Foci Formation Research Articles

DISTRIBUTION PATTERNS OF METACERCARIAE OF THE TREMATODA <i>APOPHALLUS </i><i>MUEHLINGI</i> (JÄGERSKIÖLD, 1899) IN UNDERYEARLINGS IN AN INVASIVE POPULATION OF <i>CLUPEONELLA CULTRIVENTRIS</i> (NORDMANN, 1840) FROM THE GORKY RESERVOIR (UPPER VOLGA BASIN)

After the successful naturalization of the prosobranch mollusk Lithoglyphus naticoides (C. Pfeiffer, 1828) and a number of associated trematodes in the Gorky Reservoir (in 2005-2015) an intensive spread of apophallesis in fish of this water body has been observed. As one of the second intermediate hosts of the trematode causing this disease, Apophallus muehlingi (Jägerskiöld, 1899), a freshwater form of the Black Sea-Caspian kilka Clupeonella cultriventris (Nordmann, 1840) was recorded. The aim of the work is to study the consequences of dispersal of C. cultriventris for the formation of large permanent foci of apophallesis in the Upper Volga. For comparison, data on the spread of this type of "black-spot" disease in the populations of C. cultriventris from the Gorky Reservoir and an unregulated area of the Lower Volga (downstream the cascade of Volga reservoirs, pelagic trawl coordinates from 47°08ʹN, 47°17ʹE to 46°51ʹN, 47°41ʹE) were used. The high rates of occurrence of A. muehlingi metacercariae (the infection prevalence P, %) are recorded in the fall along the entire area of the Gorky Reservoir: from the northernmost site near Rybinsk (58°01ʹN, 39°06ʹE) to the dam area (56°41ʹN, 43°21ʹE). It is found that the distribution pattern of A. muehling metacercariae varies in the sections of the reservoir differing in hydrological characteristics. In the middle (transitional type) section of the reservoir the prevalence of A. muehlingi metacercariae in groups of the smallest Black Sea-Caspian kilka fingerlings (with a body length of up to 45 mm), is close to the maximally possible (30.00-94.49%). In the upper (river-type) section of the reservoir, the prevalence of metacercariae in this group of fish does not exceed 24.39%. The minimum values of this indicator are found in the lower (lake-type) section of the reservoir (6.76-14.84%). Judging by the values of the abundance index and the coefficient of variation in the infection intensity, the aggregation of metacercariae is more pronounced in the groups of faster growing underyearlings and the Black Sea-Caspian kilka yearlings as compared to the groups of the smallest fingerlings. In the Black Sea-Caspian kilka fingerlings samples from the Lower Volga in 2020 reveals no statistically significant differences between the infection prevalence values (13.33-18.11%) in the different size groups. Thus, although the main second intermediate hosts of A. muehlingi are various fish species of the Cyprinidae family, in some cases the presence of C. cultriventris in the water body ecosystem can significantly increase the likelihood of apophallesis spread. In conditions of the Upper Volga, due to the high migration activity of large individuals of the Black Sea-Caspian kilka, a continuous zone of apophallesis can form without clearly defined boundaries between the individual foci of disease.

Yap1-mediated Flr1 expression reveals crosstalk between oxidative stress signaling and caffeine resistance in Saccharomyces cerevisiae.

Caffeine, a methylxanthine derivative, affects various physiological conditions such as cell growth, proliferation, and energy metabolism. A genome-wide screening for genes required for caffeine resistance in Schizosaccharomyces pombe revealed several candidates, including Pap1 and downstream target genes involved in caffeine efflux. We found that Yap1, a budding yeast AP-1 homolog required for oxidative stress response, has a caffeine tolerance function. Although the Yap1 mutant is not sensitive to caffeine, overexpression of Yap1 renders cells resistant to high concentrations of caffeine. Caffeine sensitivity of mutants lacking two multidrug transporters, Pdr5 or Snq2, is completely recovered by Yap1 overexpression. Among Yap1-dependent target genes, FLR1, a fluconazole-resistant gene, is necessary but not sufficient for caffeine tolerance. Low concentrations of hydrogen peroxide induce Yap1 activation, which restores cell viability against caffeine toxicity. Intriguingly, oxidative stress-mediated cellular adaptation to caffeine toxicity requires Yap1, but not Flr1. Moreover, caffeine is involved in reduction of intracellular reactive oxygen species (ROS), as well as mutation rate and Rad52 foci formation. Altogether, we identified novel reciprocal crosstalk between ROS signaling and caffeine resistance.

