Proliferative Activity Of Tumor Cells Research Articles

Atox1-cyclin D1 loop activity is critical for survival of tumor cells with inactivated TP53.

The search for relevant molecular targets is one of the main tasks of modern tumor chemotherapy. To successfully achieve this, it is necessary to have the most complete understanding of the functioning of a transcriptional apparatus of the cell, particularly related to proliferation. The p53 protein plays an important role in regulating processes such as apoptosis, repair, and cell division, and the loss of its functionality often accompanies various types of tumors and contributes to the development of chemoresistance. Additionally, the proliferative activity of tumor cells is closely related to the metabolism of transition metals. For example, the metallochaperone Atox1 - a copper transporter protein - acts as a transcription activator for cyclin D1, promoting progression through the G1/S phase of the cell cycle. On the other hand, p53 suppresses cyclin D1 at the transcriptional level, thereby these proteins have divergent effects on cell cycle progression. However, the contribution of the interaction between these proteins to cell survival is poorly understood. This work demonstrates that not only exists a positive feedback loop between Atox1 and cyclin D1 but also that the activity of this loop depends on the status of the TP53 gene. Upon inactivation of TP53 in A549 and HepG2 cell lines, the expression of ATOX1 and CCND1 genes is enhanced, and their suppression in these cells leads to pronounced apoptosis. This fundamental observation may be useful in selecting more precise interventions for combined therapy of p53-negative tumors.

Prognostic evaluation of the Ki-67 labeling system in histological grading of non-small round cell sarcoma: a supplementary analysis of a randomized controlled trial, JCOG1306.

Soft tissue sarcoma (STS) has various histological types and is rare, making it difficult to evaluate the malignancy of each histological type. Thus, comprehensive histological grading is most important in the pathological examination of STS. The Fédération Nationale des Centres de Lutte Contre le Cancer (FNCLCC) grading system is most commonly used in daily pathological analysis of STS. Among the FNCLCC grading system parameters, mitotic count is a key morphological parameter reflecting the proliferative activity of tumor cells, although its reproducibility may be lacking. Here, we compared the prognostic utility of the conventional and modified FNCLCC grading systems in JCOG1306. We analyzed 140 patients with non-small round cell sarcoma. We performed Ki-67 immunostaining using open biopsy specimens before preoperative chemotherapy in all patients. We assessed histological grade in individual cases by conventional FNCLCC grading (tumor differentiation, mitotic count, and necrosis) and modified FNCLCC grading using the Ki-67 labeling index instead of mitotic count. We conducted univariable and multivariable Cox regression analyses to investigate the influence of grade on overall survival. In univariable analysis, prognosis was worse for patients with conventional FNCLCC Grade 3 tumors compared with Grade 1 or 2 tumors (hazard ratio [HR] 4.21, 95% confidence interval [CI] 1.47-12.05, P=0.008). Moreover, prognosis was worse in patients with modified FNCLCC Grade 3 tumors compared with Grade 1 or 2 tumors (HR 4.90, 95% CI 1.64-14.65, P=0.004). In multivariable analysis including both conventional and modified FNCLCC grading, the modified grading more strongly affected overall survival (HR 6.70, 95% CI 1.58-28.40, P=0.010). The modified FNCLCC grading system was superior to the conventional system in predicting the prognosis of patients with non-small round cell sarcoma according to this supplementary analysis of data from the randomized controlled trial JCOG1306.

THE EFFECT OF GLUCOSE ON THE SURVIVAL AND METABOLIC PLASTICITY OF LEWIS LUNG CARCINOMA CELLS DURING THEIR ADHESIVE AND DE-ADHESIVE GROWTH

High proliferative activity of tumor cells requires a sufficient amount of energy and plastic substrates, in particular glucose [1]. Intensive consumption of this metabolite by tumor cells leads to its depletion in the intercellular environment and creates an uneven distribution of glucose in the tumor [2-3]. As a result, some cells can survive can adapt to glucose deficiency by reprogramming their metabolism and/or by migrating to the vasculature. Tumor cells that lose adhesive contact, become resistant to anoikis and acquire metastasis potential [4]. The ability to undergo such reprogramming may significantly depend on the characteristics of the deficient microenvironment preceding the transition of tumor cells from an adhesive to a de-adhesive state.

Treatment strategy and post-treatment management of colorectal neuroendocrine tumor.

