Therapy-induced senescent cancer cells exhibit complement activation and increased complement regulatory protein expression.

Therapy-induced senescence (TIS) is a primary response to chemotherapy, contributing to untoward treatment outcomes such as evasion of immunosurveillance. Despite the established role of the complement system in the immune response to cancer, the role of complement in mediating the immune response against senescent tumor cells remains poorly understood. To explore this relationship, we exposed lung adenocarcinoma (A549), breast adenocarcinoma (MCF7) and pancreatic carcinoma (Panc-1) cell lines to sublethal doses of either etoposide or doxorubicin to trigger TIS. Identification of TIS was based on morphological changes, upregulation of the senescence-associated ß-galactosidase, p21Cip1 induction and lamin B1 downregulation. Using immunofluorescence microscopy, quantitative PCR, ELISA of conditioned media and in silico analysis, we investigated complement activation, complement protein expression, C3 levels in the conditioned media of senescent cells and secreted complement proteins as part of the senescence-associated secretory phenotype (SASP), respectively. In cell lines undergoing TIS, complement-related changes included (i) activation of the terminal pathway, evidenced by the deposition of C5b-9 on senescent cells; (ii) an increase in the expression of CD59 and complement factor H and (iii) in A549 cells, an elevation in the expression of C3 with its secretion into the medium. In addition, increased C3 expression was observed in breast cancer samples expressing TIS hallmarks following exposure to neoadjuvant chemotherapy. In conclusion, TIS led to the activation of complement, upregulation of complement regulatory proteins and increased C3 expression. Complement appears to play a role in shaping the cancer microenvironment upon senescence induction.

Association of Human Papilloma Virus, Cytomegalovirus, and Epstein-Barr Virus with Breast Cancer in Jordanian Women.

Background and Objectives: The investigation of oncogenic viruses and their potential association with breast cancer (BC) remains an intriguing area of study. The current work aims to assess evidence of three specific viruses, human papillomavirus (HPV), cytomegalovirus (CMV), and Epstein-Barr virus (EBV) in BC samples and to explore their relationship with relevant clinicopathological variables. Materials and Methods: The analysis involved BC samples from 110 Jordanian female patients diagnosed with BC and breast tissue samples from 30 control patients with no evidence of breast malignancy, investigated using real-time PCR. The findings were then correlated with various clinico-pathological characteristics of BC. Results: HPV was detected in 27 (24.5%), CMV in 15 (13.6%), and EBV in 18 (16.4%) BC patients. None of the control samples was positive for HPV or CMV while EBV was detected in only one (3.3%) sample. While (HPV/EBV), (HPV/CMV), and (EBV/CMV) co-infections were reported in 1.8%, 2.7%, and 5.5%, respectively, coinfection with the three viruses (HPV/CMV/EBV) was not reported in our cohort. A statistically significant association was observed between HPV status and age (p = 0.047), and between clinical stage and CMV infection (p = 0.015). Conclusions: Our findings indicate the presence or co-presence of HPV, CMV, and EBV in the BC subpopulation, suggesting a potential role in its development and/or progression. Further investigation is required to elucidate the underlying mechanisms that account for the exact role of oncoviruses in breast carcinogenesis.

Angiotensin II type 1 receptor blockade attenuates gefitinib-induced cardiac hypertrophy via adjusting angiotensin II-mediated oxidative stress and JNK/P38 MAPK pathway in a rat model.

