seed development, and probable function in gametocytogenesis in case of malaria parasite. This enables readers to view the cold shock protein family comprehensively.

Granzyme B Functionalized Nanoparticles Targeting Membrane Hsp70-Positive Tumors for Multimodal Cancer Theranostics.

Functionalized superparamagnetic iron oxide nanoparticles (SPIONs) have emerged as potential clinical tools for cancer theranostics. Membrane-bound 70 kDa heat shock protein (mHsp70) is ubiquitously expressed on the cell membrane of various tumor types but not normal cells and therefore provides a tumor-specific target. The serine protease granzyme B (GrB) that is produced as an effector molecule by activated T and NK cells has been shown to specifically target mHsp70 on tumor cells. Following binding to Hsp70, GrB is rapidly internalized into tumor cells. Herein, it is demonstrated that GrB functionalized SPIONs act as a contrast enhancement agent for magnetic resonance imaging and induce specific tumor cell apoptosis. Combinatorial regimens employing stereotactic radiotherapy and/or magnetic targeting are found to further enhance the therapeutic efficacy of GrB-SPIONs in different tumor mouse models.

Combination of Anti-Cancer Drugs with Molecular Chaperone Inhibitors.

Most molecular chaperones belonging to heat shock protein (HSP) families are known to protect cancer cells from pathologic, environmental and pharmacological stress factors and thereby can hamper anti-cancer therapies. In this review, we present data on inhibitors of the heat shock response (particularly mediated by the chaperones HSP90, HSP70, and HSP27) either as a single treatment or in combination with currently available anti-cancer therapeutic approaches. An overview of the current literature reveals that the co-administration of chaperone inhibitors and targeting drugs results in proteotoxic stress and violates the tumor cell physiology. An optimal drug combination should simultaneously target cytoprotective mechanisms and trigger the imbalance of the tumor cell physiology.

Heat shock protein 70 and tumor-infiltrating NK cells as prognostic indicators for patients with squamous cell carcinoma of the head and neck after radiochemotherapy: A multicentre retrospective study of the German Cancer Consortium Radiation Oncology Group (DKTK-ROG).

Tumor cells frequently overexpress heat shock protein 70 (Hsp70) and present it on their cell surface, where it can be recognized by pre-activated NK cells. In our retrospective study the expression of Hsp70 was determined in relation to tumor-infiltrating CD56+ NK cells in formalin-fixed paraffin embedded (FFPE) tumor specimens of patients with SCCHN (N = 145) as potential indicators for survival and disease recurrence. All patients received radical surgery and postoperative cisplatin-based radiochemotherapy (RCT). In general, Hsp70 expression was stronger, but with variable intensities, in tumor compared to normal tissues. Patients with high Hsp70 expressing tumors (scores 3-4) showed significantly decreased overall survival (OS; p = 0.008), local progression-free survival (LPFS; p = 0.034) and distant metastases-free survival (DMFS; p = 0.044), compared to those with low Hsp70 expression (scores 0-2), which remained significant after adjustment for relevant prognostic variables. The adverse prognostic value of a high Hsp70 expression for OS was also observed in patient cohorts with p16- (p = 0.001), p53- (p = 0.0003) and HPV16 DNA-negative (p = 0.001) tumors. The absence or low numbers of tumor-infiltrating CD56+ NK cells also correlated with significantly decreased OS (p = 0.0001), LPFS (p = 0.0009) and DMFS (p = 0.0001). A high Hsp70 expression and low numbers of tumor-infiltrating NK cells have the highest negative predictive value (p = 0.00004). In summary, a strong Hsp70 expression and low numbers of tumor-infiltrating NK cells correlate with unfavorable outcome following surgery and RCT in patients with SCCHN, and thus serve as negative prognostic markers.

Phloretin increases the anti-tumor efficacy of intratumorally delivered heat-shock protein 70 kDa (HSP70) in a murine model of melanoma.