Following the increase in colorectal neuroendocrine tumors (NETs), there is a consequent increase in the importance of their appropriate treatment and post-treatment management. It is widely accepted that colorectal NETs sized ≥20mm and those with muscularis propria invasion are indicated for radical surgery, and those sized <10mm without the invasion are indicated for local resection. No consensus has been reached regarding the treatment strategy for those sized 10-19mm without the invasion. Endoscopic resection has become a primary option for the local resection of colorectal NETs. For rectal NETs sized <10mm, modified endoscopic mucosal resection, such as endoscopic submucosal resection with ligation device and endoscopic mucosal resection with a cap-fitted panendoscope, seems favorable because of its ability to achieve a high R0 resection rate, safety, and convenience. Endoscopic submucosal dissection can also be helpful for these lesions; however, this procedure may be more effective for large lesions or those in the colon. Management following local resection of colorectal NETs is based on the pathological evaluation of factors associated with metastasis, including tumor size, invasion depth, tumor cell proliferative activity (NET grading), presence of lymphovascular invasion, and resection margins. There remain unclear issues in managing cases with NET grading ≥2, positive lymphovascular invasion, and positive resection margins following local resection. In particular, there is confusion regarding managing positive lymphovascular invasion because positivity has become remarkably high with the increased use of the immunohistochemical/special staining. Further evidence based on long-term clinical outcomes is required to address these issues.

Prediction of Ki-67 expression in gastrointestinal stromal tumors using radiomics of plain and multiphase contrast-enhanced CT.

To study the value of radiomics models based on plain and multiphase contrast-enhanced CT to predict Ki-67 expression in gastrointestinal stromal tumors (GISTs). A total of 215 patients with GISTs were retrospectively analyzed, including 150 patients in one hospital as the training set and 65 patients in another hospital as the external verification set. The tumor at the largest level of CT images was delineated as the region of interest (ROI). The maximum diameter of the ROI was defined as the tumor size. A total of 851 radiomics features were extracted from each ROI by 3D Slicer Radiomics. After dimensionality reduction, three machine learning classification algorithms including logistic regression (LR), random forest (RF), and support vector machine (SVM) were used for Ki-67 expression prediction. Using a multivariable logistic model, a nomogram was established to predict the expression of Ki-67 individually. Delong tests showed that the SVM models had the highest accuracy in the arterial phase (Z value 0.217-1.139) and venous phase (Z value 0.022-1.396). For the plain phase, LR and SVM models had the highest accuracy (Z value 0.874-1.824, 1.139-1.763). For the delayed phase, LR models had the highest accuracy (Z value 0.056-1.824). For the combined phase, RF models had the highest accuracy (Z value 0.232-1.978). There was no significant difference among the above models for KI-67 expression prediction (Z value 0.022-1.978). A nomogram was developed with a C-index of 0.913 (95% CI, 0.878 to 0.956). Radiomics of both plain and enhanced CT images could accurately predict the expression of Ki-67 in GIST. For patients who were not suitable to use contrast agents, plain scan could be used as an alternative. CT radiomics could accurately predict the expression of Ki-67 in GIST, which has a great clinical value in reflecting the proliferative activity of tumor cells and helping determine whether a patient is suitable for adjuvant therapy with imatinib. • Radiomics of both plain and enhanced CT images could accurately predict the expression of Ki-67 in GIST. • For patients who were not suitable to use contrast agents, plain scan could be used as an alternative. • A radiomics nomogram was developed to allow personalized preoperative evaluation with high accuracy.

Response to primary chemoradiotherapy of locally advanced oropharyngeal carcinoma is determined by the degree of cytotoxic T cell infiltration within tumor cell aggregates.

Effective anti-tumor immune responses are mediated by T cells and require organized, spatially coordinated interactions within the tumor microenvironment (TME). Understanding coordinated T-cell-behavior and deciphering mechanisms of radiotherapy resistance mediated by tumor stem cells will advance risk stratification of oropharyngeal cancer (OPSCC) patients treated with primary chemoradiotherapy (RCTx). To determine the role of CD8 T cells (CTL) and tumor stem cells for response to RCTx, we employed multiplex immunofluorescence stains on pre-treatment biopsy specimens from 86 advanced OPSCC patients and correlated these quantitative data with clinical parameters. Multiplex stains were analyzed at the single-cell level using QuPath and spatial coordination of immune cells within the TME was explored using the R-package Spatstat. Our observations demonstrate that a strong CTL-infiltration into the epithelial tumor compartment (HR for overall survival, OS: 0.35; p<0.001) and the expression of PD-L1 on CTL (HR: 0.36; p<0.001) were both associated with a significantly better response and survival upon RCTx. As expected, p16 expression was a strong predictor of improved OS (HR: 0.38; p=0.002) and correlated with overall CTL infiltration (r: 0.358, p<0.001). By contrast, tumor cell proliferative activity, expression of the tumor stem cell marker CD271 and overall CTL infiltration, regardless of the affected compartment, were not associated with response or survival. In this study, we could demonstrate the clinical relevance of the spatial organization and the phenotype of CD8 T cells within the TME. In particular, we found that the infiltration of CD8 T cells specifically into the tumor cell compartment was an independent predictive marker for response to chemoradiotherapy, which was strongly associated with p16 expression. Meanwhile, tumor cell proliferation and the expression of stem cell markers showed no independent prognostic effect for patients with primary RCTx and thus requires further study.