Gefitinib is a tyrosine kinase inhibitor (TKI) of the epidermal growth factor receptor (EGFR), used for the treatment of advanced or metastatic non-small cell lung cancer. Recently, studies proved that Gefitinib-induced cardiotoxicity through induction of oxidative stress leads to cardiac hypertrophy. The current study was conducted to understand the mechanisms underlying gefitinib-induced cardiac hypertrophy through studying the roles of angiotensin II (AngII), oxidative stress, and mitogen-activated protein kinase (MAPK) pathway. Male Wistar albino rats were treated with valsartan, gefitinib, or both for four weeks. Blood samples were collected for AngII and cardiac markers measurement, and hearts were harvested for histological study and biochemical analysis. Gefitinib caused histological changes in the cardiac tissues and increased levels of cardiac hypertrophy markers, AngII and its receptors. Blocking of AngII type 1 receptor (AT1R) via valsartan protected hearts and normalized cardiac markers, AngII levels, and the expression of its receptors during gefitinib treatment. valsartan attenuated gefitinib-induced NADPH oxidase and oxidative stress leading to down-regulation of JNK/p38-MAPK pathway. Collectively, AT1R blockade adjusted AngII-induced NADPH oxidase and JNK/p38-MAPK leading to attenuation of gefitinib-induced cardiac hypertrophy. This study found a pivotal role of AngII/AT1R signaling in gefitinib-induced cardiac hypertrophy, which may provide novel approaches in the management of EGFRIs-induced cardiotoxicity.

Protective effect of valsartan against doxorubicin-induced cardiotoxicity: Histopathology and metabolomics in vivo study.

Doxorubicin (DOX) treatment has been associated with cardiotoxicity. Therefore, it is crucial to search for a therapeutic that can effectively mitigate DOX-induced cardiotoxicity. This study was conducted to investigate the protective effects of valsartan (VAL) against DOX-induced cardiotoxicity. Sprague-Dawley rats were divided into four treatment groups: Group I: Control, Group II: VAL (30 mg/kg, ip), Group III: DOX (15 mg/kg, ip), and Group IV: VAL + DOX (30 + 15 mg/kg, ip). All groups were treated every other day for 14 days. Blood was isolated for biochemical and metabolomics studies, and sections of the heart were also analyzed for histopathological and immunohistochemical alterations to detect changes in P53, BAX, BCL-2, and P62 expression. The combination of VAL + DOX resulted in a marked decrease in cardiac biomarker enzymes (aminotransferase and creatine phosphokinase) compared to DOX monotherapy. In addition, the histopathological examination of the VAL + DOX combination revealed a low percentage of fibrosis and inflammation. Immunohistochemical expression of p53 and BAX was significantly reduced, whereas BCL-2 expression was significantly increased in the VAL + DOX treatment group compared to DOX monotherapy. Also, the combination of VAL + DOX reverses the negative effect of DOX on nuclear p62 expression. Analysis of serum metabolites showed that DOX monotherapy reduced the number of several amino acids, whereas the combination of VAL + DOX restored these metabolic pathways. This study revealed the potential cardioprotective effect of VAL, which may provide novel and promising approaches for managing cardiotoxicity induced by DOX.

EGFR Inhibitor Gefitinib Induces Cardiotoxicity through the Modulation of Cardiac PTEN/Akt/FoxO3a Pathway and Reactive Metabolites Formation: In Vivo and in Vitro Rat Studies.

Gefitinib (GEF) is a selective inhibitor of the epidermal growth factor receptor (EGFR) used to treat non-small cell lung cancer. Yet, few cases of cardiotoxicity have been reported. However, the role of the PTEN/Akt/FoxO3a pathway, which mediates GEF anticancer activity, in GEF cardiotoxicity remains unclear. For this purpose, in vitro H9c2 cells and in vivo rat cardiomyocytes were utilized as study models. Treatment of H9c2 cells and Sprague-Dawley rats with GEF significantly induced the expression of hypertrophic and apoptotic markers at mRNA and protein levels with an increased plasma level of troponin. This was accompanied by induction of autophagy and mitochondrial dysfunction in H9c2 cells. Inhibition of cardiac EGFR activity and Akt cellular content of in vitro and in vivo rat cardiomyocytes by GEF increased PTEN and FoxO3a gene expression and cellular content. Importantly, treatment of H9c2 cells with PI3K/Akt inhibitor increased PTEN and FoxO3a mRNA expression associated with potentiation of GEF cardiotoxicity. In addition, by using LC-MS/MS, we showed that GEF is metabolized in the rat heart microsomes into one cyanide- and two methoxylamine-adduct reactive metabolites, where their formation was entirely blocked by CYP1A1 inhibitor, α-naphthoflavone. The current study concludes that GEF induces cardiotoxicity through modulating the expression and function of the cardiac PTEN/AKT/FoxO3a pathway and the formation of CYP1A1-mediated reactive metabolites.