Possibilities of combined intranasal drugs in the treatment of patients with rhinosinusitis

Rhinosinusitis – an inflammatory process in the nasal cavity and paranasal sinuses is a global problem of practical medicine, which remains relevant for many years. The most common cause of acute rhinosinusitis is a viral infection, only in 0.2–5 % of patients the disease is further complicated by the formation of a bacterial focus of inflammation. Treatment of patients withARS involves the use of etiotropic, pathogenetic and symptomatic therapy. National clinical guidelines consider the use of systemic and topical etiotropic drugs, intranasal glucocorticosteroids, topical decongestants, irrigation- elimination procedures, mucoactive therapy, standardized herbal medicine, drainage of the paranasal sinuses, antihistamines (with concomitant allergies) and some other activities. Local administration of drugs in patients with rhinosinusitis provides targeted delivery of the active substance to the pathological focus and the creation of its required concentration, the virtual absence of systemic action due to low absorption, as well as a favorable softening, moisturizing, enveloping effect of auxiliary components. One of these drugs is the combined nasal spray containing tuaminoheptane sulfate and acetylcysteine. The effectiveness and safety of the use of this combination has been demonstrated by the results of a fairly large amount of research. The mild and sufficiently long vasoconstrictive effect of tuaminoheptane sulfate, the direct mucolytic, antioxidant and anti-inflammatory effects of acetylcysteine and the high safety of this combination determine its therapeutic value in children from 6 years of age and adult patients with rhinitis and rhinosinusitis.

Radiosensitizing Effect of Trabectedin on Human Soft Tissue Sarcoma Cells.

Trabectedin is used for the treatment of advanced soft tissue sarcomas (STSs). In this study, we evaluated if trabectedin could enhance the efficacy of irradiation (IR) by increasing the intrinsic cell radiosensitivity and modulating tumor micro-environment in fibrosarcoma (HS 93.T), leiomyosarcoma (HS5.T), liposarcoma (SW872), and rhabdomyosarcoma (RD) cell lines. A significant reduction in cell surviving fraction (SF) following trabectedin + IR compared to IR alone was observed in liposarcoma and leiomyosarcoma (enhancement ratio at 50%, ER50: 1.45 and 2.35, respectively), whereas an additive effect was shown in rhabdomyosarcoma and fibrosarcoma. Invasive cells' fraction significantly decreased following trabectedin ± IR compared to IR alone. Differences in cell cycle distribution were observed in leiomyosarcoma and rhabdomyosarcoma treated with trabectedin + IR. In all STS lines, trabectedin + IR resulted in a significantly higher number of γ-H2AX (histone H2AX) foci 30 min compared to the control, trabectedin, or IR alone. Expression of ATM, RAD50, Ang-2, VEGF, and PD-L1 was not significantly altered following trabectedin + IR. In conclusion, trabectedin radiosensitizes STS cells by affecting SF (particularly in leiomyosarcoma and liposarcoma), invasiveness, cell cycle distribution, and γ-H2AX foci formation. Conversely, no synergistic effect was observed on DNA damage repair, neoangiogenesis, and immune system.

Targeting a Highly Deregulated eIF4F Translation Initiation Complex Sensitises IM-Resistant Cells to Tyrosine Kinase Inhibitors and Effectively Suppresses BCR-ABL1 Protein Expression