R-DA-Edoch/R-DHAP Alteration Could Overcome High Risk Factors in Younger Mantle Cell Lymphoma

Background: Mantle-cell lymphoma (MCL) is a rare subtype of non-Hodgkin's lymphoma, with a median overall survival of only 3~5 years in the era of immunochemotherapy. Although first-line intensive chemotherapy with rituximab (R) and high dose cytarabine combined with autologous transplantation (ASCT) has significantly prolonged the survival of patients with MCL, some patients still die within few years. Our published data show that Chinese patients with MCL are younger and have a higher proportion of TP53 deletion and MYC abnormality [Oncotarget, 2015 Dec 8;6(39):42362-71]. Considering the efficacy and tolerability of the dose-enhanced immunochemotherapy R-DA-EDOCH regimen in patients with multiple high-risk aggressive B-cell lymphomas and the potential higher proliferative activity of tumor cells in Chinese patients, we conducted a prospective clinical study of alternate R-DA-EDOCH/R-DHAP induction therapy for young patients with newly-diagnosed MCL (NCT02858804). Here, we analyzed the efficacy and safety of this regimen. Methods: The main inclusion criteria for patients with newly-diagnosed MCL included age ≤ 65 years, Ann Arbor stage II~IV and ECOG ≤ 1. During induction, patients received alternate R-DA-EDOCH/R-DHAP for two cycles. Patients who achieved less than partial remission would quit the trial, while the other patients would continue another one cycle of treatment. After finishing the 3 cycles of inductive treatment, patients would decide whether to continue ASCT consolidation therapy or another cycle of consolidation. R maintenance therapy was recommended to all patients. The primary endpoint of this study was the progression-free survival (PFS). Key secondary endpoints were complete response rate (CRR) and objective response rate (ORR) at the end of induction (EOI), overall survival (OS), minimal residual disease (MRD) status and the incidence of adverse events (AEs). Results: In this study, we recruited 55 patients with a median age of 53 years (range, 37~65 years) and a male/female ratio of 2.2:1. Totally, 61.8% of patients were of at least one risk factor (HR) according to risk stratification, pathological classification and molecular genetic abnormalities. All patients received at least two cycles of alternate R-DA-EDOCH/R-DHAP immunochemotherapy, and one patient dropped out due to the poor response. The ORR of the alternate R-DA-EDOCH/R-DHAP inductive treatment was 98% (95% CI 90-100), and the CRR was 89% (95%CI 78-96). There was no significant difference in CRR and ORR among patients with different prognostic characteristics. Moreover, patients rapidly achieved MRD-negative and the rate of MRD-negative after 2 cycles of treatment, the end of the inductive therapy and the end of the overall treatment was 79.2%, 88.7% and 90.6%, respectively. The median follow-up of the cohort was 31 months (range 4-86). The 3-year PFS and OS rates were 66.3% and 85.4%, respectively. Sequential ASCT consolidation and/or R maintenance slightly increased CRR but significantly prolonged PFS of patients (3-year PFS rate: ASCT+R 90.4% versus R 64.6% versus without ASCT/R 33.3%, P<0.001). There was no significance difference in PFS and OS between patients with (n=34) and without (n=9) HR (PFS P=0.537, OS P=0.408). For patients receiving ASCT and R maintenance, OS was even longer in patients with HR (mOS not reach versus 47m, P=0.007). Univariate analysis showed that age≥60 years, blastoid/pleomorphic pathologic morphology and TP53 double-allele events were still the risk factors for PFS in this therapeutic pattern. However, high tumor proliferative activity (such as Ki-67≥30% and MYC abnormality) and specific genetic abnormalities (such as 17p deletion, 9p deletion, etc) showed no significant adverse prognostic influence. In particular, patients who achieved MRD-negative by treatment had longer PFS (P<0.001). The alternate R-DA-EDOCH/R-DHAP regimen was well tolerated, and no patients were discontinued due to intolerance of treatment-related AEs. AEs were mainly hematologic and at least one hematologic AEs occurred in 85.5% of the patients. Grade 3-4 leukopenia and neutropenia were found in 49.1% and 58.2% of the patients, respectively. Conclusion: The alternate R-DA-EDOCH/R-DHAP induction therapy showed great efficacy and tolerability in young patients with newly diagnosed MCL and could overcome the adverse prognostic impact of some high-risk factors. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Functionalized Macrophage Exosomes with Panobinostat and PPM1D-siRNA for Diffuse Intrinsic Pontine Gliomas Therapy.