In vivo Assessment of Combined Effects of Glibenclamide and Losartan in Diabetic Rats.

OBJECTIVE: Diabetic complications involve multiple pathological pathways, including hyperglycemia-induced oxidative stress and inflammation. Combination therapy is usually employed to improve treatment outcomes and to lower potential adverse effects. In this study, we evaluated the effects of antidiabetic and antihypertensive agents, glibenclamide (GLI) and losartan (LT), on diabetes mellitus (DM)-associated metabolic changes in rats. MATERIALS AND METHODS: Streptozotocin-induced diabetic animals were orally treated with GLI 5 mg/kg and/or LT 25 mg/kg for 4 weeks. Blood glucose, insulin, aspartate aminotransferase, alanine aminotransferase, urinary creatinine, and urea levels were measured. Serum, liver, and kidney values of inflammatory markers, such as interleukin-1ß, tumor necrosis factor alpha, and interleukin-6 were assessed, along with lipid peroxidation products (e.g., thiobarbituric acid reactive substances), endogenous antioxidants (e.g., glutathione), as well as antioxidant enzyme activities (e.g., catalase, superoxide dismutase, and glutathione peroxidase). Finally, histological changes in liver and kidney tissues were evaluated. RESULTS: DM markedly induced systemic, hepatic, and renal inflammation and lowered antioxidant defense mechanisms. Treatment of diabetic rats with either GLI or LT significantly improved liver and kidney functions and histological structure. Moreover, both medications reduced signs of oxidative stress and inflammation in blood, liver, and kidney samples. Combining GLI and LT showed similar protective potential against systemic, hepatic, and renal oxidative stress and inflammation. CONCLUSION: Adding LT to GLI therapy revealed prospective antioxidant and anti-inflammatory action, while no synergistic or additive effects were observed.

Protective effect of rutin supplementation against cisplatin-induced Nephrotoxicity in rats.

BACKGROUND: Cisplatin (CP) is commonly used in the treatment of different types of cancer but nephrotoxicity has been a major limiting factor. Therefore, the present study aimed to study the possible protective effect of rutin against nephrotoxicity induced by cisplatin in rats. METHODS: Forty male Wistar albino rats were randomly divided into 4 groups. Rats of group 1 control group intraperitoneal (i.p.) received 2.5 ml/kg, group 2 CP group received single dose 5 mg/kg cisplatin i.p. group 3 rutin group orally received 30 mg/kg rutin group 4 (CP plus rutin) received CP and rutin as in group 2 and 3. Kidneys were harvested for histopathology and for the study the gene expression of c-Jun N-terminal kinases (JNK), Mitogen-activated protein kinase 4 (MKK4), MKK7, P38 mitogen-activated protein kinases (P38), tumor necrosis factors alpha (TNF-α), TNF Receptor-Associated Factor 2 (TRAF2), and interleukin-1 alpha (IL-1-α). RESULTS: The cisplatin single dose administration to rats induced nephrotoxicity associated with a significant increase in blood urea nitrogen (BUN) and serum creatinine and significantly increase Malondialdehyde (MDA) in kidney tissues by 230 ± 5.5 nmol/g compared to control group. The animal treated with cisplatin showed a significant increase in the expression levels of the IL-1α (260%), TRFA2 (491%), P38 (410%), MKK4 (263%), MKK7 (412%), JNK (680%) and TNF-α (300%) genes compared to control group. Additionally, histopathological examination showed that cisplatin-induced interstitial congestion, focal mononuclear cell inflammatory, cell infiltrate, acute tubular injury with reactive atypia and apoptotic cells. Rutin administration attenuated cisplatin-induced alteration in gene expression and structural and functional changes in the kidney. Additionally, histopathological examination of kidney tissues confirmed gene expression data. CONCLUSION: The present study suggested that the anti-oxidant and anti-inflammatory effect of rutin may prevent CP-induced nephrotoxicity via decreasing the oxidative stress, inhibiting the interconnected ROS/JNK/TNF/P38 MAPK signaling pathways, and repairing the histopathological changes against cisplatin administration.

