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Saeed R Khan

Publications and source records attributed to Saeed R Khan.

At least 19 recordsLinked to original sources
PubMed2006

A boronic-chalcone derivative exhibits potent anticancer activity through inhibition of the proteasome.

Chalcones and their derivatives have been shown to have potent anticancer activity. However, the exact mechanisms of cytotoxic activity remain to be established. In this study, we have evaluated a series of boronic chalcones for their anticancer activity and mechanisms of action. Among the eight chalcone derivatives tested, 3,5-bis-(4-boronic acid-benzylidene)-1-methyl-piperidin-4-one (AM114) exhibited most potent growth inhibitory activity with IC50 values of 1.5 and 0.6 microM in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and colony formation assay, respectively. The cytotoxic activity of AM114 was shown to be associated with the accumulation of p53 and p21 proteins and induction of apoptosis. Mechanistic studies showed that AM114 treatment inhibited the chymotrypsin-like activity of the 20S proteasome in vitro, leading to a significant accumulation of ubiquitinated p53 and other cellular proteins in whole cells. In vitro studies showed that AM114 did not significantly disrupt the interaction of p53 and murine double minute 2 protein. It is noteworthy that AM114 as a single agent was preferentially toxic to cells with wild-type p53 expression, whereas combination of this compound with ionizing radiation (IR) significantly enhanced the cell-killing activity of IR in both wild-type p53 and p53-null cells. Together, these results indicate that the boronic chalcone derivative AM114 induces significant cytotoxic effect in cancer cells through the inhibition of the cellular proteasome and provide a rationale for the further development of this class of compounds as novel cancer chemotherapeutic agents.

Antineoplastic Agents↗
PubMed2006

Benzoylphenylurea sulfur analogues with potent antitumor activity.

A novel series of BPU analogues were synthesized and evaluated for antitumor activity. In particular, BPU sulfur analogues 6n and 7d were shown to possess up to 10-fold increased potency, when compared to 1 (NSC-639829), against cancer cell lines. 6n was more effective than 1 in causing apoptosis of MCF-7 cells. When compared to other drugs with a similar mechanism of action, 6n retained significant ability to inhibit tubulin assembly, with an IC(50) of 2.1 microM.

Antineoplastic Agents↗
PubMed2006

Adhesion force between calcium oxalate monohydrate crystal and kidney epithelial cells and possible relevance for kidney stone formation.

AFM interaction force measurements have been performed between calcium oxalate monohydrate crystal (COM) colloidal probes and monolayers of renal epithelial cells (on a polymer substrate) in artificial urine (AU) solutions. The adhesion force was measured for the COM/MDCK cell interaction, while no adhesion force was found for the COM/LLC-PK(1) cell interaction. Long-range repulsive forces for both lines of cells were measured in the range of 2-3 mum. After removal of the cell from the substrate by the AU flow, the basal membrane (BM), with a thickness of 100-200 nm, remained on the substrate. In this case, the shorter-range repulsive forces were found on the extending (approaching) portion of force/indentation curves. Similar to the COM/MDCK cell interaction, the retracting portions of curves for COM/basal membranes have shown the existence of the attractive force of adhesion for the interaction of COM with a BM of MDCK cells, while no adhesion was found for COM/BM LLC-PK(1) cells interaction. No adhesion force was found for the interaction of a BM (of any cells) with the silicon nitride tip. Besides the hydrodynamic reasons, the adhesion difference between LLC-PK(1) and MDCK cells possibly explains the preferential deposition of crystals only in collecting ducts (lined with MDCK-type cells) and the lack of the crystal deposition in the proximal tubules (lined with LLC-PK(1)-type cells). Previous treatments of cells with oxalate alone increased the adhesion force COM/BM MDCK; however, even after oxalate treatment there was small or no adhesion between COM and BM LLC-PK(1) cells. Note that the adhesion force for COM/BM MDCK is practically independent of the probe velocity, i.e., does not have the viscous origin. Evaluation of the adhesion energy shows that this force should be related to the ionic or hydrogen bonds of samples.