Diffuse intrinsic pontine glioma (DIPG) is a rare and fatal pediatric brain tumor. Mutation of p53‐induced protein phosphatase 1 (PPM1D) in DIPG cells promotes tumor cell proliferation, and inhibition of PPM1D expression in DIPG cells with PPM1D mutation effectively reduces the proliferation activity of tumor cells. Panobinostat effectively kills DIPG tumor cells, but its systemic toxicity and low blood–brain barrier (BBB) permeability limits its application. In this paper, a nano drug delivery system based on functionalized macrophage exosomes with panobinostat and PPM1D‐siRNA for targeted therapy of DIPG with PPM1D mutation is prepared. The nano drug delivery system has higher drug delivery efficiency and better therapeutic effect than free drugs. In vivo and in vitro experimental results show that the nano drug delivery system can deliver panobinostat and siRNA across the BBB and achieve a targeted killing effect of DIPG tumor cells, resulting in the prolonged survival of orthotopic DIPG mice. This study provides new ideas for the delivery of small molecule drugs and gene drugs for DIPG therapy.

Effects of Exogenous ATP on Melanoma Growth and Tumor Metabolism in C57BL/6 Mice.

Altered energy metabolism (glucose, lipid, amino acid) is a hallmark of cancer growth that provides the theoretical basis for the development of metabolic therapies as cancer treatments. ATP is one of the major biochemical constituents of the tumor microenvironment. ATP promotes tumor progression or suppression depending on various factors, including concentration and tumor type. Here we evaluated the antitumor effect of extracellular ATP on melanoma and the potential underlying mechanisms. A subcutaneous tumor model in mice was used to investigate the antitumor effects of ATP. Major lymphocyte cell changes and intratumoral metabolic changes were assessed. Metabolomic analysis (1H nuclear magnetic resonance spectroscopy) was performed on tumor samples. We measured the activities of lactate dehydrogenase A (LDHA) and LDHB in the excised tumors and serum and found that ATP and its metabolites affected the proliferation of and LDHA activity in B16F10 cells, a murine melanoma cell line. In addition, treatment with ATP dose-dependently reduced tumor size in melanoma-bearing mice. Moreover, flow cytometry analysis demonstrated that the antitumor effect of ATP was not achieved through changes in T-cell or B-cell subsets. Metabolomics analysis revealed that ATP treatment simultaneously reduced multiple intratumoral metabolites related to energy metabolism as well as serum and tumor LDHA activities. Furthermore, both ATP and its metabolites significantly suppressed both tumor cell proliferation and LDHA activity in the melanoma cell line. Our results in vivo and in vitro indicate that exogenous ATP inhibits melanoma growth in association with altered intratumoral metabolism.

ET-7 Roles for hENT1 and dCK in gemcitabine sensitivity and malignancy of meningioma

Background: High-grade meningiomas are aggressive tumors with high morbidity and mortality rates that frequently recur even after surgery and adjuvant radiotherapy. However, limited information is currently available on the biology of these tumors, and no alternative adjuvant treatment options exist. Although we previously demonstrated that high-grade meningioma cells were highly sensitive to gemcitabine in vitro and in vivo, the underlying molecular mechanisms remain unknown. Methods: We examined the roles of hENT1 (human equilibrative nucleoside transporter 1) and dCK (deoxycytidine kinase) in the gemcitabine sensitivity and growth of meningioma cells in vitro. Tissue samples from meningiomas (26 WHO grade I and 21 WHO grade II/III meningiomas) were immunohistochemically analyzed for hENT1 and dCK as well as for Ki-67 as a marker of proliferative activity. Results: hENT1 and dCK, which play critical roles in the intracellular transport and activation of gemcitabine, respectively, were responsible for the high gemcitabine sensitivity of high-grade meningioma cells and were strongly expressed in high-grade meningiomas. hENT1 expression was required for the proliferation and survival of high-grade meningioma cells and dCK expression. Furthermore, high hENT1 and dCK expression levels correlated with stronger tumor cell proliferative activity and shorter survival in meningioma patients. Conclusions: The present results suggest that hENT1 is a key molecular factor influencing the growth capacity and gemcitabine sensitivity of meningioma cells and also that hENT1, together with dCK, may be a viable prognostic marker for meningioma patients as well as a predictive marker of their responses to gemcitabine.