Combined antiproliferative effects of the aminoalkylindole WIN55,212-2 and radiation in breast cancer cells.

The potential antitumor activity of cannabinoid receptor agonists, such as the aminoalklylindole WIN55,212-2 (WIN2), has been studied extensively, but their potential interaction with conventional cancer therapies, such as radiation, remains unknown. In the present work, the influence of WIN2 on the antiproliferative activity of radiation in human (MCF-7 and MDA-MB231) and murine (4T1) breast cancer cells was investigated. The antiproliferative effects produced by combination of WIN2 and radiation were more effective than either agent alone. The stereoisomer of WIN2, WIN55,212-3 (WIN3), failed to inhibit growth or potentiate the growth-inhibitory effects of radiation, indicative of stereospecificity. Two other aminoalkylindoles, pravadoline and JWH-015 [(2-methyl-1-propyl-1H-indol-3-yl)-1-naphthalenyl-methanone], also enhanced the antiproliferative effects of radiation, but other synthetic cannabinoids (i.e., nabilone, CP55,940 [(+)-rel-5-(1,1-dimethylheptyl)-2-[(1R,2R,5R)-5-hydroxy-2-(3-hydroxypropyl)cyclohexyl]-phenol], and methanandamide) or phytocannabinoids [i.e., Δ9-tetrahydrocannabinol (THC) and cannabidiol] did not. The combination treatment of WIN2 + radiation promoted both autophagy and senescence but not apoptosis or necrosis. WIN2 also failed to alter radiation-induced DNA damage or the apparent rate of DNA repair. Although the antiproliferative actions of WIN2 were mediated through noncannabinoid receptor-mediated pathways, the observation that WIN2 interfered with growth stimulation by sphingosine-1-phosphate (S1P) implicates the potential involvement of S1P/ceramide signaling pathways. In addition to demonstrating that aminoalkylindole compounds could potentially augment the effectiveness of radiation treatment in breast cancer, the present study suggests that THC and nabilone are unlikely to interfere with the effectiveness of radiation therapy, which is of particular relevance to patients using cannabinoid-based drugs to ameliorate the toxicity of cancer therapies.

Cisplatin Provokes Peripheral Nociception and Neuronal Features of Therapy-Induced Senescence and Calcium Dysregulation in Rats.

Therapy-Induced Senescence (TIS) is a form of senescence that is typically described in malignant cells in response to the exposure of cancer chemotherapy or radiation but can also be precipitated in non-malignant cells. TIS has been shown to contribute to the development of several cancer therapy-related adverse effects; however, evidence on its role in mediating chemotherapy-induced neurotoxicity, such as Chemotherapy-induced Peripheral Neuropathy (CIPN), is limited. We here show that cisplatin treatment over two cycles (cumulative dose of 23 mg/kg) provoked mechanical allodynia and thermal hyperalgesia in Sprague-Dawley rats. Isolation of dorsal root ganglia (DRG) from the cisplatin-treated rats demonstrated robust SA-ß-gal upregulation at both day 8 (after the first cycle) and day 18 (after the second cycle), decreased lmnb1 expression, increased expression of cdkn1a and cdkn2a, and of several factors of the Senescence-associated Secretory Phenotype (SASP) (Il6, Il1b, and mmp9). Moreover, single-cell calcium imaging of cultured DRGs revealed a significant increase in terms of the magnitude of KCl-evoked calcium responses in cisplatin-treated rats compared to vehicle-treated rats. No significant change was observed in terms of the magnitude of capsaicin-evoked calcium responses in cisplatin-treated rats compared to vehicle-treated rats but with decreased area under the curve of the responses in cisplatin-treated rats. Further evidence to support the contribution of TIS to therapy adverse effects is required but should encourage the use of senescence-modulating agents (senotherapeutics) as novel palliative approaches to mitigate chemotherapy-induced neurotoxicity.

Characterization of BCL-X L , MCL-1, and BAX Protein Expression in Response to Neoadjuvant Chemotherapy in Breast Cancer.

