Abdelkader E. Ashour

Carbonic anhydrase inhibitors: Benzenesulfonamides incorporating cyanoacrylamide moieties are low nanomolar/subnanomolar inhibitors of the tumor-associated isoforms IX and XII

A series of benzenesulfonamides incorporating cyanoacrylamide moieties (tyrphostine analogues) have been obtained by reaction of sulfanilamide with ethylcyanoacetate followed by condensation with aromatic/heterocyclic aldehydes, isothiocyanates or diazonium salts. The new compounds have been investigated as inhibitors of the metalloenzyme carbonic anhydrase (CA, EC 4. 2.1.1), and more specifically against the cytosolic human (h) isoforms hCA I and II, as well as the transmembrane, tumor-associated ones CA IX and XII, which are validated antitumor targets. Most of the new benzenesulfonamides were low nanomolar or subnanomolar CA IX/XII inhibitors whereas they were less effective as inhibitors of CA I and II. The structure-activity relationship for this class of effective CA inhibitors is also discussed. Generally, electron donating groups in the starting aldehyde reagent favored CA IX and XII inhibition, whereas halogeno, methoxy and dimethylamino moieties led to very potent CA XII inhibitors.

Metformin Rescues the Myocardium from Doxorubicin-Induced Energy Starvation and Mitochondrial Damage in Rats

Clinical use of doxorubicin (DOX) is limited by its cardiotoxic side effects. Recent studies established that metformin (MET), an oral antidiabetic drug, possesses an antioxidant activity. However, whether it can protect against DOX-induced energy starvation and mitochondrial damage has not been reported. Our results, in a rat model of DOX-induced cardiotoxicity, show that DOX treatment significantly increased serum levels of LDH and CK-MB, indicators of cardiac injury, and induced expression of hypertrophic gene markers. DOX also caused marked decreases in the cardiac levels of glutathione, CoA-SH and ATP, and mRNA expression of catalase and NQO-1. These biochemical changes were associated with myocardial histopathological and ultrastructural deteriorations, as observed by light and electron microscopy, respectively. Cotreatment with MET (500 mg/kg) eliminated all DOX-induced biochemical, histopathological, and ultrastructural changes. These findings demonstrate that MET successfully prevents DOX-induced cardiotoxicity in vivo by inhibiting DOX-induced oxidative stress, energy starvation, and depletion of intramitochondrial CoA-SH.

High gene expression of semaphorin 5A in pancreatic cancer is associated with tumor growth, invasion and metastasis

Semaphorin 5A (SEMA5A) is an axonal regulator molecule, which belongs to the Semaphorin family of proteins. Previously, we identified SEMA5A as a putative marker for aggressive pancreatic tumors. However, the expression, localization and functional significance of SEMA5A in pancreatic tumors remain unclear. In our study, we hypothesized that SEMA5A expression modulates pancreatic tumor growth and metastasis. We analyzed the constitutive expression and localization of SEMA5A in patient pancreatic tumors (n = 33) and unmatched normal pancreatic (n = 8) tissues and human pancreatic cancer cell lines (n = 16) with different histopathological characteristics. We observed significantly higher expression of SEMA5A protein expression (p < 0.05) in human pancreatic tumor tissue samples compared to normal pancreatic tissues. Similarly, the pancreatic cancer cell lines with higher tumorigenic and metastatic potentials as xenografts in nude mice expressed higher levels of SEMA5A mRNA compared to those with lower tumorigenic and metastatic potentials. Furthermore, we examined the functional role of SEMA5A in pancreatic tumor growth and invasion. Ectopic expression of mouse full‐length Sema5A in Panc1 (SEMA5A negative) cells significantly (p < 0.05) enhanced tumorigenesis, growth and metastasis in vivo as well as proliferation, invasiveness and homotypic aggregation in vitro. Together, these data demonstrate that the expression of SEMA5A in pancreatic cancer cells regulates tumorigenesis, growth, invasion and metastasis, and it also suggests a novel target for diagnosis and treatment of pancreatic cancer.

