Carbonic Anhydrase Inhibitors Induce Ferroptosis through Inhibition of AKT/FTH1 Signaling in Ewing Sarcoma Tumor Cells.

Ewing sarcoma (ES) is one of the most frequent types of malignant tumors among children. The active metabolic state of ES cells presents a new potential target for therapeutic interventions. As a primary regulator of cellular homeostasis, carbonic anhydrases (CAs; EC 4.2.1.1) have emerged as promising molecular targets for the development of anticancer drugs. Within the present study, we tested the commercial drug acetazolamide and our previously discovered inhibitors to target the CAII isoform, which was overexpressed and positively correlated with ES patient relapse. We employed molecular biology tests to identify effective inhibitors of CAII that can induce ferroptosis by downregulating FTH1 expression in ES cells. In vitro, we have also demonstrated their ability to reduce cell proliferation, decrease invasion, and induce apoptosis- or autophagy-related cell death. Using Western blotting, we confirmed the induction of cathepsin B in cells treated with CA inhibitors. It was found that the suppression of cathepsin B expression during the treatment reduces the anticancer efficacy of selected CAII inhibitors. These experiments highlighted profound antitumor activity of CAII inhibitors attributive to their remarkable ability to trigger ferroptosis in Ewing sarcoma cells without causing substantial host damage. The obtained results suggest that cytosolic CAII may be a prospective target for ES treatment, and CAII inhibitors can be considered as potential single-agent or combination antitumor agents to be used in the treatment of ES.

Development and characterization of chimera of yellow fever virus vaccine strain and Tick-Borne encephalitis virus.

Tick-borne encephalitis virus (TBEV) is one of the most threatening pathogens which affects the human central nervous system (CNS). TBEV circulates widely in Northern Eurasia. According to ECDC, the number of TBE cases increase annually. There is no specific treatment for the TBEV infection, thus vaccination is the main preventive measure. Despite the existence of several inactivated vaccines currently being licensed, the development of new TBEV vaccines remains a leading priority in countries endemic to this pathogen. Here we report new recombinant virus made by infectious subgenomic amplicon (ISA) approach using TBEV and yellow fever virus vaccine strain (YF17DD-UN) as a genetic backbone. The recombinant virus is capable of effective replication in mammalian cells and induce TBEV-neutralizing antibodies in mice. Unlike the original vector based on the yellow fever vaccine strain, chimeric virus became neuroinvasive in doses of 107-106 PFU and can be used as a model of flavivirus neuroinvasiveness, neurotropism and neurovirulence. These properties of hybrid structures are the main factors limiting their practical use as vaccines platforms.

The spectrum of cell death in sarcoma.

The balance between cell death and cell survival is a highly coordinated process by which cells break down and remove unnecessary or harmful materials in a controlled, highly regulated, and compartmentalized manner. Cell exposure to various stresses, such as oxygen starvation, a lack of nutrients, or exposure to radiation, can initiate autophagy. Autophagy is a carefully orchestrated process with multiple steps, each regulated by specific genes and proteins. Autophagy proteins impact cellular maintenance and cell fate in response to stress, and targeting this process is one of the most promising methods of anti-tumor therapy. It is currently not fully understood how autophagy affects different types of tumor cells, which makes it challenging to predict outcomes when this process is manipulated. In this review, we will explore the mechanisms of autophagy and investigate it as a potential and promising therapeutic target for aggressive sarcomas.

Dynamics of SARS-CoV-2 Major Genetic Lineages in Moscow in the Context of Vaccine Prophylaxis.

Findings collected over two and a half years of the COVID-19 pandemic demonstrated that the level immunity resulting from vaccination and infection is insufficient to stop the circulation of new genetic variants. The short-term decline in morbidity was followed by a steady increase. The early identification of new genetic lineages that will require vaccine adaptation in the future is an important research target. In this study, we summarised data on the variability of genetic line composition throughout the COVID-19 pandemic in Moscow, Russia, and evaluated the virological and epidemiological features of dominant variants in the context of selected vaccine prophylaxes. The prevalence of the Omicron variant highlighted the low effectiveness of the existing immune layer in preventing infection, which points to the necessity of optimising the antigens used in vaccines in Moscow. Logistic growth curves showing the rate at which the new variant displaces the previously dominant variants may serve as early indicators for selecting candidates for updated vaccines, along with estimates of efficacy, reduced viral neutralising activity against the new strains, and viral load in previously vaccinated patients.