The use of chemotherapy has improved the overall treatment of breast cancer, which is frequently administered in the form of neoadjuvant chemotherapy (NAC). Apoptosis is an established cell stress response to NAC in preclinical models; however, there is limited understanding of its role in clinical cancer, specifically, its contribution to favorable pathologic responses in breast cancer therapy. Here, we aimed to characterize the change in protein expression of 3 apoptosis-associated biomarkers, namely, BCL-X L , MCL-1, and BAX in breast cancer in response to NAC. For this, we utilized a set of 68 matched invasive breast cancer FFPE samples that were collected before (pre) and after (post) the exposure to NAC therapy that were characterized by incomplete pathologic response. Immunohistochemistry (IHC) analysis suggested that most of the samples show a decrease in the protein expression of all 3 markers following exposure to NAC as 90%, 69%, and 76% of the matched samples exhibited a decrease in expression for BCL-X L , MCL-1, and BAX, respectively. The median H-score of BCL-X L post-NAC was 150/300 compared with 225/300 pre-NAC ( P value <0.0001). The median H-score of MCL-1 declined from 200 pre-NAC to 160 post-NAC ( P value <0.0001). The median H-score of BAX protein expression decreased from 260 pre-NAC to 190 post-NAC ( P value <0.0001). There was no statistically significant association between the expression of these markers and stage, grade, and hormone receptor profiling (luminal status). Collectively, our data indicate that the expression of apoptosis regulatory proteins changes following exposure to NAC in breast cancer tissue, developing a partial pathologic response.

Hydroxychloroquine ameliorates dasatinib-induced liver injury via decrease in hepatic lymphocytes infiltration.

Dasatinib is an effective treatment for chronic myeloid leukemia. However, cases of idiosyncratic hepatotoxicity were reported. This study was conducted to investigate the chemopreventive effects of hydroxychloroquine against dasatinib-induced hepatotoxicity. Balb/c mice were randomly assigned into four groups; vehicle control (5% DMSO, i.p., n = 6), dasatinib (50 mg/kg; i.p., n = 6), hydroxychloroquine (10 mg/kg, i.p., n = 6), and hydroxychloroquine + dasatinib (10 mg/kg + 50 mg/kg; i.p., n = 6). Treatments were given once every 2 days for 14 days. Serum and histopathological assessments of liver architecture and fibrosis were performed using H&E, Masson's trichrome, and reticulin staining. The infiltration of lymphocytes was assessed using immunohistochemistry. The gene expression of antioxidant enzymes (CAT, SOD-2, GPX-1) was assessed using real-time quantitative PCR. Dasatinib showed a significant increase in liver injury biomarkers (AST and ALT) with higher lymphocytes infiltration (as indicated by CD3+, CD4+, CD8+, and CD20+ immunohistochemistry). Hepatic tissue of Dasatinib group exhibited significant downregulation in the gene expression of antioxidant enzymes (CAT, SOD-2, and GPX-1) compared to the control group. However, the combination of hydroxychloroquine with dasatinib showed a slight increase in AST and ALT. Also, hydroxychloroquine + dasatinib treated mice showed a significant reduction in lymphocytes infiltration as compared to dasatinib. The results showed that dasatinib induces an immune response leading to an increase in lymphocytes infiltration which promotes hepatocyte destruction and persistent liver injury. The results also suggest that hydroxychloroquine ameliorates dasatinib-induced hepatotoxicity via reduction in hepatic infiltration of T and B immune cells.

A three-marker signature identifies senescence in human breast cancer exposed to neoadjuvant chemotherapy.