Background We previously reported that Abelson helper integration site-1 (AHI-1) is highly deregulated in chronic myeloid leukemia (CML) leukemic stem cells (LSCs) and mediates tyrosine kinase inhibitor (TKI) resistance. AHI-1 physically interacts with BCR-ABL1 and its SH3 domain interacts with BCR-ABL1 substrates, like dynamin-2, which regulate leukemic properties of CML LSCs. However, the molecular and biological roles of AHI-1 and its interacting partners in mediating TKI resistance remain largely unknown. Objective To investigate the molecular functions of AHI-1 and its SH3 domain in regulation of TKI resistance, using advanced antibody microarray analysis. Methods A high content antibody microarray was performed in BCR-ABL1+ cells co-transduced with wild-type (WT) AHI-1 or the deletion of AHI-1 SH3 domain (SH3Δ) with or without imatinib (IM). Changes in antibody signals for protein expression or phosphorylation were determined using limma and pathway enrichment analysis by g:Profiler. eIF4G1 genetic inhibition by lentiviral-mediated shRNA or pharmacological inhibition by SBI-756, was performed in TKI-resistant cells to assay translation initiation activity by proximity ligation assay (PLA), o-propargyl-puromycin (OPP), and polysome profiling. Results The antibody microarray analysis revealed that WT AHI-1 cells have the greatest number of changes in the phospho-proteome and proteome compared to BCR-ABL1+ cells and AHI-1 SH3Δ cells with and without IM. Pathway enrichment analysis identified that the targets with significantly increased differential antibody signal after IM treatment in WT AHI-1 cells were related to the regulation of translation initiation complex (p < 0.0001). Interestingly, our RNA-seq dataset analysis further identified several eukaryotic initiation factor 4F (eIF4F) complex members to be significantly increased in CD34+ CML patient cells compared to normal bone marrow, particularly eIF4G1, the scaffold protein of the eIF4F complex (2-fold, p = 0.001). WT AHI-1 cells also showed increased expression and phosphorylation of eIF4G1 (>2-fold) and eIF4B (>2-fold), a cofactor that regulates the helicase activity of the eIF4F complex, and cyclin D3 (a downstream protein of eIF4F translational activity) as compared to BCR-ABL1+ cells, by immunoblotting. These results were similarly demonstrated in IM-resistant cells as compared to IM-sensitive cells (2-5-fold, p < 0.05). Mechanistically, eIF4G1 knockdown by shRNA impaired survival (5-fold, p < 0.0001) and increased TKI sensitivity in IM-resistant cells (p < 0.0021). These cells showed reduced levels of eIF4F complex formation by PLA assay (p = 0.013) and reduced protein expression of cyclin D3 (40%, p = 0.0036) and BCR-ABL1 (70%). Similarly, IM-resistant cells were more sensitive to SBI-756 treatment, an eIF4G1 inhibitor (50% reduction, p = 0.0001) than IM-sensitive cells (30% reduction); these effects were enhanced by a combination of SBI-756 and IM (80%, p < 0.02). Furthermore, SBI-756 treatment reduced PLA foci formation (p = 0.014) and global protein synthesis rates (p < 0.0001). Polysome profiling demonstrated that IM-resistant cells had higher levels of translation activity, which was inhibited by SBI-756 treatment. Most interestingly, SBI-756 treatment reduced protein expression of BCR-ABL1 (70%) and cyclin D3 (40%) in these cells. Conclusion We have uncovered that the eIF4F complex, the key regulator of the mRNA-ribosome recruitment phase of translation initiation, has increased activity in IM-resistant cells. eIF4G1 inhibitor treatment sensitises IM-resistant cells to TKI and reduces BCR-ABL1 protein expression, providing potential treatment strategies to overcome TKI resistance.

DNAR-06. LOSS OF HISTONE H3 THREONINE 45 PHOSPHORYLATION DECREASES H3K36ME3 TO ABROGATE THE RADIATION-INDUCED DNA DAMAGE IN GLIOBLASTOMA MULTIFORME

Abstract Glioblastoma multiforme (GBM) is the most common and fatal adult brain tumour. As radioresistance is a common occurrence and contributes to poor patient prognosis, exploiting the mechanistic basis of GBM radioresistance to develop new therapeutic approaches remains an unmet clinical need. Chromatin post-translational modifications (PTMs) can augment cancer radioresistance. Several chromatin PTMs are regulated by PI3K/AKT, which is frequently hyperactivated in GBM. Previous research demonstrated that the PI3K effector AKT phosphorylates histone H3 at threonine 45 (pH3T45) in response to DNA damage. To genetically dissect the function of H3T45 phosphorylation in the DNA damage response, we engineered the PI3K pathway-activated U87MG GBM cell line to stably overexpress wildtype H3, or a phospho-null H3T45A mutant. We show that in both the presence and absence of irradiation, cells harbouring the phospho-null H3T45A mutant decreases H3K36me3 on free and nucleosome-incorporated histones, which may suggest a deficiency in DNA repair protein recruitment. By surveying irradiation-induced 53BP1 foci formation, we demonstrate that loss of pH3T45 compromises damage repair through NHEJ by delaying the onset of repair signaling. We also observe a defect in DNA damage resolution, which enhances apoptosis to reduce cell clonogenicity and proliferative capacity. Irradiation does not further reduce clonogenicity and proliferation in the phospho-null H3T45A-expressing cells. Taken together, these data suggest a critical function for PI3K-directed H3T45 phosphorylation in DNA damage repair and that loss of pH3T45 may contribute to GBM radioresistance. We anticipate that further characterization of this phosphorylation event will inform the understanding of GBM radioresistance and may suggest novel therapeutic strategies.