Estimation of anti-orthopoxvirus immunity in Moscow residents and potential risks of spreading Monkeypox virus.

WHO has declared the outbreak of monkeypox as a public health emergency of international concern. In less than three months, monkeypox was detected in more than 30 000 people and spread to more than 80 countries around the world. It is believed that the immunity formed to smallpox vaccine can protect from monkeypox infection with high efficiency. The widespread use of Vaccinia virus has not been carried out since the 1980s, which raises the question of the level of residual immunity among the population and the identification of groups requiring priority vaccination. We conducted a cross-sectional serological study of remaining immunity among Moscow residents. To do this, a collection of blood serum samples of age group over 30 years old was formed, an in-house ELISA test system was developed, and a virus neutralization protocol was set up. Serum samples were examined for the presence of IgG antibodies against Vaccinia virus (n=2908), as well as for the ability to neutralize plaque formation with a Vaccinia virus MNIIVP-10 strain (n=299). The results indicate the presence of neutralizing antibody titer of 1/20 or more in 33.3 to 53.2% of people older than 45 years. Among people 30-45 years old who probably have not been vaccinated, the proportion with virus neutralizing antibodies ranged from 3.2 to 6.7%. Despite the higher level of antibodies in age group older than 66 years, the proportion of positive samples in this group was slightly lower than in people aged 46-65 years. The results indicate the priority of vaccination in groups younger than 45, and possibly older than 66 years to ensure the protection of the population in case of spread of monkeypox among Moscow residents. The herd immunity level needed to stop the circulation of the virus should be at least 50.25 - 65.28%.

Sputnik V Effectiveness against Hospitalization with COVID-19 during Omicron Dominance.

Mass vaccination campaigns against COVID-19 affected more than 90% of the population in most developed countries. The new epidemiologic wave of COVID-19 has been ongoing since the end of 2021. It is caused by a virus variant B.1.1.529, also known as "Omicron" and its descendants. The effectiveness of major vaccines against Omicron is not known. The purpose of this study is to evaluate the efficacy of the Sputnik V vaccine. The main goal is to assess its protection against hospitalization in the period of Omicron dominance. We conducted our study based on a large clinical center in Moscow (Russia) where 1112 patients were included. We used the case-population method to perform the calculations. The data we obtained indicate that the Omicron variant causes at least 90% of infections in the studied cohort. The effectiveness of protection against hospitalization with COVID-19 in our study was 85.9% (95% CI 83.0-88.0%) for those who received more than one dose. It was 87.6% (95% CI 85.4-89.5%) and 97.0% (95% CI 95.9-97.8%) for those who received more than two or three doses. The effectiveness in cases of more severe forms was higher than for less severe ones. Thus, present study indicates the high protective efficacy of vaccination against hospitalization with COVID-19 in case of Omicron lineage.

Detection of SARS-CoV-2 in Neonatal Autopsy Tissues and Placenta.

Severe coronavirus disease in neonates is rare. We analyzed clinical, laboratory, and autopsy findings from a neonate in the United States who was delivered at 25 weeks of gestation and died 4 days after birth; the mother had asymptomatic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and preeclampsia. We observed severe diffuse alveolar damage and localized SARS-CoV-2 by immunohistochemistry, in situ hybridization, and electron microscopy of the lungs of the neonate. We localized SARS-CoV-2 RNA in neonatal heart and liver vascular endothelium by using in situ hybridization and detected SARS-CoV-2 RNA in neonatal and placental tissues by using reverse transcription PCR. Subgenomic reverse transcription PCR suggested viral replication in lung/airway, heart, and liver. These findings indicate that in utero SARS-CoV-2 transmission contributed to this neonatal death.

Bone marrow findings of idiopathic Multicentric Castleman disease: A histopathologic analysis and systematic literature review.