Regulation of TNF-α and NF-κB activation through the JAK/STAT signaling pathway downstream of histamine 4 receptor in a rat model of LPS-induced joint inflammation.

Histamine 4 receptor (H4R) is a novel target for the pharmacological modulation of histamine-mediated immune signals during inflammatory diseases. The purpose of this study was to assess the effects of the H4R agonist 4-methylhistamine dihydrochloride (4-MeH) and antagonist JNJ7777120 (JNJ) in the inflamed rat knee. Animals were fasted for 18h before a single dose of 4-MeH or JNJ (30mg/kg) was administered intraperitoneally (i.p.), both followed by intra-articular (i.a.) injection of LPS 2h later. Blood and synovial fluid were collected after a short incubation period and TNF-α, NF-κB, and IkB-α levels were measured via flow cytometry. Additionally, we assessed the effects of H4R engagement on the expression of IL-1β, TNF-α, and NF-κB mRNAs and the protein levels of TNF-α, NF-κB, JAK-1, and STAT-3 in the inflamed knee tissue. These results revealed increased TNF-α and NF-κB expression and decreased IkB-α levels in both the LPS alone and 4-MeH treated groups in whole blood and synovial fluid. Further, IL-1β, TNF-α, and NF-κB mRNA levels were significantly increased and western blot analysis confirmed increased expression of TNF-α, NF-κB, JAK-1, and STAT-3 in both LPS and 4-MeH treatment groups. Furthermore, these increases were completely inhibited in the inflamed knee tissue of the JNJ-treated group. Thus, the inhibition of inflammatory mediators and signaling pathways by the H4R antagonist JNJ suggests the anti-arthritic importance of this molecule.

Salubrious effects of dexrazoxane against teniposide-induced DNA damage and programmed cell death in murine marrow cells

The intention of the present study was to answer the question whether the catalytic topoisomerase-II inhibitor, dexrazoxane, can be used as a modulator of teniposide-induced DNA damage and programmed cell death (apoptosis) in the bone marrow cells in vivo. The alkaline single cell gel electrophoresis, scoring of chromosomal aberrations, micronuclei and mitotic activity were undertaken in the current study as markers of DNA damage. Apoptosis was analysed by the occurrence of a hypodiploid DNA peak and caspase-3 activity. Oxidative stress marker such as intracellular reactive oxygen species production, lipid peroxidation, reduced and oxidised glutathione were assessed in bone marrow as a possible mechanism underlying this amelioration. Dexrazoxane was neither genotoxic nor apoptogenic in mice at the tested dose. Moreover, for the first time, it has been shown that dexrazoxane affords significant protection against teniposide-induced DNA damage and apoptosis in the bone marrow cells in vivo and effectively suppresses the apoptotic signalling triggered by teniposide. Teniposide induced marked biochemical alterations characteristic of oxidative stress including accumulation of intracellular reactive oxygen species, enhanced lipid peroxidation, accumulation of oxidised glutathione and reduction in the reduced glutathione level. Prior administration of dexrazoxane ahead of teniposide challenge ameliorated these biochemical alterations. It is thus concluded that pretreatment with dexrazoxane attenuates teniposide-induced oxidative stress and subsequent DNA damage and apoptosis in bone marrow cells. Based on our data presented, strategies can be developed to decrease the teniposide-induced DNA damage in normal cells using dexrazoxane. Therefore, dexrazoxane can be a good candidate to decrease the deleterious effects of teniposide in the bone marrow cells of cancer patients treated with teniposide.