Changes in the Fecal Metabolome Accompany an Increase in Aberrant Crypt Foci in the Colon of C57BL/6 Mice Fed with a High-Fat Diet.

High-fat diet (HFD)-induced obesity is a risk factor for colon cancer. Our previous data show that compared to an AIN-93 diet (AIN), a HFD promotes azoxymethane (AOM)-induced colonic aberrant crypt foci (ACF) formation and microbial dysbiosis in C57BL/6 mice. To explore the underlying metabolic basis, we hypothesize that AOM treatment triggers a different fecal metabolomic profile in C57BL/6 mice fed the HFD or the AIN. We found that 65 of 196 identified metabolites were significantly different among the four groups of mice (AIN, AIN + AOM, HFD, and HFD + AOM). A sparse partial least squares discriminant analysis (sPLSDA) showed that concentrations of nine fecal lipid metabolites were increased in the HFD + AOM compared to the HFD, which played a key role in overall metabolome group separation. These nine fecal lipid metabolite concentrations were positively associated with the number of colonic ACF, the cell proliferation of Ki67 proteins, and the abundance of dysbiotic bacteria. These data suggest that the process of AOM-induced ACF formation may increase selective fecal lipid concentrations in mice fed with a HFD but not an AIN. Collectively, the accumulation of these critical fecal lipid species may alter the overall metabolome during tumorigenesis in the colon.

PRIMPOL competes with RAD51 to resolve G-quadruplex-induced replication stress via its interaction with RPA.

PRIMPOL (primase-polymerase) is a recently discovered DNA primase-polymerase involved in DNA damage tolerance and replication stress response in eukaryotic cells. However, the detailed mechanism of the PRIMPOL response to replication stress remains elusive. Here, we demonstrate that replication-related factors, including replication protein A (RPA), regulate the accumulation of PRIMPOL in subnuclear foci in response to replication stress induced by replication inhibitors. Moreover, PRIMPOL works at G-quadruplexes (G4s) in human cells to resolve the replication stress induced by G4s. The formation of PRIMPOL foci persists throughout the cell cycle. We further demonstrate that PRIMPOL competes with RAD51 to resolve G4-induced replication stress. In conclusion, our results provide novel insight into the mechanism of PRIMPOL in G4s to resolve replication stress and competition between PRIMPOL (repriming)- and RAD51 (fork reversal)-mediated pathways, which indicates a new strategy to improve the tumor response to DNA-damaging chemotherapy by targeting the PRIMPOL pathway.

Lack of peroxisomal catalase affects heat shock response in Caenorhabditis elegans.

Exact mechanisms of heat shock-induced lifespan extension, although documented across species, are still not well understood. Here, we show that fully functional peroxisomes, specifically peroxisomal catalase, are needed for the activation of canonical heat shock response and heat-induced hormesis in Caenorhabditis elegans Although during heat shock, the HSP-70 chaperone is strongly up-regulated in the WT and in the absence of peroxisomal catalase (ctl-2(ua90)II), the small heat shock proteins display modestly increased expression in the mutant. Nuclear foci formation of HSF-1 is reduced in the ctl-2(ua90)II mutant. In addition, heat-induced lifespan extension, observed in the WT, is absent in the ctl-2(ua90)II strain. Activation of the antioxidant response and pentose phosphate pathway are the most prominent changes observed during heat shock in the WT worm but not in the ctl-2(ua90)II mutant. Involvement of peroxisomes in the cell-wide cellular response to transient heat shock reported here gives new insight into the role of organelle communication in the organism's stress response.