Adhesiveness↗
PubMed2006

Role of lipids in urinary stones: studies of calcium oxalate precipitation at phospholipid langmuir monolayers.

This article reviews the authors' experiments on calcium oxalate growth at lipid monolayers. Calcium oxalate is the principal mineral component of most urinary stones. Membrane constituents associate either actively or passively with calcific minerals during stone formation, and it has been proposed that lipid assemblies play a significant role, possibly providing sites for the initial nucleation event. Langmuir monolayers allow systematic studies of the heterogeneous precipitation of calcium oxalate at lipid assemblies. The influences of the chemical identity of the lipid headgroup, the organization of the monolayer, and the presence of heterogeneities and phase boundaries within the monolayer have been explored.

Calcium Oxalate↗
PubMed2006

Anticancer activities of novel chalcone and bis-chalcone derivatives.

A series of novel chalcones and bis-chalcones containing boronic acid moieties has been synthesized and evaluated for antitumor activity against the human breast cancer MDA-MB-231 (estrogen receptor-negative) and MCF7 (estrogen receptor-positive) cell lines and against two normal breast epithelial cell lines, MCF-10A and MCF-12A. These molecules inhibited the growth of the human breast cancer cell lines at low micromolar to nanomolar concentrations, with five of them (1-4, 9) showing preferential inhibition of the human breast cancer cell lines. Furthermore, bis-chalcone 8 exhibited a more potent inhibition of colon cancer cells expressing wild-type p53 than of an isogenic cell line that was p53-null.

Antineoplastic Agents↗
PubMed2006

Renal tubular damage/dysfunction: key to the formation of kidney stones.

Supersaturation is the driving force behind crystal formation in the kidneys. It can, however, result only in the formation of crystals which can be harmlessly expelled. For stone formation, crystals must form in the kidneys and be retained there, which is indeed a rare occurrence. Crystalluria is common while stone formation is not. Only pathological changes in the kidneys including renal injury and dysfunction can accomplish crystal retention. Lethal epithelial cellular injury promotes crystal nucleation, aggregation and retention. Sub-lethal injury or dysfunctional cells may produce ineffective crystallization modulators and localized areas of supersaturation in the interstitium. The former will affect crystallization in the urine while the latter may cause precipitation in the interstitium and development of Randall's plaques.

Animals↗
PubMed2006

Reactive oxygen species mediated calcium oxalate crystal-induced expression of MCP-1 in HK-2 cells.

Under severe hyperoxaluric conditions calcium oxalate crystals often deposit in the renal interstitium and produce localized inflammation. We have proposed that renal epithelial cells exposed to CaOx crystals produce chemoattractants such as monocyte chemoattractant protein-1 (MCP-1). MCP-1 synthesis is mediated by reactive oxygen species (ROS). HK-2 cells of human renal epithelial line were exposed to CaOx crystals for different lengths of time. The culture media was tested for cell injury marker LDH, and subjected to enzyme-linked immunosorbent assay to determine the secretion of MCP-1 protein. Cell expression of MCP-1 was assessed by Western blot analysis. Gene expression was determined by reverse transcriptase-polymerase chain reaction. The data clearly showed that the HK-2 cells express MCP-1 gene and protein. The MCP-1 mRNA expression was increased following exposure to CaOx crystals, which was reduced upon treatment with free radical scavengers, catalase and superoxide dismutase. Results indicate that CaOx crystals strongly induce MCP-1 synthesis and secretion by the HK-2 cells and production is mediated by intracellular ROS production. Based on these and other data, antioxidant therapy and blockade of rennin-angiotensin system may prove beneficial for the prevention of end stage renal disease caused by hyperoxaluria and CaOx crystal deposition.

Blotting, Western↗
PubMed2006

Oxalate ions and calcium oxalate crystal-induced up-regulation of osteopontin and monocyte chemoattractant protein-1 in renal fibroblasts.