Idiopathic multicentric Castleman disease (iMCD) is a polyclonal lymphoproliferative disorder characterized by constitutional symptoms, generalized lymphadenopathy, cytopenias, and multi-organ dysfunction due to excessive cytokines, notably Interleukin-6. Idiopathic multicentric Castleman disease is often sub-classified into iMCD-TAFRO, which is associated with thrombocytopenia (T), anasarca (A), fever/elevated C-reactive protein (F), renal dysfunction (R), and organomegaly (O), and iMCD not otherwise specified (iMCD-NOS), which is typically associated with thrombocytosis and hypergammaglobulinemia. The diagnosis of iMCD is challenging as consensus clinico-pathological diagnostic criteria were only recently established and include several non-specific lymph node histopathological features. Identification of further clinico-pathological features commonly found in iMCD could contribute to more accurate and timely diagnoses. We set out to characterize bone marrow (BM) histopathological features in iMCD, assess differences between iMCD-TAFRO and iMCD-NOS, and determine if these findings are specific to iMCD. Examination of BM specimens from 24 iMCD patients revealed a high proportion with hypercellularity, megakaryocytic atypia, reticulin fibrosis, and plasmacytosis across patients with both iMCD-NOS and iMCD-TAFRO with significantly more megakaryocytic hyperplasia (p = 0.001) in the iMCD-TAFRO cases. These findings were also consistent with BM findings from 185 published cases of iMCD-NOS and iMCD-TAFRO. However, these findings are relatively nonspecific as they can be seen in various other infectious, malignant, and autoimmune diseases.

Isoforms of autophagy-related proteins: role in glioma progression and therapy resistance.

Autophagy is the process of recycling and utilization of degraded organelles and macromolecules in the cell compartments formed during the fusion of autophagosomes with lysosomes. During autophagy induction the healthy and tumor cells adapt themselves to harsh conditions such as cellular stress or insufficient supply of nutrients in the cell environment to maintain their homeostasis. Autophagy is currently seen as a form of programmed cell death along with apoptosis and necroptosis. In recent years multiple studies have considered the autophagy as a potential mechanism of anticancer therapy in malignant glioma. Although, subsequent steps in autophagy development are known and well-described, on molecular level the mechanism of autophagosome initiation and maturation using autophagy-related proteins is under investigation. This article reviews current state about the mechanism of autophagy, its molecular pathways and the most recent studies on roles of autophagy-related proteins and their isoforms in glioma progression and its treatment.

SARS-CoV-2 Infects Endothelial Cells In Vivo and In Vitro.

SARS-CoV-2 infection can cause fatal inflammatory lung pathology, including thrombosis and increased pulmonary vascular permeability leading to edema and hemorrhage. In addition to the lung, cytokine storm-induced inflammatory cascade also affects other organs. SARS-CoV-2 infection-related vascular inflammation is characterized by endotheliopathy in the lung and other organs. Whether SARS-CoV-2 causes endotheliopathy by directly infecting endothelial cells is not known and is the focus of the present study. We observed 1) the co-localization of SARS-CoV-2 with the endothelial cell marker CD31 in the lungs of SARS-CoV-2-infected mice expressing hACE2 in the lung by intranasal delivery of adenovirus 5-hACE2 (Ad5-hACE2 mice) and non-human primates at both the protein and RNA levels, and 2) SARS-CoV-2 proteins in endothelial cells by immunogold labeling and electron microscopic analysis. We also detected the co-localization of SARS-CoV-2 with CD31 in autopsied lung tissue obtained from patients who died from severe COVID-19. Comparative analysis of RNA sequencing data of the lungs of infected Ad5-hACE2 and Ad5-empty (control) mice revealed upregulated KRAS signaling pathway, a well-known pathway for cellular activation and dysfunction. Further, we showed that SARS-CoV-2 directly infects mature mouse aortic endothelial cells (AoECs) that were activated by performing an aortic sprouting assay prior to exposure to SARS-CoV-2. This was demonstrated by co-localization of SARS-CoV-2 and CD34 by immunostaining and detection of viral particles in electron microscopic studies. Moreover, the activated AoECs became positive for ACE-2 but not quiescent AoECs. Together, our results indicate that in addition to pneumocytes, SARS-CoV-2 also directly infects mature vascular endothelial cells in vivo and ex vivo, which may contribute to cardiovascular complications in SARS-CoV-2 infection, including multipleorgan failure.

A Case Report with Severe Thrombocytopenia Induced by Axitinib.

Axitinib is an oral, second-generation tyrosine kinase inhibitor that is selective for vascular endothelial growth factor receptors (VEGFR). This agent is approved as monotherapy or in combination with immune checkpoint inhibitors for the treatment of metastatic renal cell carcinoma. Axitinib is associated with a safety profile very similar to other anti-VEGFR inhibitors but usually with fewer hematologic adverse events, due to the selectivity for VEGF. In this report, we presented a rare case of grade 4 axitinib-induced thrombocytopenia, not observed with other antiangiogenic therapies. We discuss the differential diagnostic work-up, the necessary multidisciplinary approach, and the successful management of the case.