Multivalent binding of the hub protein LC8 at a newly discovered site in 53BP1

Tumor suppressor p53 binding protein 1 (53BP1) is a scaffolding protein involved in poly-ADP ribose polymerase inhibitor hypersensitivity in BRCA1-negative cancers. 53BP1 plays a critical role in the DNA damage response and relies on its oligomerization to create foci that promote repair of DNA double-strand breaks. Previous work shows that mutation of either the oligomerization domain or the dynein light chain 8 (LC8)-binding sites of 53BP1 results in reduced accumulation of 53BP1 at double-strand breaks. Mutation of both abolishes focus formation almost completely. Here, we show that, contrary to current literature, 53BP1 contains three LC8-binding sites, all of which are conserved in mammals. Isothermal titration calorimetry measuring binding affinity of 53BP1 variants with LC8 shows that the third LC8-binding site has a high affinity and can bind LC8 in the absence of other sites. NMR titrations confirm that the third site binds LC8 even in variants that lack the other LC8-binding sites. The third site is the closest to the oligomerization domain of 53BP1, and its discovery would challenge our current understanding of the role of LC8 in 53BP1 function.

Amorfrutin B Protects Mouse Brain Neurons from Hypoxia/Ischemia by Inhibiting Apoptosis and Autophagy Processes Through Gene Methylation- and miRNA-Dependent Regulation

Amorfrutin B is a selective modulator of the PPARγ receptor, which has recently been identified as an effective neuroprotective compound that protects brain neurons from hypoxic and ischemic damage. Our study demonstrated for the first time that a 6-h delayed post-treatment with amorfrutin B prevented hypoxia/ischemia-induced neuronal apoptosis in terms of the loss of mitochondrial membrane potential, heterochromatin foci formation, and expression of specific genes and proteins. The expression of all studied apoptosis-related factors was decreased in response to amorfrutin B, both during hypoxia and ischemia, except for the expression of anti-apoptotic BCL2, which was increased. After post-treatment with amorfrutin B, the methylation rate of the pro-apoptotic Bax gene was inversely correlated with the protein level, which explained the decrease in the BAX/BCL2 ratio as a result of Bax hypermethylation. The mechanisms of the protective action of amorfrutin B also involved the inhibition of autophagy, as evidenced by diminished autophagolysosome formation and the loss of neuroprotective properties of amorfrutin B after the silencing of Becn1 and/or Atg7. Although post-treatment with amorfrutin B reduced the expression levels of Becn1, Nup62, and Ambra1 during hypoxia, it stimulated Atg5 and the protein levels of MAP1LC3B and AMBRA1 during ischemia, supporting the ambiguous role of autophagy in the development of brain pathologies. Furthermore, amorfrutin B affected the expression levels of apoptosis-focused and autophagy-related miRNAs, and many of these miRNAs were oppositely regulated by amorfrutin B and hypoxia/ischemia. The results strongly support the position of amorfrutin B among the most promising anti-stroke and wide-window therapeutics.

Upregulation of Long Noncoding RNA MALAT1 in Colorectal Cancer Promotes Radioresistance and Aggressive Malignance.

Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), a conserved transcript with 8000 nt, is highly associated with malignancy of numerous cancer types. However, the function of MALAT1 plays in regulating the response to radiotherapy in colorectal cancer (CRC) remains unclear. Thus, the object of this study is to investigate the functions of MALAT1 in CRC radioresistance. First, the expression of MALAT1 in colon adenocarcinoma (COAD) was analyzed through the Cancer Genome Atlas (TCGA) database. Then, we detected the expression level of MALAT1 in tumor tissues and CRC cell lines and analyzed the relevance of MALAT1 and clinicopathological parameters. In the end, the effect of silencing MALAT1 on the radiosensitivity of CRC cells was investigated, and its potential mechanism was preliminarily illustrated. The analysis of TCGA data showed that MALAT1 was closely related to the type of tumor, and high expression of MALAT1 was remarkably relevant to poor outcome. MALAT1 was highly expressed in CRC tissues and cell lines and related to tumor stages. Knockdown of MALAT1 could significantly suppress colony survival, proliferation, and migration and increase apoptosis, G2/M phase arrest, and formation of gamma-H2AX foci in HCT116, whether in combination with X-rays or not. Moreover, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis demonstrated that the regulated proteins were principally enriched in the glycosaminoglycan degradation pathway after silencing MALAT1. Our results implied that MALAT1 was highly expressed in CRC and associated with tumor stage and prognosis. Silencing MALAT1 can increase HCT116 cell radiosensitivity, which may be potentially influenced by glycosaminoglycan degradation pathway.