Albuca Bracteate Polysaccharides Synergistically Enhance the Anti-Tumor Efficacy of 5-Fluorouracil Against Colorectal Cancer by Modulating β-Catenin Signaling and Intestinal Flora

The first-line treatment for colorectal cancer (CRC) is 5-fluorouracil (5-FU). However, the efficacy of this treatment is sometimes limited owing to chemoresistance as well as treatment-associated intestinal mucositis and other adverse events. Growing evidence suggests that certain phytochemicals have therapeutic and cancer-preventing properties. Further, the synergistic interactions between many such plant-derived products and chemotherapeutic drugs have been linked to improved therapeutic efficacy. Polysaccharides extracted from Albuca bracteata (Thunb.) J.C.Manning and Goldblatt (ABP) have been reported to exhibit anti-oxidant, anti-inflammatory, and anti-tumor properties. In this study, murine CRC cells (CT26) and a murine model of CRC were used to examine the anti-tumor properties of ABP and explore the mechanism underlying the synergistic interactions between ABP and 5-FU. Our results revealed that ABP could inhibit tumor cell proliferation, invasion, and migratory activity in vitro and inhibited tumor progression in vivo by suppressing β-catenin signaling. Additionally, treatment with a combination of ABP and 5-FU resulted in better outcomes than treatment with either agent alone. Moreover, this combination therapy resulted in the specific enrichment of Ruminococcus, Anaerostipes, and Oscillospira in the intestinal microbiota and increased fecal short-chain fatty acid (SCFA) levels (acetic acid, propionic acid, and butyric acid). The improvement in the intestinal microbiota and the increase in beneficial SCFAs contributed to enhanced therapeutic outcomes and reduced the adverse effects of 5-FU. Together, these data suggest that ABP exhibits anti-neoplastic activity and can effectively enhance the efficacy of 5-FU in CRC treatment. Therefore, further research on the application of ABP in the development of novel anti-tumor drugs and adjuvant compounds is warranted and could improve the outcomes of CRC patients.

Combined Application of Pan-AKT Inhibitor MK-2206 and BCL-2 Antagonist Venetoclax in B-Cell Precursor Acute Lymphoblastic Leukemia.

Aberrant PI3K/AKT signaling is a hallmark of acute B-lymphoblastic leukemia (B-ALL) resulting in increased tumor cell proliferation and apoptosis deficiency. While previous AKT inhibitors struggled with selectivity, MK-2206 promises meticulous pan-AKT targeting with proven anti-tumor activity. We herein, characterize the effect of MK-2206 on B-ALL cell lines and primary samples and investigate potential synergistic effects with BCL-2 inhibitor venetoclax to overcome limitations in apoptosis induction. MK-2206 incubation reduced AKT phosphorylation and influenced downstream signaling activity. Interestingly, after MK-2206 mono application tumor cell proliferation and metabolic activity were diminished significantly independently of basal AKT phosphorylation. Morphological changes but no induction of apoptosis was detected in the observed cell lines. In contrast, primary samples cultivated in a protective microenvironment showed a decrease in vital cells. Combined MK-2206 and venetoclax incubation resulted in partially synergistic anti-proliferative effects independently of application sequence in SEM and RS4;11 cell lines. Venetoclax-mediated apoptosis was not intensified by addition of MK-2206. Functional assessment of BCL-2 inhibition via Bax translocation assay revealed slightly increased pro-apoptotic signaling after combined MK-2206 and venetoclax incubation. In summary, we demonstrate that the pan-AKT inhibitor MK-2206 potently blocks B-ALL cell proliferation and for the first time characterize the synergistic effect of combined MK-2206 and venetoclax treatment in B-ALL.

ULTRASTRUCTURAL AND IMMUNOHISTOCHEMICAL STUDIES OF REACTIVE AND ADAPTIVE TRANSFORMATIONS OF THE GREATER OMENTUM AT TUMOR LESIONS OF THE OVARIES

The greater omentum is an organ in which metastases of malignant neoplasms of the female reproductive system are often formed. The issues of adaptive and reactive transformations of the greater omentum under conditions of a tumor process in the organs of the female reproductive system have been insufficiently studied. The aim of the study was to elucidate morphological and functional ultrastructural and immune-histochemical changes in the greater omentum in women with ovarian tumor lesions. Using the methods of light and electron microscopy, immunohistochemistry and morphometry, the ovaries of 48 women were examined, who were diagnosed with poorly differentiated serous-papillary adenocarcinoma stage II (20 patients, group 1), stage III (28 patients, group 2). In stage III patients, numerous tumor metastases were detected in the greater omentum. The results of the study showed that in the patients of the 2nd group, in the peri-tumor areas of the greater omentum, the vessels of the microvasculature were dilated, in the connective tissue, moderate edema and leukocyte infiltration were observed. Immuno-histochemical markers showed an increase in the number of immunocompetent cells, mainly T-lymphocytes in the tumor and in the peri-tumor areas. In the tumor and in the surrounding areas of the greater omentum, the process of neoplasm of blood capillaries was noted, differences in the density of collagen fibrils in different parts of the serous membrane were revealed, and the presence of numerous holes was shown, the sizes of which were reduced in areas with metastases. The results obtained show the adaptive and reparative capabilities of the structures of the greater omentum during the tumor process in the organs of the female reproductive system, indicate the important role of the greater omentum in providing protective reactions in the abdominal cavity. In response to the invasion of tumor cells, the processes of angiogenesis are activated in the greater omentum, and the number of immunocompetent cells increases in neo-lymphogenesis. The materials of the study indicate a significant plasticity and reactivity of the greater omentum under conditions of a tumor process in the body. However, the presence of a pronounced proliferative activity of tumor cells in tumor metastases indicates that these adaptive capacities are insufficient to resist tumor invasion into the greater omentum.