Simultaneous Targeting of PARP1 and RAD52 Triggers Dual Synthetic Lethality in BRCA-Deficient Tumor Cells.

PARP inhibitors (PARPis) have been used to induce synthetic lethality in BRCA-deficient tumors in clinical trials with limited success. We hypothesized that RAD52-mediated DNA repair remains active in PARPi-treated BRCA-deficient tumor cells and that targeting RAD52 should enhance the synthetic lethal effect of PARPi. We show that RAD52 inhibitors (RAD52is) attenuated single-strand annealing (SSA) and residual homologous recombination (HR) in BRCA-deficient cells. Simultaneous targeting of PARP1 and RAD52 with inhibitors or dominant-negative mutants caused synergistic accumulation of DSBs and eradication of BRCA-deficient but not BRCA-proficient tumor cells. Remarkably, Parp1-/-;Rad52-/- mice are normal and display prolonged latency of BRCA1-deficient leukemia compared with Parp1-/- and Rad52-/- counterparts. Finally, PARPi+RAD52i exerted synergistic activity against BRCA1-deficient tumors in immunodeficient mice with minimal toxicity to normal cells and tissues. In conclusion, our data indicate that addition of RAD52i will improve therapeutic outcome of BRCA-deficient malignancies treated with PARPi.

Tyrosine kinase inhibitor-induced defects in DNA repair sensitize FLT3(ITD)-positive leukemia cells to PARP1 inhibitors.

Mutations in FMS-like tyrosine kinase 3 (FLT3), such as internal tandem duplications (ITDs), can be found in up to 23% of patients with acute myeloid leukemia (AML) and confer a poor prognosis. Current treatment options for FLT3(ITD)-positive AMLs include genotoxic therapy and FLT3 inhibitors (FLT3i's), which are rarely curative. PARP1 inhibitors (PARP1i's) have been successfully applied to induce synthetic lethality in tumors harboring BRCA1/2 mutations and displaying homologous recombination (HR) deficiency. We show here that inhibition of FLT3(ITD) activity by the FLT3i AC220 caused downregulation of DNA repair proteins BRCA1, BRCA2, PALB2, RAD51, and LIG4, resulting in inhibition of 2 major DNA double-strand break (DSB) repair pathways, HR, and nonhomologous end-joining. PARP1i, olaparib, and BMN673 caused accumulation of lethal DSBs and cell death in AC220-treated FLT3(ITD)-positive leukemia cells, thus mimicking synthetic lethality. Moreover, the combination of FLT3i and PARP1i eliminated FLT3(ITD)-positive quiescent and proliferating leukemia stem cells, as well as leukemic progenitors, from human and mouse leukemia samples. Notably, the combination of AC220 and BMN673 significantly delayed disease onset and effectively reduced leukemia-initiating cells in an FLT3(ITD)-positive primary AML xenograft mouse model. In conclusion, we postulate that FLT3i-induced deficiencies in DSB repair pathways sensitize FLT3(ITD)-positive AML cells to synthetic lethality triggered by PARP1i's. Therefore, FLT3(ITD) could be used as a precision medicine marker for identifying AML patients that may benefit from a therapeutic regimen combining FLT3 and PARP1i's.

Novel Fluorinated Porphyrins Sensitize Tumor Cells to Photodamage in Normoxia and Hypoxia: Synthesis and Biocompatible Formulations.

BACKGROUND: Hypoxia renders tumor cells refractory to treatment. One way to overcome this problem is the design of drug delivery systems that contain the antitumor agent within an oxygen supply medium. OBJECTIVE: to evaluate whether the perfluorocarbon liquids (capable of retaining up to 50% v/v amounts of O2 gas) can be tools for delivery of photosensitizers to hypoxic tumors. METHOD: We synthesized a series of compounds in which fluoroaliphatic or fluoroaromatic moieties were conjugated to the porphyrin ring in meso-positions. Two derivatives were tested as the solutions prepared either from a dimethylformamide stock ('free' formulation) or from a perfluorocarbon emulsion in which the photosensitizer is entrapped in the oxygenated medium. RESULTS: In the emulsion the hydrophobic photosensitizer and the gas transporting liquid represented a biocompatible composition. Free formulations or perfluorocarbon emulsions of fluorinated porphyrins evoked little-to-null dark cytotoxicity. In contrast, each formulation triggered cell death upon light activation. Photodamage in the presence of fluorinated porphyrins was achievable not only at normoxic (20.9% O2 v/v) conditions but also in hypoxia (0.5% O2). With new compounds dissolved in the medium the cell photodamage in hypoxia was negligible whereas a significant photodamage was achieved with the emulsions of fluorinated porphyrins. The derivative with the fluoroalkyl substituent was more potent than its structurally close analog carrying the fluoroaryl moiety. CONCLUSION: Our new fluorinated porphyrin derivatives, especially their emulsions in which the photosensitizer and the oxygenated medium are coupled into one phase, can be perspective for photoelimination of hypoxic tumor cells.