Enhanced DNA repair through droplet formation and p53 oscillations

Living organisms are constantly exposed to DNA damage, and optimal repair is therefore crucial. A characteristic hallmark of the response is the formation of sub-compartments around the site of damage, known as foci. Following multiple DNA breaks, the transcription factor p53 exhibits oscillations in its nuclear concentration, but how this dynamics can affect the repair remains unknown. Here, we formulate a theory for foci formation through droplet condensation and discover how oscillations in p53, with its specific periodicity and amplitude, optimize the repair process by preventing Ostwald ripening and distributing protein material in space and time. Based on the theory predictions, we reveal experimentally that the oscillatory dynamics of p53 does enhance the repair efficiency. These results connect the dynamical signaling of p53 with the microscopic repair process and create a new paradigm for the interplay of complex dynamics and phase transitions in biology.

&lt;i&gt;Yersinia pestis&lt;/i&gt; Strains of the 1.ORI Line as Etiological Agent of the Plague Pandemic III

Yersinia pestis strains of the 1.ORI lineage originate from China as a result of evolution of the 1.ANT phylogenetic branch. Strains of the biovar orientalis are divided into three major lines of evolution: 1.ORI1, 1.ORI2, 1.ORI3. Lines 1.ORI1 and 1.ORI2 originated in China and then spread across the east and west coasts of India, respectively. Strains of the biovar orientalis have widely spread throughout the world, mainly as a result of introduction by sea. This way, the 1.ORI1 line was imported onto the territory of North America. 1.ORI2 line has spread to Southeast Asia, Africa, Europe, and South America. In addition, the strains of the biovar orientalis were brought to the territory of Australia, however, the formation of natural foci did not occur. The spread of strains to new territories during the third plague pandemic, as a rule, took place with the participation of one strain, which caused epizootics among synanthropic rodents. After that, outbreaks were recorded among the population of port cities, followed by drifting into the countryside and the formation of natural foci under suitable natural conditions. In the absence of such, the plague pathogen was eliminated from natural biotopes, and the formation of a natural focus did not occur. In recent decades, most cases of human plague in the world have been caused by strains of the biovar orientalis (1.ORI). However, the emergence and spread of the evolutionary line “1” is insufficiently studied. Currently, there is a lack of both historical data and strains that are ancestors of modern strains in many countries to clarify the details of the irradiation of strains of the biovar orientalis. As a result, the concepts of dissemination of many evolution branches of the strains, biovar orientalis are in the form of hypotheses to date. In this work, the collection and analysis of literature data on the history and epidemiology of plague over the third pandemic, a search for a connection between epidemic manifestations and the appurtenance of the strains that caused them to certain phylogenetic lineages was carried out.

Planispine A Sensitized Cancer Cells to Cisplatin by Inhibiting the Fanconi Anemia Pathway.

The use of cisplatin as a chemotherapeutic drug is impeded by the development of drug resistance. Combination therapies of a chemosensitizer for cisplatin have been studied, but with little success, and the search for an effective combination therapy is continuing. Our earlier reports have shown that Zanthoxylum armatum DC. extract enhances the apoptotic effect of cisplatin in cancer cell lines. In this study, we purified and identified the bioactive phytocompound through bio-assay-guided purification, using column chromatography and HPLC. Chemical characterization using NMR and mass spectrometry revealed the compound as planispine A, with molecular structure C25H30O6 and molecular weight, 426.16 g/mol. Planispine A was found to inhibit cancer cell proliferation in a dose-dependent manner and to sensitize the cancer cells to cisplatin-augmented apoptotic cell death, in a caspase-dependent manner. A combination of planispine A and cisplatin induced S-phase cell cycle arrest, and reduced the expression of survival proteins such as cyclin D1. Interestingly, planispine A inhibits the Fanconi anemia pathway, as shown by reduced FANCD2 foci formation and FANCD2 monoubiquitination, which revealed the molecular mechanism of chemo-sensitization of cancer cells to cisplatin. Evaluation of this combination therapy in cisplatin-resistant tumors may lead to more efficient cisplatin treatment.