PURPOSE: Despite the beneficial effects of chemotherapy, therapy-induced senescence (TIS) manifests itself as an undesirable byproduct. Preclinical evidence suggests that tumor cells undergoing TIS can re-emerge as more aggressive divergents and contribute to recurrence, and thus, senolytics were proposed as adjuvant treatment to eliminate senescent tumor cells. However, the identification of TIS in clinical samples is essential for the optimal use of senolytics in cancer therapy. In this study, we aimed to detect and quantify TIS using matched breast cancer samples collected pre- and post-exposure to neoadjuvant chemotherapy (NAC). METHODS: Detection of TIS was based on the change in gene and protein expression levels of three senescence-associated markers (downregulation of Lamin B1 and Ki-67 and upregulation of p16INK4a). RESULTS: Our analysis revealed that 23 of 72 (31%) of tumors had a shift in the protein expression of the three markers after exposure to NAC suggestive of TIS. Gene expression sets of two independent NAC-treated breast cancer samples showed consistent changes in the expression levels of LMNB1, MKI67 and CDKN2A. CONCLUSIONS: Collectively, our study shows a more individualized approach to measure TIS hallmarks in matched breast cancer samples and provides an estimation of the extent of TIS in breast cancer clinically. Results from this work should be complemented with more comprehensive identification approaches of TIS in clinical samples in order to adopt a more careful implementation of senolytics in cancer treatment.

Aneugenic and clastogenic alterations in the DBA/IJ mouse model of rheumatoid arthritis treated with rituximab, an anti-CD20 antibody.

Rheumatoid arthritis (RA), an autoimmune disorder in which the immune system attacks healthy cells, is associated with elevated risk of lymphoma. Rituximab, a treatment for non-Hodgkin's lymphoma, has been approved as a treatment for RA. We studied the effects of rituximab on chromosomal stability in collagen-induced arthritis DBA/1J animal models. Micronucleus levels were increased in the mouse models, mainly due to chromosome loss, as detected by fluorescence in situ hybridization; rituximab-treated arthritic mice had significantly less micronucleus formation. Serum 8-hydroxydeoxyguanosine, a DNA oxidative stress marker, was increased in the mice models but reduced following rituximab administration.

NOXA expression is downregulated in human breast cancer undergoing incomplete pathological response and senescence after neoadjuvant chemotherapy.

Neoadjuvant chemotherapy (NAC) is a frequently utilized approach to treat locally advanced breast cancer, but, unfortunately, a subset of tumors fails to undergo complete pathological response. Apoptosis and therapy-induced senescence (TIS) are both cell stress mechanisms but their exact role in mediating the pathological response to NAC is not fully elucidated. We investigated the change in expression of PAMIP1, the gene encoding for the pro-apoptotic protein, NOXA, following NAC in two breast cancer gene datasets, and the change in NOXA protein expression in response to NAC in 55 matched patient samples (pre- and post-NAC). PAMIP1 expression significantly declined in post-NAC in the two sets, and in our cohort, 75% of the samples exhibited a downregulation in NOXA post-NAC. Matched samples that showed a decline in NOXA post-NAC were examined for TIS based on a signature of downregulated expression of Lamin-B1 and Ki-67 and increased p16INK4a, and the majority exhibited a decrease in Lamin B1 (66%) and Ki-67 (80%), and increased p16INK4a (49%). Since our cohort consisted of patients that did not develop complete pathological response, such findings have clinical implications on the role of TIS and NOXA downregulation in mediating suboptimal responses to the currently established NAC.

The Combination of Radiation with PARP Inhibition Enhances Senescence and Sensitivity to the Senolytic, Navitoclax, in Triple Negative Breast Tumor Cells.

Despite significant advances in the treatment of triple-negative breast cancer, this disease continues to pose a clinical challenge, with many patients ultimately suffering from relapse. Tumor cells that recover after entering into a state of senescence after chemotherapy or radiation have been shown to develop a more aggressive phenotype, and to contribute to disease recurrence. By combining the PARP inhibitor (PARPi), talazoparib, with radiation, senescence was enhanced in 4T1 and MDA-MB-231 triple-negative breast cancer cell lines (based on SA-ß-gal upregulation, increased expression of CDKN1A and the senescence-associated secretory phenotype (SASP) marker, IL6). Subsequent treatment of the radiation- and talazoparib-induced senescent 4T1 and MDA-MB231 cells with navitoclax (ABT-263) resulted in significant apoptotic cell death. In immunocompetent tumor-bearing mice, navitoclax exerted a modest growth inhibitory effect when used alone, but dramatically interfered with the recovery of 4T1-derived tumors induced into senescence with ionizing radiation and talazoparib. These findings support the potential utility of a senolytic strategy in combination with the radiotherapy/PARPi combination to mitigate the risk of disease recurrence in triple-negative breast cancer.