Molecular cytogenetic evaluation of the mechanism of micronuclei formation induced by camptothecin, topotecan, and irinotecan

We used the conventional bone marrow micronucleus test complemented with the fluorescent in situ hybridization with the minor satellite DNA probe to investigate the mechanisms of induction of micronuclei in mice treated with camptothecin and its clinical antineoplastic analogues topotecan and irinotecan. All experiments were performed with male Swiss albino mice. Single doses of 1 mg/kg camptothecin or 0.6 mg/kg topotecan were injected intraperitoneally and bone marrow was sampled at 30 hr (camptothecin) or 24 hr (topotecan) after treatment. A dose of 60 mg/kg irinotecan was injected intravenously, once every fourth day for 13 days and bone marrow was sampled 24 hr after the last treatment. In animals treated with camptothecin, a total of 1.07% micronuclei were found and 70% of them were centromere‐negative, indicating their formation by DNA strand breaks and reflecting the predominant clastogenic activity of camptothecin. Exposure to topotecan and irinotecan yielded 1.71 and 0.83% micronuclei, respectively. About 52.7 and 48.8% of the induced micronuclei, respectively, were centromere‐positive, indicating their formation by whole chromosomes and reflecting the aneugenic activity of both compounds. Correspondingly, about 47.3 and 51.2% of the induced micronuclei, respectively were centromere‐negative, demonstrating that topotecan and irinotecan not only induce chromosome loss but also DNA strand breaks. Both the clastogenic and aneugenic potential of these drugs can lead to the development of secondary tumors and abnormal reproductive outcomes. Therefore, the clinical use of these agents must be weighed against the risks of secondary malignancies in cured patients and persistent genetic damage of their potential offspring. Environ. Mol. Mutagen. 2009.

Spleen but not tumor infiltration by dendritic and T cells is increased by intravenous adenovirus-Flt3 ligand injection

Dendritic cell (DC) expansion is regulated by the hematopoietic growth factor fms-like tyrosine kinase 3 ligand (Flt3L). DCs are critical to the control of tumor growth and metastasis, and there is a positive correlation between intratumoral DC infiltration and clinical outcome. In this report, we first demonstrate that single intravenous (i.v.) injections of adenovirus (Adv)-Flt3L significantly increased splenic dendritic, B, T and natural killer (NK) cell numbers in both normal and mammary tumor-bearing mice. In contrast, the numbers of DCs and T cells infiltrating the tumors were not increased. Consistent with the minimal effect on immune cell infiltration, i.v. Adv-Flt3L injections had no therapeutic activity against orthotopic mammary tumors. In addition, we noted tumor and Adv-Flt3L expansion of Gr1+CD11b+ immature myeloid suppressor cells (IMSCs), which may inhibit the therapeutic efficacy of Adv-Flt3L-expanded DCs.

Secreted semaphorin 5A suppressed pancreatic tumour burden but increased metastasis and endothelial cell proliferation

Background:Our earlier reports demonstrated that membrane-bound semaphorin 5A (SEMA5A) is expressed in aggressive pancreatic cancer cells and tumours, and promotes tumour growth and metastasis. In this study, we examine whether (1) pancreatic cancer cells secrete SEMA5A and (2) that secreted SEMA5A modulates certain phenotypes associated with tumour progression, angiogenesis and metastasis through various other molecular factors and signalling proteins.Methods and results:In this study, we show that human pancreatic cancer cell lines secrete the extracellular domain (ECD) of SEMA5A (SEMA5A-ECD) and overexpression of mouse Sema5A-ECD in Panc1 cells (not expressing SEMA5A; Panc1-Sema5A-ECD; control cells - Panc1-control) significantly increases their invasion in vitro via enhanced ERK phosphorylation. Interestingly, orthotopic injection of Panc1-Sema5A-ECD cells into athymic nude mice results in a lower primary tumour burden, but enhances the micrometastases to the liver as compared with Panc1-control cells. Furthermore, there is a significant increase in proliferation of endothelial cells treated with conditioned media (CM) from Panc1-Sema5A-ECD cells and a significant increase in microvessel density in Panc1-Sema5A-ECD orthotopic tumours compared with those from Panc1-control cells, suggesting that the increase in liver micrometastases is probably due to increased tumour angiogenesis. In addition, our data demonstrate that this increase in endothelial cell proliferation by Sema5A-ECD is mediated through the angiogenic molecules – interleukin-8 and vascular endothelial growth factor.Conclusion:Taken together, these results suggest that a bioactive, secreted form of Sema5A-ECD has an intriguing and potentially important role in its ability to enhance pancreatic tumour invasiveness, angiogenesis and micrometastases.