Effects of Irradiation Combined with Trastuzumab on Myocardial Cell Line H9c2 In Vitro

<h3>Purpose/Objective(s)</h3> The current clinical guidelines do not make clear restrictions or affirmation on whether chest radiotherapy can be delivered currently with trastuzumab. We aim to analyze the effect of trastuzumab combined with dose gradient X-ray irradiation on H9c2 cardiomyocytes and detect the relevant mechanism. <h3>Materials/Methods</h3> Trastuzumab was given at the same time with X-ray irradiation of H9c2 cardiomyocytes with dose gradient (0, 2, 5, 10, 15gy). Flow cytometry was used to detect the proportion of apoptosis, a cell counting kit was used to detect the viability of cells, and WB was used to detect apoptosis related proteins for analyzing the damage effect of irradiation combined with trastuzumab on cardiomyocytes. To analyze the mechanism of trastuzumab aggravating myocardial radiotoxicity by detecting ROS content, Akt protein phosphorylation and DNA damage. <h3>Results</h3> 1. Flow cytometry and WB suggested that irradiation could promote the apoptosis of H9c2 cardiomyocytes, which was positively correlated with the dose of irradiation; after trastuzumab treatment, apoptosis was further improved. 2. A cell counting kit assay showed that irradiation combined with trastuzumab could superimpose the inhibitory effect on cardiomyocyte activity. 3. Compared with irradiation alone, irradiation combined with trastuzumab significantly inhibited the phosphorylation of Akt molecules. 4. Trastuzumab could promote the aggregation of ROS in cells by Flow cytometry, and enhance DNA damage induced by irradiation by the formation of fluorescence focus of γ - H2AX. <h3>Conclusion</h3> 1. Radiation injury to isolated cardiomyocytes was dose-dependent. Meanwhile, trastuzumab aggravated the toxicity of irradiation on cardiomyocytes. 2. Trastuzumab may aggravate myocardial radiation injury by inhibiting Akt phosphorylation, promoting ROS accumulation in cells and promoting DNA damage.

Synergistic Prostate Cancer Targeted Radiosensitization by Gold Nanoparticles and Histone Deacetylase Inhibitor Romidepsin

<h3>Purpose/Objective(s)</h3> Histone deacetylase inhibitors (HDACi), such as Romidepsin (FK228) have been confirmed as effective anticancer agents for liquid cancers, but systemic toxicity hinders their application in solid tumors. Gold nanoparticles (GNPs) with tumor specific ligands can be actively accumulated in the targeted tumor, elevating HDACi delivery efficiency. Moreover, GNPs demonstrate their exceptional competence in improving radiosensitization through locally enhanced radiation scatter in the tumor. In this study, we examined the synergistic consequence of prostate cancer treatment with FK228 conjugated GNPs and investigate the molecular mechanism underlying radiosensitization. <h3>Materials/Methods</h3> GNPs were thiolated with N-succinimidyl 3-(2-pyridyldithio)-propionate (SPDP). FK228 and αPSMA antibody were reduced with tris(2-carboxyethyl)-phosphine and mixed with GNPs at a 10:1 ratio, respectively. Dynamic light scattering (DLS) and enzyme linked immunosorbent assay (ELISA) confirmed conjugation, and targeting efficiency was assessed with immunofluorescence (IF) staining of GNPs. PSMA-targeted GNPs (pGNP), and pGNP+FK228 (pfGNP) were treated to LNCaP cells at varying concentrations and monitored on a real time cell analysis (RTCA) system. Western blot (WB) was performed on HDACi pathways. Finally, 2Gy at 160kVp was delivered to cells. 1hr post irradiation, cells were fixed and stained with γH2AX antibodies. Foci count and size were quantified. <h3>Results</h3> DLS showed shift in GNP hydrodynamic size from 49nm to 52nm when conjugated to αPSMA antibody or antibody and FK228. Zeta potential changed from -8mv to -14mV and -20mV in pGNP and pfGNP, respectively. ELISA revealed successful conjugation of αPSMA antibodies with ∼4 antibodies per GNP. IF demonstrated that pfGNP have three-fold higher accumulation on LNCaP cells compared to untargeted GNPs. Based on RTCA, pGNP showed cell killing starting at 1mg/mL at 24hrs, while pfGNP started cell killing at 10µg/mL, but the initiation ranged from 72hrs to 24hrs in a dose dependent manner. At non-toxic concentrations of FK228 and pfGNP, WB revealed both treatments induced a lower expression of androgen receptor and increased expression of ATF3, while pGNP did not. γH2AX assay revealed an increased formation of foci with the treatment of pGNP (158%), FK228 (125%), and pfGNP (170%). There was no statistically significant increase in the foci count between pGNP and pfGNP. However, measurements of total surface area per foci revealed a statistically significant increase in foci size from FK228 and pfGNP compared to pGNP. <h3>Conclusion</h3> FK228 and PSMA-antibodies were successfully conjugated to GNPs using SPDP chemistry. Furthermore, pfGNP demonstrated increased prostate cancer affinity and induced similar cellular signaling as free FK228. Finally, conjugation of FK228 to GNPs synergistically improved prostate cancer radiosensitization through greater induction of double stranded DNA breaks in vitro.