Treatment with the anti-CD20 monoclonal antibody rituximab mitigates gonadal disruptions in the collagen-induced arthritis in male DBA/1 J mouse model.

Rheumatoid arthritis (RA), which is driven by persistent activation of the immune system, primarily affects the joints. Several reports have estimated the risk of gonadal disruptions in arthritic patients, with potential attributable risk factors such as treatments with the disease-modifying antirheumatic drugs and the influence of the disease itself. The FDA approved rituximab, a therapy for non-Hodgkin's lymphoma, for management of RA in February 2006. However, the influence of repeated treatment with rituximab on gonadal function in RA has not been reported yet. Thus, the aim of the presents study is to evaluate whether repeated treatment with the clinically relevant dose of rituximab may change the gonadal disruptions in collagen-induced arthritis in male DBA/1 J mouse, a model of RA. Testicular disruptions, as determined by the sperm DNA strand breaks, spermatocyte chromosomal analysis and spermiogram examination have been conducted by the use of standard techniques. Additionally, we aimed to test whether the anti-rheumatic effect of rituximab also decreases the cellular oxidant-antioxidant imbalance in arthritic male DBA/1 J mice. Repeated treatment of naïve control DBA/1 J mice with rituximab did not exhibit any significant deleterious effects. Moreover, repeated administration of rituximab to the arthritic DBA/1 J mice suppressed disease severity and decreased testicular disruptions. Rituximab treatment also diminished gonadal oxidative stress, through decreasing reactive oxygen species generation and restoring the reduced glutathione level in arthritic DBA/1 J mice. In conclusion, rituximab is a safe therapeutic agent and can mitigate gonadal disruptions induced by arthritis, which insinuates the importance for arthritic patients especially at reproductive age.

In vitro and in vivo antitumor studies of potential anticancer agents of platinum(II) complexes of dicyclopentadiene and dithiocarbamates§.

Three platinum(II) complexes of dicyclopentadiene (DCP) and dithiocarbamates (DTCs), namely, [Pt(η4-DCP)(Me2DTC)]PF6 (1), [Pt(η4-DCP)(Et2DTC)]PF6 (2), and [Pt(η4-DCP)(Bz2DTC)]PF6 (3) [Me2DTC = dimethyldithiocarbamate, Et2DTC = diethyldithiocarbamate, and Bz2DTC = dibenzyldithiocarbamate] were prepared and characterized by elemental analysis, IR, 1H, and 13C Nuclear Magnetic Resonance spectroscopy. The spectroscopic data indicated the coordination of both DCP and DTC ligands to platinum(II). The solution chemistry of complex 1 revealed that the complexes are stable in both dimethyl sulfoxide (DMSO) and 1:1 mixture of DMSO:H2O. In vitro cytotoxicity of the complexes relative to cisplatin was tested using MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay, against CHL-1 (human melanoma cancer cells), MDA-MB-231 (breast cancer cells), A549 (lung cancer cells), and B16 (murine melanoma cancer cells). The antiproliferative effect of all three prepared complexes was found to be significantly higher than cisplatin. Furthermore, flow cytometric analysis of complex 1 showed that the complex induced apoptosis, oxidative stress, mitochondrial potential depolarization and cell cycle arrest in a concentration-dependent pattern in the CHL-1 cells. Confirmation of apoptosis via gene expression analysis demonstrated down-regulation of anti-apoptotic genes and up-regulation of pro-apoptotic genes in the CHL-1 cells. Wound-healing assays also lent support to the strong cytotoxicity of the complexes. In vivo studies showed a significant reduction of tumor volume at the end of the experiment. In addition, the drug did not change the weight of the mice. In conclusion, complex 1 inhibited cell proliferation in vitro and reduced tumor growth in vivo.

Inhibition of interleukin-2-inducible T-cell kinase causes reduction in imiquimod-induced psoriasiform inflammation through reduction of Th17 cells and enhancement of Treg cells in mice.