Shikonin Reduces Growth of Docetaxel-Resistant Prostate Cancer Cells Mainly through Necroptosis.

Simple SummaryProstate carcinoma (PCa) is the most common tumor in men with an increasing age-associated risk. Several therapy strategies, one of which is docetaxel (DX) chemotherapy, have been established. However, due to the development of therapy resistance, in which chemotherapy no longer effectively combats the cancer, advanced, metastasized PCa with a poor prognosis may become manifested and therapy inevitably fails. Thus, new treatment options are urgently needed. Shikonin (SHI), from Traditional Chinese Medicine, has revealed promising antitumor activity in several tumor entities. In the current study, the impact of SHI on four therapy-sensitive and four respective DX-resistant PCa cell lines was determined. SHI induced growth inhibition mainly by necroptosis, a type of cell death, in all the tested therapy-sensitive, but more importantly, DX-resistant PCa cell lines. Corresponding molecular alterations contributing to growth inhibition after SHI exposure were found. SHI could, therefore, be a promising additive in treating advanced PCa.The prognosis for advanced prostate carcinoma (PCa) remains poor due to development of therapy resistance, and new treatment options are needed. Shikonin (SHI) from Traditional Chinese Medicine has induced antitumor effects in diverse tumor entities, but data related to PCa are scarce. Therefore, the parental (=sensitive) and docetaxel (DX)-resistant PCa cell lines, PC3, DU145, LNCaP, and 22Rv1 were exposed to SHI [0.1–1.5 μM], and tumor cell growth, proliferation, cell cycling, cell death (apoptosis, necrosis, and necroptosis), and metabolic activity were evaluated. Correspondingly, the expression of regulating proteins was assessed. Exposure to SHI time- and dose-dependently inhibited tumor cell growth and proliferation in parental and DX-resistant PCa cells, accompanied by cell cycle arrest in the G2/M or S phase and modulation of cell cycle regulating proteins. SHI induced apoptosis and more dominantly necroptosis in both parental and DX-resistant PCa cells. This was shown by enhanced pRIP1 and pRIP3 expression and returned growth if applying the necroptosis inhibitor necrostatin-1. No SHI-induced alteration in metabolic activity of the PCa cells was detected. The significant antitumor effects induced by SHI to parental and DX-resistant PCa cells make the addition of SHI to standard therapy a promising treatment strategy for patients with advanced PCa.

Tumor cell proliferation (Ki-67) expression and its prognostic significance in histological subtypes of lung adenocarcinoma

ObjectivesKi-67 is a key molecular marker to indicate the proliferative activity of tumor cells in lung cancer. However, Ki-67 expression and its prognostic significance in histological subtypes of lung adenocarcinoma (LUAD) remain unclear. Materials and MethodsWe retrospectively analyzed 1028 invasive LUAD patients who underwent surgery treatment between January 2012 and April 2020 in our department. Associations between Ki-67 expression and histological subtypes of LUAD, as well as other clinicopathological characteristics, were evaluated. The prognostic role of Ki-67 in LUAD subtypes was further assessed using log-rank test and univariate/multivariate Cox proportional hazards regression analyses. ResultsKi-67 expression differed across LUAD histological subtypes. The solid-predominant adenocarcinoma (SPA, 46.31 ± 24.72) had the highest expression level of Ki-67, followed by micropapillary (MPA, 31.71 ± 18.14), papillary (PPA, 22.09 ± 19.61), acinar (APA, 19.73 ± 18.71) and lepidic-predominant adenocarcinoma (LPA, 9.86 ± 8.10, P < 0.001). Tumors with solid or micropapillary components also had a higher Ki-67 expression than those without solid or micropapillary components. Besides, males, smokers, larger tumor size, lymph node metastasis and EGFR wild type were correlated with elevated Ki-67 expression. Univariate analysis indicated that increased Ki-67 expression and MPA/SPA subtypes were significantly associated with a poorer prognosis. Notably, the survival differences between LUAD subtypes vanished after adjusting for tumor size and Ki-67 expression in multivariate analysis, while Ki-67 was an independent prognostic factor of LUAD. Patients with MPA/SPA had non-inferior overall and disease-free survival than LPA/APA/PPA patients with a Ki-67 expression comparable to MPA/SPA subjects. ConclusionKi-67 expression varied considerably according to the predominant histological subtypes of LUAD. Ki-67 expression level and tumor size contributed to the survival differences between LUAD histological subtypes.