OBJECTIVE: To examine the responses of renal fibroblasts to high oxalate (Ox) and calcium Ox (CaOx) crystals, as the latter are found in the renal interstitium of patients with primary or enteric hyperoxaluria, and in animals with experimental CaOx nephrolithiasis, and are associated with tubulointerstitial inflammation (TI). TI might begin with the production of chemoattractants by the renal epithelial cells exposed to high Ox and/or CaOx crystals; as Ox levels are also high in the renal interstitium and crystal deposition in nephrolithiasis might start in the interstitium, we hypothesized that renal fibroblasts might also be involved in the development of TI. MATERIALS AND METHODS: We exposed renal fibroblast cells of line NRK 49F in vitro to Ox ions (500 micromol/L) or CaOx monohydrate crystals (67 microg/cm(2)). We assessed the production of osteopontin and monocyte chemoattractant protein-1 (MCP-1), and expression of their mRNA, in the cells. We also determined the cellular malondialdehyde content as a marker of reactive oxygen species (ROS)-induced lipid peroxidation, and Trypan blue staining and the release of lactate dehydrogenase as markers of injury. RESULTS: Similar to renal epithelial cells, renal fibroblasts were stimulated by exposure to Ox and CaOx crystals. They showed signs of injury and ROS-induced lipid peroxidation. The mRNA expression and production of osteopontin and MCP-1 increased significantly. CONCLUSIONS: These results indicate that fibroblasts respond to high Ox and CaOx crystals by up-regulating specific pathways producing pro-inflammatory conditions. Migration of monocytes/macrophages to sites of interstitial crystal deposits can lead to localized interstitial inflammation and fibrosis.

Animals↗
PubMed2005

Citrate and vitamin E blunt the shock wave-induced free radical surge in an in vitro cell culture model.

Free radical formation plays a major role in shock wave lithotripsy induced renal damage. Moreover, previous studies suggest that free radicals may also promote de novo calcium oxalate crystallization of previously damaged urothelium. Citrate is a known inhibitor of renal stone formation and has also been used as a free radical scavenger. Using an in vitro model with Madin-Darby canine kidney (MDCK) cells, we investigated the influence of two free radical scavengers, citrate and vitamin E, on the prevention of the shock wave-induced free radical surge. Suspensions of MDCK cells were placed in containers for shock wave exposure. Six groups of six containers each were examined: (a) no scavengers 0 shocks, (b) no scavengers 100 shocks, (c) citrate 0 shocks, (d) citrate 100 shocks, (e) vitamin E 0 shocks, (f) vitamin E 100 shocks. An unmodified HM3 was used to deliver 100 shocks at 24 kV. The cell groups that were not shocked acted as the control group and were handled identically, except for the lack of shock wave exposure. After shock wave administration, the containers were emptied and cell suspensions were immediately centrifuged. The supernatant was examined for lactate dehydrogenase (LDH) and 8-isoprostane (8-IP), markers of cellular injury and free radical formation, respectively. Intracellular LDH uniformly increased in all groups exposed to shock wave energy. Similarly, 8-IP increased in all shocked groups. However, the 8-IP increase was significantly reduced when the free radical scavengers were employed. As citrate is a well-known inhibitor of calcium nephrolithiasis, its mechanism of action may be further enhanced, based on its ability to reduce free radical formation, by a protective effect on the urothelium. These data further support the use of citrate based medications during the peri-operative period of shock wave lithotripsy, not only to inhibit stone formation and facilitate fragment passage, but also to reduce the incidence of shock wave induced renal damage. Further studies are warranted to clinically test this hypothesis.

Animals↗
PubMed2005

Oxalate induced expression of monocyte chemoattractant protein-1 (MCP-1) in HK-2 cells involves reactive oxygen species.