Recombinant HSP70 chaperone exerts a profound anticancer effect when administered intratumorally. This action is based on the ability of HSP70 to penetrate tumor cells and extract its endogenous homolog. To enhance the efficacy of HSP70 cycling, we employed phloretin, a flavonoid that enhances the pore-forming activity of the chaperone on artificial membranes. Phloretin increased the efficacy of HSP70 penetration in B16 mouse melanoma cells and K-562 human erythroblasts; this was accompanied with increased transport of the endogenous HSP70 to the plasma membrane. Importantly, treatment with HSP70 combined with phloretin led to the elevation of cell sensitivity to cytotoxic lymphocytes by 16-18 % compared to treatment with the chaperone alone. The incubation of K-562 cells with biotinylated HSP70 and phloretin increased the amount of the chaperone released from cells, suggesting that chaperone cycling could trigger a specific anti-tumor response. We studied the effect of the combination of HSP70 and phloretin using B16 melanoma and a novel method of HSP70-gel application. We found that the addition of phloretin to the gel reduced tumor weight almost fivefold compared with untreated mice, while the life span of the animals extended from 25 to 39 days. The increased survival was corroborated by the activation of innate and adaptive immunity; interestingly, HSP70 was more active in induction of CD8+ cell-mediated toxicity and γIFN production while phloretin contributed largely to the CD56+ cell response. In conclusion, the combination of HSP70 with phloretin could be a novel treatment for efficient immunotherapy of intractable cancers such as skin melanoma.

Detection of experimental myocardium infarction in rats by MRI using heat shock protein 70 conjugated superparamagnetic iron oxide nanoparticle.

Superparamagnetic iron-oxide based contrast agents can provide important diagnostic information regarding the assessment of cardiac inflammatory diseases. The aim of the study was to analyze whether nanoparticles conjugated to recombinant 70-kDa heat shock protein (Hsp70-SPION) can be applied for the detection of acute myocardium infarct by MRI. Cellular experiments demonstrated increased CD40-mediated uptake of Hsp70-SPIONs in comparison to non-conjugated SPIONs. Following induction of an acute infarct in rats by ligation of the left anterior descending artery SPIONs and Hsp70-SPION conjugates were injected intravenously on day 4. The animals underwent sequential MRI that showed the presence of the particles in the infarcted zone. Subsequent biodistribution analyses with the help of method on non-linear magnetic response indicated the preferential accumulation of the Hsp70-SPIONs in the heart tissue that was further confirmed with histological analyses. The study demonstrated that an acute infarct can be visualized by MRI using Hsp70-functionalized SPION conjugates. FROM THE CLINICAL EDITOR: Superparamagnetic iron oxides nanoparticles (SPIONs) have been studied extensively as a contrast agent for MRI. Their tissue specificity can be further enhanced by conjugation with various ligands. In this study, the authors conjugated superparamagnetic nanoparticles to 70-kDa heat shock protein (Hsp70-SPION) to investigate the feasibility for the detection of acute myocardium infarct. The positive findings would suggest that this approach might be used clinically in the future.

Changes of fractional anisotropy (FA) and apparent diffusion coefficient (ADC) in the model of experimental acute hydrocephalus in rabbits.

BACKGROUND: To study the integrity of white matter, we investigated the correlation between the changes in neuroradiological and morphological parameters in an animal model of acute obstructive hydrocephalus. METHODS: Hydrocephalus was induced in New Zealand rabbits (n = 10) by stereotactic injection of kaolin into the lateral ventricles. Control animals received saline in place of kaolin (n = 10). The progression of hydrocephalus was assessed using magnetic resonance imaging. Regional fractional anisotropy (FA) and the apparent diffusion coefficient (ADC) were measured in several white matter regions before and after the infusion of kaolin. Morphology of myelinated nerve fibers as well as of the blood-brain barrier were studied with the help of transmission electron microscopy (TEM) and light microscopy. RESULTS: Compared with control animals, kaolin injection into the ventricles resulted in a dramatic increase in ventricular volume with compression of basal cisterns, brain shift and periventricular edema (as observed on magnetic resonance imaging [MRI]). The values of ADC in the periventricular and periaqueductal areas significantly increased in the experimental group (P < 0.05). FA decreased by a factor of 2 in the zones of periventricular, periaqueductal white matter and corpus collosum. Histological analysis demonstrated the impairment of the white matter and necrobiotic changes in the cortex. Microsctructural alterations of the myelin fibers were further proved with the help of TEM. Blood-brain barrier ultrastructure assessment showed the loss of its integrity. CONCLUSIONS: The study demonstrated the correlation of the neuroradiological parameters with morphological changes. The abnormality of the FA and ADC parameters in the obstructive hydrocephalus represents a significant implication for the diagnostics and management of hydrocephalus in patients.

Effective immunotherapy of rat glioblastoma with prolonged intratumoral delivery of exogenous heat shock protein Hsp70.