Ruxolitinib-induced defects in DNA repair cause sensitivity to PARP inhibitors in myeloproliferative neoplasms.

Myeloproliferative neoplasms (MPNs) often carry JAK2(V617F), MPL(W515L), or CALR(del52) mutations. Current treatment options for MPNs include cytoreduction by hydroxyurea and JAK1/2 inhibition by ruxolitinib, both of which are not curative. We show here that cell lines expressing JAK2(V617F), MPL(W515L), or CALR(del52) accumulated reactive oxygen species-induced DNA double-strand breaks (DSBs) and were modestly sensitive to poly-ADP-ribose polymerase (PARP) inhibitors olaparib and BMN673. At the same time, primary MPN cell samples from individual patients displayed a high degree of variability in sensitivity to these drugs. Ruxolitinib inhibited 2 major DSB repair mechanisms, BRCA-mediated homologous recombination and DNA-dependent protein kinase-mediated nonhomologous end-joining, and, when combined with olaparib, caused abundant accumulation of toxic DSBs resulting in enhanced elimination of MPN primary cells, including the disease-initiating cells from the majority of patients. Moreover, the combination of BMN673, ruxolitinib, and hydroxyurea was highly effective in vivo against JAK2(V617F)+ murine MPN-like disease and also against JAK2(V617F)+, CALR(del52)+, and MPL(W515L)+ primary MPN xenografts. In conclusion, we postulate that ruxolitinib-induced deficiencies in DSB repair pathways sensitized MPN cells to synthetic lethality triggered by PARP inhibitors.

Giant cystic sacral schwannoma mimicking tarlov cyst: a case report.

PURPOSE: To present a rare case of a giant schwannoma of the sacrum mimicking a Tarlov cyst. METHODS: A 58-year-old woman had a 1-year history of low back pain. MRI revealed a large cystic mass in the sacral canal with bony erosion. Radiological diagnosis of Tarlov cyst was made. RESULTS: The patient underwent surgical treatment for the lesion, which revealed a solid mass. Histopathological examination of the tumor confirmed the diagnosis of schwannoma. The postoperative course was uneventful and the patient has had significant improvement in her pain 1 month postoperatively. CONCLUSION: Giant cystic schwannoma of the sacrum is a very rare diagnosis overlooked by practitioners for more common cystic etiologies, but its treatment is significantly different. Care should be taken to include this diagnosis in a differential for a cystic sacral mass.

Effect of Quadrivalent Human Papillomavirus Vaccination on Oral Squamous Cell Papillomas.

IMPORTANCE: Cutaneous verruca vulgaris lesions (warts) and oral squamous cell papillomas are common lesions caused by human papillomavirus (HPV). Multiple reports have described cases of wart resolution following quadrivalent HPV vaccination. We report the case of a patient with chronic oral papillomas with resolution after quadrivalent HPV vaccination and perform a review of the literature. OBSERVATIONS: An immunocompetent man in his 60s presented with chronic verrucous papules on the lips, tongue, and buccal mucosa refractory to multiple excisions. Biopsy showed squamous cell papilloma, and DNA sequencing revealed HPV-32. He received the quadrivalent HPV vaccine resulting in clearance of all lesions after 3 months. We found 8 reported cases of disseminated, recurrent warts with resolution after quadrivalent HPV vaccination. Improvement was seen within 4 weeks of vaccination, and resolution after 3 to 8 months. CONCLUSIONS AND RELEVANCE: We report the case of recurrent oral papillomas caused by HPV-32 with complete resolution after quadrivalent HPV vaccination and reviewed reports of resolution of recalcitrant and disseminated warts after vaccination. Production of cross-protective immunoglobulins and cytotoxic T cells is a possible mechanism. There remains a critical need for randomized clinical trials to assess efficacy of quadrivalent HPV vaccination for treatment of oral squamous papillomas and cutaneous verruca vulgaris.