Oxalate is a toxic end product of metabolism largely because of its propensity to crystallize and form calcium oxalate, which is insoluble at physiologic pH and often deposits at very unfortunate sites, notably the kidneys. In the current study, we investigated the oxalate-induced injury and up-regulation of monocyte-chemoattractant protein-1 (MCP-1) in HK-2 cells, a proximal tubular epithelial cell line derived from normal human kidney. The cells were exposed to oxalate ions for different lengths of time. The culture media was tested for LDH release, a cell injury marker. mRNA was isolated from the cells and subjected to reverse transcriptase-polymerase chain reaction. The data showed that oxalate exposure resulted in cell injury in a time and concentration dependent manner. The MCP-1 mRNA increased following exposure to oxalate and was reduced upon treatment with free radical scavengers, catalase and superoxide dismutase. These data support the importance of reactive oxygen species in the induction of expression of MCP-1 in renal epithelial cells. To our knowledge, this is the first report of MCP-1 expression and its upregulation by oxalate exposure in HK-2 cells.

Base Sequence↗
PubMed2005

Hyperoxaluria-induced oxidative stress and antioxidants for renal protection.

Renal cellular exposure to oxalate (Ox) and/or CaOx crystals leads to the production of reactive oxygen species (ROS), development of oxidative stress followed by injury and inflammation. Renal injury and inflammation appear to play a significant role in stone formation. ROS are produced from many sources and involve a variety of signaling pathways. Tissue culture and animal model studies show that treatments with anti-oxidants and free radical scavengers reduce Ox/CaOx crystal induced injuries. In addition, CaOx crystal deposition in kidneys is significantly reduced by treatments with antioxidants and free radical scavengers, indicating their efficacy. These results point towards a great potential for the therapeutic application of antioxidants and free radical scavengers to reduce stone recurrence particularly after shock wave lithotripsy, which is itself known to generate ROS and cause renal damage.

Animals↗
PubMed2005

Bis(pivaloyloxymethyl) thymidine 5'-phosphate is a cell membrane-permeable precursor of thymidine 5'-phosphate in thymidine kinase deficient CCRF CEM cells.

Bis(pivaloyloxymethyl) thymidine 5-phosphate (POM(2)-dTMP) has been investigated as a membrane-permeable prodrugs of dTMP. The growth inhibitory activity of POM(2)-TMP has been compared with thymidine (TdR) in wild type CCRF CEM cells (CEM) and a strain that lacks TdR kinase (CEM tk-). After 72 h incubation at 37 degrees C, TdR showed significant antiproliferative activity (IC(50)=27 microM) against CEM cells but was weakly effective (IC(50)=730 microM) against the mutant cell line. By comparison, bis(pivaloyloxymethyl) thymidine 5'-monophosphate (POM(2)-dTMP) was equally inhibitory (IC(50)=5 microM) to both cell lines. The growth inhibitory effects were reversed by deoxycytidine. Cellular [methyl-(3)H]dTTP pools increased linearly over 2h during incubation of CEM or CEM tk- with 5 microM POM(2)-[methyl-(3)H]dTMP. The incorporation of [methyl-(3)H]TdR into HClO(4)-insoluble cell residue by CEM tk- was <0.1% that of CEM and did not increase over 1h. In contrast, CEM tk- incorporated radioactivity from POM(2)-dTMP into acid insoluble residue at a rate 59% that of CEM. These results demonstrate that POM(2)-dTMP can penetrate into cells and serve as a source of dTMP.

Carbon Radioisotopes↗
PubMed2005

Oxalate-inducible AMBP gene and its regulatory mechanism in renal tubular epithelial cells.

The AMBP [A1M (alpha1-microglobulin)/bikunin precursor] gene encodes two plasma glycoproteins: A1M, an immunosuppressive lipocalin, and bikunin, a member of plasma serine proteinase inhibitor family with prototypical Kunitz-type domain. Although previously believed to be constitutively expressed exclusively in liver, the present study demonstrates the induction of this gene by oxalate in porcine proximal tubular LLC-PK1 cells and rat kidney. In liver, the precursor protein is cleaved in the Golgi network by a furin-like enzyme to release constituent proteins, which undergo glycosylation before their export from the cell. In the renal tubular cells, A1M and bikunin co-precipitate, indicating lack of cleavage of the precursor protein. As the expression of the AMBP gene is regulated by A1M-specific cis elements and transcription factors, A1M protein was studied as a representative of AMBP gene expression in renal cells. Oxalate treatment (500 microM) resulted in a time- and dose-dependent induction of A1M protein in LLC-PK1 cells. Of the four transcription factors, HNF-4 (hepatocyte nuclear factor-4) has been reported previously to be a major regulator of AMBP gene expression in liver. Electrophoretic mobility-shift assay, supershift assay, immunoreactivity assay and transfection-based studies showed the presence of an HNF-4 or an HNF-4-like protein in the kidney, which can affect the expression of the AMBP gene. In situ hybridization and immunocytochemical studies showed that the expression of this gene in kidney was mainly restricted to cells lining the renal tubular system.