Chaperone Hsp70 can activate adaptive immunity suggesting its possible application as an antitumor vaccine. To assess the therapeutic capacity of Hsp70 we administered purified chaperone into a C6 glioblastoma brain tumor and explored the viability and tumor size as well as interferon gamma (IFNγ) production and cytotoxicity of lymphocytes in the treated animals. Targeted intratumoral injection of Hsp70 resulted in its distribution within the area of glioblastoma, and caused significant inhibition of tumor progression as confirmed by magnetic resonance imaging. The delay in tumor growth corresponded to the prolonged survival of tumor-bearing animals of up to 31 days versus 20 days in control. Continuous administration of Hsp70 with an osmotic pump increased survival even further (39 days). Therapeutic efficacy was associated with infiltration to glioblastoma of NK cells (Ly-6c+) and T lymphocytes (CD3+, CD4+ and CD8+) as well as with an increase in the activity of NK cells (granzyme B production) and CD8+ T lymphocytes as shown by IFNγ ELISPOT assay. Furthermore, we found that Hsp70 treatment caused concomitantly, with a tenfold elevated IFNγ production, an increase in anti-C6 tumor cytotoxicity of lymphocytes. In conclusion, continuous intratumoral delivery of Hsp70 demonstrates high therapeutic potential and therefore could be applied in the treatment of glioblastoma.

Tumor Dormancy and Reactivation: The Role of Heat Shock Proteins.

Tumors are a heterogeneous group of cell masses originating in various organs or tissues. The cellular composition of the tumor cell mass interacts in an intricate manner, influenced by humoral, genetic, molecular, and tumor microenvironment cues that dictate tumor growth or suppression. As a result, tumors undergo a period of a dormant state before their clinically discernible stage, which surpasses the clinical dormancy threshold. Moreover, as a genetically imprinted strategy, early-seeder cells, a distinct population of tumor cells, break off to dock nearby or extravasate into blood vessels to secondary tissues, where they form disseminated solitary dormant tumor cells with reversible capacity. Among the various mechanisms underlying the dormant tumor mass and dormant tumor cell formation, heat shock proteins (HSPs) might play one of the most important roles in how the dormancy program plays out. It is known that numerous aberrant cellular processes, such as malignant transformation, cancer cell stemness, tumor invasion, metastasis, angiogenesis, and signaling pathway maintenance, are influenced by the HSPs. An accumulating body of knowledge suggests that HSPs may be involved in the angiogenic switch, immune editing, and extracellular matrix (ECM) remodeling cascades, crucial genetically imprinted strategies important to the tumor dormancy initiation and dormancy maintenance program. In this review, we highlight the biological events that orchestrate the dormancy state and the body of work that has been conducted on the dynamics of HSPs in a tumor mass, as well as tumor cell dormancy and reactivation. Additionally, we propose a conceptual framework that could possibly underlie dormant tumor reactivation in metastatic relapse.

Tumor Microenvironment Modulation by Cancer-Derived Extracellular Vesicles.

The tumor microenvironment (TME) plays an important role in the process of tumorigenesis, regulating the growth, metabolism, proliferation, and invasion of cancer cells, as well as contributing to tumor resistance to the conventional chemoradiotherapies. Several types of cells with relatively stable phenotypes have been identified within the TME, including cancer-associated fibroblasts (CAFs), tumor-associated macrophages (TAMs), neutrophils, and natural killer (NK) cells, which have been shown to modulate cancer cell proliferation, metastasis, and interaction with the immune system, thus promoting tumor heterogeneity. Growing evidence suggests that tumor-cell-derived extracellular vesicles (EVs), via the transfer of various molecules (e.g., RNA, proteins, peptides, and lipids), play a pivotal role in the transformation of normal cells in the TME into their tumor-associated protumorigenic counterparts. This review article focuses on the functions of EVs in the modulation of the TME with a view to how exosomes contribute to the transformation of normal cells, as well as their importance for cancer diagnosis and therapy.

Comparison In Vitro Study on the Interface between Skin and Bone Cell Cultures and Microporous Titanium Samples Manufactured with 3D Printing Technology Versus Sintered Samples.