Roles for hENT1 and dCK in gemcitabine sensitivity and malignancy of meningioma.

BackgroundHigh-grade meningiomas are aggressive tumors with high morbidity and mortality rates that frequently recur even after surgery and adjuvant radiotherapy. However, limited information is currently available on the biology of these tumors, and no alternative adjuvant treatment options exist. Although we previously demonstrated that high-grade meningioma cells were highly sensitive to gemcitabine in vitro and in vivo, the underlying molecular mechanisms remain unknown.MethodsWe examined the roles of hENT1 (human equilibrative nucleoside transporter 1) and dCK (deoxycytidine kinase) in the gemcitabine sensitivity and growth of meningioma cells in vitro. Tissue samples from meningiomas (26 WHO grade I and 21 WHO grade II/III meningiomas) were immunohistochemically analyzed for hENT1 and dCK as well as for Ki-67 as a marker of proliferative activity.ResultshENT1 and dCK, which play critical roles in the intracellular transport and activation of gemcitabine, respectively, were responsible for the high gemcitabine sensitivity of high-grade meningioma cells and were strongly expressed in high-grade meningiomas. hENT1 expression was required for the proliferation and survival of high-grade meningioma cells and dCK expression. Furthermore, high hENT1 and dCK expression levels correlated with stronger tumor cell proliferative activity and shorter survival in meningioma patients.ConclusionsThe present results suggest that hENT1 is a key molecular factor influencing the growth capacity and gemcitabine sensitivity of meningioma cells and also that hENT1, together with dCK, may be a viable prognostic marker for meningioma patients as well as a predictive marker of their responses to gemcitabine.

Impact of the acidic environment on gene expression and functional parameters of tumors in vitro and in vivo

BackgroundThe low extracellular pH (pHe) of tumors resulting from glycolytic metabolism is a stress factor for the cells independent from concomitant hypoxia. The aim of the study was to analyze the impact of acidic pHe on gene expression on mRNA and protein level in two experimental tumor lines in vitro and in vivo and were compared to hypoxic conditions as well as combined acidosis+hypoxia.MethodsGene expression was analyzed in AT1 prostate and Walker-256 mammary carcinoma of the rat by Next Generation Sequencing (NGS), qPCR and Western blot. In addition, the impact of acidosis on tumor cell migration, adhesion, proliferation, cell death and mitochondrial activity was analyzed.ResultsNGS analyses revealed that 147 genes were uniformly regulated in both cell lines (in vitro) and 79 genes in both experimental tumors after 24 h at low pH. A subset of 25 genes was re-evaluated by qPCR and Western blot. Low pH consistently upregulated Aox1, Gls2, Gstp1, Ikbke, Per3, Pink1, Tlr5, Txnip, Ypel3 or downregulated Acat2, Brip1, Clspn, Dnajc25, Ercc6l, Mmd, Rif1, Zmpste24 whereas hypoxia alone led to a downregulation of most of the genes. Direct incubation at low pH reduced tumor cell adhesion whereas acidic pre-incubation increased the adhesive potential. In both tumor lines acidosis induced a G1-arrest (in vivo) of the cell cycle and a strong increase in necrotic cell death (but not in apoptosis). The mitochondrial O2 consumption increased gradually with decreasing pH.ConclusionsThese data show that acidic pHe in tumors plays an important role for gene expression independently from hypoxia. In parallel, acidosis modulates functional properties of tumors relevant for their malignant potential and which might be the result of pH-dependent gene expression.