Animals↗
PubMed2005

Design, synthesis and biological evaluation of novel riccardiphenol analogs.

A novel, facile, high yield, and less cumbersome synthesis of riccardiphenol analogs is described. The synthesized compounds were characterized and assessed for its in vitro activity in a panel of human cancer cell lines of differing origin: HuCCT-1, BxPC3, Panc-1, Mia-Paca, A431, Hep2, and HN006. HuCCT-1 was derived from an intrahepatic cholangiocarcinoma; BxPC3, Mia-Paca, and Panc-1 were derived from pancreatic cancers; A431 was derived from a vulvar epithelial carcinoma; and Hep2 and HN006 were derived from squamous cell carcinomas of the head and neck. The cytotoxicity of a newly developed riccardiphenol analog against human cancer cell lines was assessed. The cancer cells exhibited varying sensitivities to the compound, with IC50 values from 30 to 50 microM. This susceptibility was particularly interesting in the case of lines such as Hep2 and BxPC3 that are resistant to classic cytotoxic drugs as well as some targeted agents. These results demonstrate that the novel riccardiphenol analog has effective action against human-derived cancer cell in vitro.

Benzopyrans↗
PubMed2005

Diphenyleneiodium (DPI) reduces oxalate ion- and calcium oxalate monohydrate and brushite crystal-induced upregulation of MCP-1 in NRK 52E cells.

BACKGROUND: Our earlier studies have demonstrated upregulation of monocyte chemoattractant protein-1 (MCP-1) in NRK52E rat renal epithelial cells by exposure to oxalate (Ox) ions and crystals of calcium oxalate monohydrate (COM) or the brushite (Br) form of calcium phosphate. The upregulation was mediated by reactive oxygen species (ROS). This study was performed to investigate whether NADPH oxidase is involved in ROS production. METHODS: Confluent cultures of NRK52E cells were exposed to Ox ions or COM and Br crystals. They were exposed for 1, 3, 6, 12, 24 and 48 h for isolation of MCP-1 mRNA and 24 h for enzyme-linked immunosorbent assay (ELISA) to determine the secretion of protein into the culture medium. We also investigated the effect of free radical scavenger, catalase, and the NADPH oxidase inhibitor diphenyleneiodium (DPI) chloride, on the Ox- and crystal-induced expression of MCP-1 mRNA and protein. The transcription of MCP-1 mRNA in the cells was determined using real-time polymerase chain reaction. Hydrogen peroxide and 8-isoprostane were measured to investigate the involvement of ROS. RESULTS: Exposure of NRK52E cells to Ox ions as well as the crystals resulted in increased expression of MCP-1 mRNA and production of the chemoattractant. Treatment with catalase reduced the Ox- and crystal-induced expression of both MCP-1 mRNA and protein. DPI reduced the crystal-induced gene expression and protein production but not Ox-induced gene expression and protein production. CONCLUSIONS: Exposure to Ox ions, and COM and Br crystals stimulates a ROS-mediated increase in MCP-1 mRNA expression and protein production. Reduction in ROS production, lipid peroxidation, low-density lipoprotein release, and inducible MCP-1 gene and protein in the presence of DPI indicates an involvement of NADPH oxidase in the production of ROS.

Animals↗
PubMed2005

Bis(carbamoyloxymethyl) esters of 2',3'-dideoxyuridine 5'-monophosphate (ddUMP) as potential ddUMP prodrugs.