Percutaneous implants osseointegrated into the residuum of a person with limb amputation need to provide mechanical stability and protection against infections. Although significant progress has been made in the biointegration of percutaneous implants, the problem of forming a reliable natural barrier at the level of the surface of the implant and the skin and bone tissues remains unresolved. The use of a microporous implant structure incorporated into the Skin and Bone Integrated Pylon (SBIP) should address the issue by allowing soft and bone tissues to grow directly into the implant structure itself, which, in turn, should form a reliable barrier to infections and support strong osseointegration. To evaluate biological interactions between dermal fibroblasts and MC3T3-E1 osteoblasts in vitro, small titanium discs (with varying pore sizes and volume fractions to achieve deep porosity) were fabricated via 3D printing and sintering. The cell viability MTT assay demonstrated low cytotoxicity for cells co-cultured in the pores of the 3D-printed and sintered Ti samples during the 14-day follow-up period. A subsequent Quantitative Real-Time Polymerase Chain Reaction (RT-PCR) analysis of the relative gene expression of biomarkers that are associated with cell adhesion (α2, α5, αV, and ß1 integrins) and extracellular matrix components (fibronectin, vitronectin, type I collagen) demonstrated that micropore sizes ranging from 200 to 500 µm of the 3D printed and sintered Ti discs were favorable for dermal fibroblast adhesion. For example, for representative 3D-printed Ti sample S6 at 72 h the values were 4.71 ± 0.08 (α2 integrin), 4.96 ± 0.08 (α5 integrin), 4.71 ± 0.08 (αV integrin), and 1.87 ± 0.12 (ß1 integrin). In contrast, Ti discs with pore sizes ranging from 400 to 800 µm demonstrated the best results (in terms of marker expression related to osteogenic differentiation, including osteopontin, osteonectin, osteocalcin, TGF-ß1, and SMAD4) for MC3T3-E1 cells. For example, for the representative 3D sample S4 on day 14, the marker levels were 11.19 ± 0.77 (osteopontin), 7.15 ± 0.29 (osteonectin), and 6.08 ± 0.12 (osteocalcin), while for sintered samples the levels of markers constituted 5.85 ± 0.4 (osteopontin), 4.45 ± 0.36 (osteonectin), and 4.46 ± 0.3 (osteocalcin). In conclusion, the data obtained show the high biointegrative properties of porous titanium structures, while the ability to implement several pore options in one structure using 3D printing makes it possible to create personalized implants for the best one-time integration with both skin and bone tissues.

Targeting host inducible-heat shock protein 70 with PES-Cl is a promising antiviral strategy against SARS-CoV-2 infection and pathogenesis.

One of the fundamental mechanisms developed by the host to contain the highly infectious and rapidly proliferating SARS-coronavirus is elevation of body temperature, a natural fallout of which is heat shock proteins over-expression. Here, for the first time, we demonstrate that the SARS-CoV-2 exploits the host Heat shock protein 70 (Hsp70) chaperone for its entry and propagation, and blocking it can combat the infection. SARS-CoV-2 infection as well as febrile temperature enhanced Hsp70 expression in host Vero E6 cells. Furthermore, heat shock or viral infection elevated the host cell autophagic response which is a prerequisite for viral propagation. In addition, Hsp70 protein demonstrated strong interaction with host Angiotensin-converting enzyme 2 (ACE2) as well as the receptor binding domain (RBD) of the SARS-CoV-2 Spike protein, indicating that interaction of Hsp70 with ACE2 and Spike protein may serve to protect them during febrile conditions. Suppressive and prophylactic treatment of Vero E6 cells with Hsp70 inhibitor PES, 2-(3-chlorophenyl) ethynesulfonamide (PES-Cl), abrogated viral infection more potently than the currently used drug Remdesivir. In conclusion, our study not only provides a fundamental insight into the role of host Hsp70 in SARS-CoV-2 pathogenesis, it paves the way for development of potent and irresistible anti-viral therapeutics.

Targeting PfProhibitin 2-Hu-Hsp70A1A complex as a unique approach towards malaria vaccine development.

Malaria parasite invasion to host erythrocytes is mediated by multiple interactions between merozoite ligands and erythrocyte receptors that contribute toward the development of disease pathology. Here, we report a novel antigen Plasmodium prohibitin "PfPHB2" and identify its cognate partner "Hsp70A1A" in host erythrocyte that plays a crucial role in mediating host-parasite interaction during merozoite invasion. Using small interfering RNA (siRNA)- and glucosamine-6-phosphate riboswitch (glmS) ribozyme-mediated approach, we show that loss of Hsp70A1A in red blood cells (RBCs) or PfPHB2 in infected red blood cells (iRBCs), respectively, inhibit PfPHB2-Hsp70A1A interaction leading to invasion inhibition. Antibodies targeting PfPHB2 and monoclonal antibody therapeutics against Hsp70A1A efficiently block parasite invasion. Recombinant PfPHB2 binds to RBCs which is inhibited by anti-PfPHB2 antibody and monoclonal antibody against Hsp70A1A. The validation of PfPHB2 to serve as antigen is further supported by detection of anti-PfPHB2 antibody in patient sera. Overall, this study proposes PfPHB2 as vaccine candidate and highlights the use of monoclonal antibody therapeutics for future malaria treatment.