KAT2-mediated acetylation switches the mode of PALB2 chromatin association to safeguard genome integrity.

The tumour suppressor PALB2 stimulates RAD51-mediated homologous recombination (HR) repair of DNA damage, whilst its steady-state association with active genes protects these loci from replication stress. Here, we report that the lysine acetyltransferases 2A and 2B (KAT2A/2B, also called GCN5/PCAF), two well-known transcriptional regulators, acetylate a cluster of seven lysine residues (7K-patch) within the PALB2 chromatin association motif (ChAM) and, in this way, regulate context-dependent PALB2 binding to chromatin. In unperturbed cells, the 7K-patch is targeted for KAT2A/2B-mediated acetylation, which in turn enhances the direct association of PALB2 with nucleosomes. Importantly, DNA damage triggers a rapid deacetylation of ChAM and increases the overall mobility of PALB2. Distinct missense mutations of the 7K-patch render the mode of PALB2 chromatin binding, making it either unstably chromatin-bound (7Q) or randomly bound with a reduced capacity for mobilisation (7R). Significantly, both of these mutations confer a deficiency in RAD51 foci formation and increase DNA damage in S phase, leading to the reduction of overall cell survival. Thus, our study reveals that acetylation of the ChAM 7K-patch acts as a molecular switch to enable dynamic PALB2 shuttling for HR repair while protecting active genes during DNA replication.

KRCC1, a modulator of the DNA damage response.

The lysine-rich coiled-coil 1 (KRCC1) protein is overexpressed in multiple malignancies, including ovarian cancer, and overexpression correlates with poor overall survival. Despite a potential role in cancer progression, the biology of KRCC1 remains elusive. Here, we characterize the biology of KRCC1 and define its role in the DNA damage response and in cell cycle progression. We demonstrate that KRCC1 associates with the checkpoint kinase 1 (CHK1) upon DNA damage and regulates the CHK1-mediated checkpoint. KRCC1 facilitates RAD51 recombinase foci formation and augments homologous recombination repair. Furthermore, KRCC1 is required for proper S-phase progression and subsequent mitotic entry. Our findings uncover a novel component of the DNA damage response and a potential link between cell cycle, associated damage response and DNA repair.

ОСОБЕННОСТИ ЛУЧЕВОЙ ДИАГНОСТИКИ КРОВОИЗЛИЯНИЙ В ОЧАГИ ПОРАЖЕНИЯ ГОЛОВНОГО МОЗГА ПРИ ОНКОГЕМАТОЛОГИЧЕСКОЙ ПАТОЛОГИИ. КЛИНИЧЕСКОЕ НАБЛЮДЕНИЕ

The acute myeloid leukemia (AML) is the malignant transformation and uncontrolled proliferation of abnormally differentiated, long-lived myeloid progenitor cells cause a large number of blast cells in the circulating blood and replacement of normal bone marrow with tumor cells, as well as damage to other organs and systems. Neurological disorders in AML diseases are among the secondary lesions of the nervous system. Morphologically, neuroleukemia is characterized by multifocal leukemic infiltrates and disorders of the myelination of nerve fibers. The formation of leukemic foci in the medulla leads to an increase in intracranial pressure, compression of cerebral and spinal structures, loss of functions of the affected area. In such situations, computed tomography of the brain may reveal lesions of the brain substance, leukemic infiltration of the meninges of the brain, intracranial hemorrhages, and cerebral infarctions.