Epithelial-mesenchymal transition in main types of gastric carcinoma

Introduction. The rapid development of basic science enabled us to significantly expand our understanding of various intercellular interactions. Epithelial-mesenchymal transition (EMT) is known to play a key role in certain tissue formation in the embryonic period. However, recent data show that EMT can also be observed in some pathological conditions, in particular, in various neoplasm development. This suggests that there are a number of alternative and fundamentally new mechanisms for the tumor formation and progression. Thus, EMT, which occurs in carcinomas, increases the invasiveness, immunoresistance, immunity to therapy, and the metastatic potential. Knowledge of EMT features and their timely recognition in morphological tumor diagnosis is of great predictive importance for patients. The aim of the research was to study the morphologi-cal features of epithelial-mesenchymal transition in the main types of gastric cancer. Materials and methods. We studied specimens of gastric carcinomas (N=64) including 31 cases of diffuse type, 19 cases of intestinal type, and 14 cases of mixed type. Results. All cases of the diffuse carcinoma group showed spread EMT features, which appeared already in the mucosa and completed with positive vimentin expression in 93.5% of cases. The malignant cell prolifera-tive activity was low; however, in 29% of cases we detected areas of moderate or even high activity. In the intestinal type gastric cancer, EMT developed as a result of tumor progression, it arose more often in the deeper layers and was incomplete and focal. As a rule, the proliferative activity of tumor cells was high and moderate. Vascular invasion occurred more often in diffuse type (90.3%), less often in mixed type (71.4%), and even less often in the intestine type (55.8%) gastric carcinoma. Conclusion. The variety of morphological features of EMT, its frequency, prevalence, completeness, and sequence in the development of various types of gastric cancer determines the features of their clinical manifestation and influences their further management. Keywords: gastric cancer, diagnosis, histological main types, EMT, morphopathology

Diffusion Kurtosis Imaging Reflects GFAP, TopoIIα, and MGMT Expression in Astrocytomas.

Preliminary study of magnetic resonance (MR) diffusion kurtosis imaging (DKI) assessing the pathological glial fibrillary acidic protein (GFAP), TopoIIα, and O 6-methylguanine-DNA methyltransferase (MGMT) expression in astrocytomas. This study was approved by the local ethics committee, and informed consent was obtained from all participants. Sixty-six cases with pathologically proven astrocytomas were enrolled in this study; of which, 34 were high grade and remaining 32 were low grade. They patients underwent conventional MRI head scan, DKI scan, and enhanced scan under the same conditions. Fractional anisotropy (FA) and mean kurtosis (MK) calculated from DKI, as well as GFAP, TopoIIα, and MGMT expression level were compared prospectively between high and low-grade astrocytomas. Spearman rank correlation analysis was used for comparing values of DKI and GFAP, TopoIIα, and MGMT expression level in the two groups. The MK values were significantly higher in high-grade astrocytomas than those in low-grade astrocytomas (P < 0.05); FA values demonstrated no significant difference between the two groups (P = 0.331). GFAP expression level was significantly lower in high-grade astrocytomas than in low-grade astrocytomas (P < 0.05). Topo-IIα expression level were significantly higher in high-grade astrocytomas than in low-grade astrocytomas (P < 0.05). There was no significant difference in MGMT expression level between the two groups (P = 0.679). MK values were negatively correlated with the expression of GFAP (r = -0.836; P = 0.03), however, they were positively correlated with the expression of Topo-IIα (r = 0.896; P = 0.01). FA values were not correlated with the expression of GFAP (r = 0.366; P = 0.05), Topo-IIα (r = -0.562; P = 0.05), and MGMT (r = -0.153; P = 0.10). MK, the DKI parameter values of astrocytomas, was significantly correlated to the expression of GFAP and TopoIIα. To a certain extent, applying DKI may provide the biological behavior of tumor cell differentiation, proliferation activity, invasion and metastasis, and can guide individual treatment.

Immunohistochemical study of P53 protein expression in different prostate cancer Gleason grading groups

Prostate cancer (PC) remains an urgent public health problem, especially in developed countries. The use of immunohistochemical research methods in addition to the morphological classification of prostate adenocarcinomas allows a more accurate diagnosis and prognosis of the disease. The aim of the study is to identify isoforms of P53 using clones of mouse antibodies (D-07 and Y5; Epitomics, USA) in prostate cancer with different proliferative activity and the degree of malignancy. Materials and Methods: The work included surgical material for prostate resection and prostatectomy, as well as biopsy specimens (56 cases in total). An immunohistochemical study was carried out with the Ki-67 marker, as well as with mouse monoclonal antibodies (D-07 and Y5) to the P53 protein, interacting with its wild and mutant isoforms. The significance of the difference in the samples was determined using the Mann-Whitney U-test, correlation relationships were determined using the Spearman coefficient. Results: Expression of P53 upon interaction with antibodies D-07 and Y5 was determined in 56.3% and 39.6%, respectively. A statistically significant direct correlation was found between the severity of P53 expression when interacting with Y5 antibodies and the degree of tumor differentiation (rs = 0.567, p 0.05), as well as between the expression level of this protein and tumor proliferative activity (rs = 0.698, p 0.05). Conclusion: Antibodies of clone D-07, interacting with both wild and mutant isoforms of P53 protein, show positive expression in adenocarcinomas of all degrees. Expression of the mutant P53 protein is most pronounced in low-differentiated carcinomas and correlates with high proliferative activity of tumor cells, which may be associated with a loss in the induction of P53-dependent apoptosis.