Mesenchymal stem cells-derived extracellular vesicles for therapeutics of renal tuberculosis.

Extrapulmonary tuberculosis with a renal involvement can be a manifestation of a disseminated infection that requires therapeutic intervention, particularly with a decrease in efficacy of conventional regimens. In the present study, we investigated the therapeutic potency of mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) in the complex anti-tuberculosis treatment (ATT). A rabbit model of renal tuberculosis (rTB) was constructed by injecting of the standard strain Mycobacterium tuberculosis H37Rv into the cortical layer of the kidney parenchyma. Isolated rabbit MSC-EVs were intravenously administered once as an addition to standard ATT (isoniazid, pyrazinamide, and ethambutol). The therapeutic efficacy was assessed by analyzing changes of blood biochemical biomarkers and levels of anti- and pro-inflammatory cytokines as well as by renal computed tomography with subsequent histological and morphometric examination. The therapeutic effect of therapy with MSC-EVs was shown by ELISA method that confirmed a statistically significant increase of the anti-inflammatory and decrease of pro-inflammatory cytokines as compared to conventional treatment. In addition, there is a positive trend in increase of ALP level, animal weigh, and normalization of ADA activity that can indicate an improvement of kidney state. A significant reduction of the area of specific and interstitial inflammation indicated positive affect of MSC-EVs that suggests a shorter duration of ATT. The number of MSC-EVs proteins (as identified by mass-spectometry analysis) with anti-microbial, anti-inflammatory and immunoregulatory functions reduced the level of the inflammatory response and the severity of kidney damage (further proved by morphometric analysis). In conclusion, MSC-EVs can be a promising tool for the complex treatment of various infectious diseases, in particularly rTB.

Membrane-bound Heat Shock Protein mHsp70 Is Required for Migration and Invasion of Brain Tumors.

Molecular chaperones, especially 70 kDa heat shock protein, in addition to their intracellular localization in cancer cells, can be exposed on the surface of the plasma membrane. We report that the membrane-associated chaperone mHsp70 of malignant brain tumors is required for high migratory and invasive activity of cancer cells. Live-cell inverted confocal microscopy of tumor samples from adult (n = 23) and pediatric (n = 9) neurooncologic patients showed pronounced protein expression on the membrane, especially in the perifocal zone. Mass spectrometry analysis of lipid rafts isolated from tumor cells confirmed the presence of the protein in the chaperone cluster (including representatives of other families, such as Hsp70, Hsc70, Hsp105, and Hsp90), which in turn, during interactome analysis, was associated with proteins involved in cell migration (e.g., Rac1, RhoC, and myosin-9). The use of small-molecule inhibitors of HSP70 (PES and JG98) led to a substantial decrease in the invasive potential of cells isolated from a tumor sample of patients, which indicates the role of the chaperone in invasion. Moreover, the use of HSP70 inhibitors in animal models of orthotopic brain tumors significantly delayed tumor progression, which was accompanied by an increase in overall survival. Data demonstrate that chaperone inhibitors, particularly JG98, disrupt the function of mHsp70, thereby providing an opportunity to better understand the diverse functions of this protein and offer aid in the development of novel cancer therapies. SIGNIFICANCE: Membrane-bound mHsp70 is required for brain tumor cell migration and invasion and therefore could be employed as a target for anticancer therapies.

Intracellular and extracellular Hsp70 chaperone as a target for cancer therapy.

PURPOSE: Heat shock protein 70 (HSPA family) is a multi-functional protein which protects individual cells from proteotoxic shock and the whole organism from microbial, viral and oncogenic pathogens. These diverse functions may depend upon 'chaperone' activity that allows Hsp70 to regulate the mechanism of damaged protein recovery or utilisation inside a cell and to be a potent adjuvant, stimulating immune activity against a variety of viral or tumour antigens. The aim of this review is to present recent data on specific roles of intracellular and extracellular Hsp70 in cancerous tissue. CONCLUSION: The data presented in this paper show that endogenous Hsp70 protects cancer cells of different origins from a variety of cytotoxic threats including cancer cell therapeutics. In contrast, however, Hsp70 released from stressed cancer cells can serve as a danger signal or may recruit cells responsible for the generation of innate and adaptive immune responses against tumour cells.