Psoriasis is a debilitating chronic skin disease with a worldwide prevalence. Its main features include well-marked silvery scales on the skin of hands and feet and back which arise due to hyperproliferation of keratinocytes and infiltration of immune cells in the skin. Multiple interactions exist between adaptive immune cells such as T cells and innate immune cells such as neutrophils and macrophages which are key players in the pathogenesis of psoriasis. Interleukin-2-inducible T-cell kinase (ITK) plays a key role in Th17 cell development through control of several transcription factors. ITK has been shown to control NFATc1, NFkB and STAT3 in CD4+ T cells. Effect of ITK inhibitor in imiquimod (IMQ)-induced psoriasiform inflammation remains to be explored. In the current examination, role of ITK signaling and its inhibition blockade were evaluated on NFATc1, NFkB and STAT3, IL-17A, TNF-α, IFN-γ, Foxp3, IL-10 in CD4+ T cells in IMQ model. Our data display that ITK signaling is involved in IMQ-induced psoriatic inflammation as paralleled by enhancement of p-ITK, NFATc1, p-NFkB and p-STAT3 in CD4+ T cells. It was associated with enhancement of Th17/Th1 cells and neutrophilic inflammation in the skin. Preventive treatment with ITK inhibitor led to a reduction in Th17/Th1 cells and enhancement of Treg cells. Overall, this study suggests that ITK signaling is an important modulator of transcription factor signaling in CD4+ T cells which is associated with Th17/Th1 cells and psoriasiform inflammation in mice. ITK signaling blockade could be a therapeutic target for the treatment of psoriatic inflammation.

Clearance of therapy-induced senescent tumor cells by the senolytic ABT-263 via interference with BCL-XL -BAX interaction.

Tumor cells undergo senescence in response to both conventional and targeted cancer therapies. The induction of senescence in response to cancer therapy can contribute to unfavorable patient outcomes, potentially including disease relapse. This possibiliy is supported by our findings that tumor cells induced into senescence by doxorubicin or etoposide can give rise to viable tumors in vivo. We further demonstrate sensitivity of these senescent tumor cells to the senolytic ABT-263 (navitoclax), therefore providing a "two-hit" approach to eliminate senescent tumor cells that persist after exposure to chemotherapy or radiation. The sequential combination of therapy-induced senescence and ABT-263 could shift the response to therapy toward apoptosis by interfering with the interaction between BCL-XL and BAX. The administration of ABT-263 after either etoposide or doxorubicin also resulted in marked, prolonged tumor suppression in tumor-bearing animals. These findings support the premise that senolytic therapy following conventional cancer therapy may improve therapeutic outcomes and delay disease recurrence.

Vorinostat is genotoxic and epigenotoxic in the mouse bone marrow cells at the human equivalent doses.

Vorinostat was approved as the first histone deacetylase inhibitor for the management of cutaneous T cell lymphoma. However, it's in vivo genetic and epigenetic effects on non-cancerous cells remain poorly understood. As genetic and epigenetic changes play a critical role in the pathogenesis of carcinogenesis, we investigated whether vorinostat induces genetic and epigenetic alterations in mouse bone marrow cells. Bone marrow cells were isolated 24 h following the last oral administration of vorinostat at the doses of 25, 50, or 100 mg/kg/day for five days (approximately equal to the recommended human doses). The cells were then used to assess clastogenicity and aneugenicity by the micronucleus test complemented by fluorescence in situ hybridization assay; DNA strand breaks, oxidative DNA strand breaks, and DNA methylation by the modified comet assay; apoptosis by annexin V/PI staining analysis and the occurrence of the hypodiploid DNA content; and DNA damage/repair gene expression by polymerase chain reaction (PCR) Array. The expression of the mRNA transcripts were also confirmed by real-time PCR and western blot analysis. Vorinostat caused structural chromosomal damage, numerical chromosomal abnormalities, DNA strand breaks, oxidative DNA strand breaks, DNA hypomethylation, and programed cell death in a dose-dependent manner. Furthermore, the expression of numerous genes implicated in DNA damage/repair were altered after vorinostat treatment. Accordingly, the genetic/epigenetic mechanism(s) of action of vorinostat may play a role in its carcinogenicity and support the continued study and development of new compounds with lower toxicity.