Grape seed proanthocyanidin extract has potent anti-arthritic effects on collagen-induced arthritis by modifying the T cell balance.

Rheumatoid arthritis (RA) is an autoimmune disease characterised by chronic inflammation of the synovial joints, joint malformations, and disability. The continuous use of conventional anti-inflammatory drugs is associated with severe adverse effects. Grape seed proanthocyanidin extract (GSPE) is considered to have protective effects against several diseases. In this study based on the mouse adjuvant-induced-arthritis (AIA) model, we examined the effects of GSPE on the key mediators of arthritic inflammation, namely T cell subsets, glucocorticoid-induced tumour necrosis factor receptor (GITR) expressing cells, CD4(+)CD25(+)Foxp3(+) regulatory T (Treg) cells, Th17 cells, Th1/Th2 cytokines, and inflammatory mediator gene expression. We treated BALB/c mice with 25, 50, or 100 mg/kg GSPE or saline daily (14 days) per orally (p.o.) at the onset of AIA. At the peak phase of AIA (day 14), the heparinised whole blood and ankle joints of all groups were collected and tested. GSPE-treated mice showed a substantial reduction in the levels of T cell subsets, GITR-expressing cells, and Th1 cytokines as well as the inflammatory mediators (MCP-1, MIP-2, and ICAM-1) that induce them compared with the vehicle-treated (saline) and arthritic mice. However, GSPE significantly upregulated the number of Tregs and Th2 cytokine producing cell number or it also induced Th17/Treg rebalance and orchestrated various pro-inflammatory and anti-inflammatory cytokines and the gene expression of their mediators that mediate cellular infiltration into the joints. This might, contribute to its anti-arthritic activity. Our results suggest that p.o. treatment with GSPE attenuated AIA in mice might offer a promising alternative/adjunct treatment for RA.

Poly(ADP-ribose) polymerase-1 inhibitor modulates T regulatory and IL-17 cells in the prevention of adjuvant induced arthritis in mice model

Rheumatoid arthritis (RA) is one of the major autoimmune diseases of global prevalence. Irrespective of much research in RA disease, no drugs with capable safety profiles are yet available. Poly(ADP-ribose) polymerase-1 (PARP-1) synthesizes and transfers ADP ribose polymers to target proteins, and regulates DNA repair and genomic integrity maintenance. PARP-1 also plays a crucial role in the progression of the inflammatory response, and its inhibition confers protection in several models of inflammatory disorders. We investigated the possible anti-arthritic effects of the PARP-1 inhibitor 5-aminoisoquinolinone (5-AIQ) in a mouse model of adjuvant induced arthritis (AIA). In this study, we examined the effects of 5-AIQ on the key mediators of arthritic inflammation, namely, edema and arthritic score, T cell subsets, regulatory T (Treg) cells, IL-17A, GITR expressing cells, NF-kB p65, IkB-α and pro and anti-inflammatory mediators mRNA expression levels. PARP-1 inhibition 5-AIQ treatment significantly attenuated the severity of AIA, reduced the arthritis scores, a substantial reduction in the levels of T cell subsets, IL-17A, NF-kB p65, GITR expressing cells, and as well as the pro-inflammatory mediators. However, 5-AIQ significantly up-regulated the number of Tregs cells, IkB-α levels and mRNA expression of anti-inflammatory mediators. Our results suggest that treatment with 5-AIQ attenuated AIA in mice might offer a promising alternative/adjunct treatment for RA.