PURPOSE: We previously reported the synthesis of bis(pivaloyloxymethyl) 2',3'-dideoxyuridine 5'-monophosphate (POM2-ddUMP) (1a) as a membrane-transport prodrug formulation of the free parent nucleotide, ddUMP. Although successful at delivering ddUMP into cells in culture, POM2-ddUMP was rapidly degraded by plasma carboxylate esterases after intravenous administration to experimental animals, and therefore has limited therapeutic potential as a systemically administered prodrug. We now report the synthesis of bis(N,N'-dimethylcarbamoyloxymethyl)- and bis(N-piperidinocarbamoyloxymethyl) 2',3'-dideoxyuridine 5'-monophosphate [DM2-ddUMP (1b) and DP2-ddUMP (1c), respectively], analogues of POM2-ddUMP that were designed to be more resistant to degradation by plasma esterases. METHODS: After entering cell by passive diffusion, it was anticipated that loss of one of the carbamoyloxymethyl groups of 1b and 1c would occur by spontaneous chemical hydrolysis to give the intermediate phosphodiesters, 2b and 2c. Cleavage of the remaining carbamoyloxymethyl groups by cellular phosphodiesterase I would generate ddUMP. 1b and 1c were prepared by condensation of 2',3'-dideoxyuridine (ddU) with the appropriate bis(N-alkylcarbamoyloxymethyl) phosphate in DMA in the presence of triphenylphosphine and diethyl azodicarboxylate (the Mitsunobo reagent). RESULTS: The half-lives of 1b and 1c when incubated at a concentration of 10(-4) M in human plasma at 37 degrees C were 3.5 h and 3.7 h, respectively, similar to the half-lives observed under the same temperature conditions in 0.05 M aqueous phosphate buffer, pH 7.4. By contrast, the half-life of the POM2 prodrug, 1a, in plasma was only 5 min. The initial products of degradation of 1b and 1c were the phosphodiesters 2b and 2c. The latter compounds gave rise to ddUMP when incubated with snake venom phosphodiesterase I. CONCLUSION: These findings support the premise inherent in the design of 1b and 1c, namely that the carbamate prodrugs are far more resistant to hydrolysis by plasma carboxylate esterases than their POM counterparts and can revert to the free parent 5'-mononucletides by successive chemical and enzymatic hydrolysis. Further studies of 1b and 1c as membrane-permeable prodrugs of ddUMP are in progress.

Animals↗
PubMed2005

Citrate provides protection against oxalate and calcium oxalate crystal induced oxidative damage to renal epithelium.

PURPOSE: Oxalate and calcium oxalate (CaOx) crystals are injurious to renal epithelial cells. The injury is caused by the production of reactive oxygen species (ROS). Citrate is a well-known inhibitor of CaOx crystallization and as such it is one of the major therapeutic agents prescribed. Since citrate increases cellular reduced nicotinamide adenine dinucleotide phosphate and glutathione (GSH), we hypothesized that exogenously administered citrate should act as an antioxidant and protect cells from oxalate induced injury. MATERIALS AND METHODS: We exposed LLC-PK1 and MDCK cells to 500 microM/ml oxalate or 150 mug/cm calcium oxalate crystals for 30, 60 and 180 minutes with or without 3 mg/ml citrate in the medium. We determined cell viability by lactate dehydrogenase release and trypan blue exclusion, ROS involvement by changes in hydrogen peroxide and GSH, and lipid peroxidation by quantifying 8-isoprostane. RESULTS: The presence of citrate was associated with significant decrease in lactate dehydrogenase release (p <0.001) and staining with trypan blue (p <0.05). In addition, there was a significant increase in GSH (p <0.005) and a decrease in the production of hydrogen peroxide (p <0.05) and 8-isoprostane (p <0.0005) secretion into the culture medium when citrate was present in the medium. CONCLUSIONS: Citrate protects cells from oxalate and CaOx crystal induced injury by preventing lipid peroxidation through a decrease in ROS production. The results provide additional data for the beneficial role of citrate therapy for CaOx nephrolithiasis.

Calcium Oxalate↗