Inhibition of human topoisomerase II by anti-neoplastic benzazolo[3,2-alpha]quinolinium chlorides

Previously we reported [20] that there is no correlation between the cytotoxic activity of four new structural analogs of the antitumor DNA intercalator 3-nitrobenzothiazolo[3,2-a]quinolinium chloride (NBQ-2) and their interaction with DNA. In the present study, we present evidence suggesting that the molecular basis for the anti-proliferative activity of these drugs is the inhibition of topoisomerase II. The NBQ-2 derivatives inhibited the relaxation of supercoiled DNA plasmid pRYG mediated by purified human topoisomerase II. Inhibition of the decatenation of kinetoplast DNA mediated by partially purified topoisomerase II extracted from the human histiocytic lymphoma U937 (a cell line previously shown to be sensitive to the drugs) was also caused by these drugs. The potency of the benzazolo[3,2-a]quinolinium drugs against topoisomerase II in vitro was the following: 7-(1-propenyl)-3-nitrobenzimidazolo[3,2-a]quinolinium chloride (NBQ-59) > 4-chlorobenzothiazolo[3,2-a]quinolinium chloride (NBQ-76) > 7-ethyl-3-nitrobenzimidazolo[3,2-a]quinolinium chloride (NBQ-48) > 7-benzyl-3-nitrobenzimidazolol[3,2-a]quinolinium chloride (NBQ-38). This rank of potency for topoisomerase II inhibition correlated very well with the cytotoxicity elicited by these drugs. Furthermore, significant levels of topoisomerase II/DNA cleavage complex induced by these drugs in vivo were detected when U937 cells were treated with NBQ-59 and NBQ-76 whereas NBQ-38 and NBQ-48 produced negligible amounts of the cleavage complex. Our results strongly suggest that topoisomerase II is the major cellular target of this family of compounds.     

A similar pattern of chromosomal alterations in prostate cancers from African-Americans and Caucasian Americans

A combination of genetic and epigenetic factors may explain the disproportionate incidence and mortality of prostate cancer among African-American males (AAMs) as compared with Caucasian American males (CAMs). We wished to determine whether primary prostate cancers from AAMs and CAMs harbor different patterns or frequencies of chromosomal alterations. Comparative genomic hybridization (CGH) was performed on clinically localized, untreated primary prostate cancers from 16 AAMs and 16 CAMs. Detailed statistical analysis was used to delineate gains and deletions with high sensitivity and specificity and to compare the frequency and pattern of alterations between the two groups of tumors. The two groups of patients had indistinguishable preoperative serum prostate-specific antigen levels, and the two groups of tumors had similar pathological stages and grades. Chromosomal gains and deletions occurred in regions known to be frequently altered in prostate cancer. Specifically, the most frequent alterations were deletions of regions on chromosomes 13q, 5q, 16q, and 8p and gains of regions on 8q and 5q. When tumors from AAMs and CAMs were compared, the frequencies of alteration (deletion, gain, or no alteration) were similar across 98.9% of the length of the genome. The patterns of alterations of the most frequently altered chromosomes were also similar between tumors from AAMs and CAMs. We concluded that primary prostate cancers from AAMs and CAMs harbor a similar pattern and frequency of chromosomal alterations. These data support the notion that sporadic prostate cancers from AAMs and CAMs develop by similar chromosomal mechanisms. Biological differences, if present, do not occur on the chromosomal level.     

Regulation of stress-induced cytokine production by pyridinylimidazoles; inhibition of CSBP kinase

Members of three classes of pyridinylimidazoles bind with varying affinities to CSBP (p38) kinase which is a member of a stress-induced signal transduction pathway. Based upon SAR and protein homology modeling, the pharmacophore and three potential modes of binding to the enzyme are presented. For a subset of pyridinylimidazoles, binding is shown to correlate with inhibition of CSBP kinase activity, whereas no significant inhibition of PKA, PKC alpha and ERK kinase activity is observed.     

NMR spectroscopic studies of the DNA-binding domain of the monomer-binding nuclear orphan receptor, human estrogen related receptor-2. The carboxyl-terminal extension to the zinc-finger region is unstructured in the free form of the protein

Unlike steroid and retinoid receptors, which associate with DNA as dimers, human estrogen related receptor-2 (hERR2) belongs to a growing subclass of nuclear hormone receptors that bind DNA with high affinity as monomers. A carboxyl-terminal extension (CTE) to the zinc-finger domain has been implicated to be responsible for determining the stoichiometry of binding by a nuclear receptor to its response element. To better understand the mechanism by which DNA specificity is achieved, the solution structure of the DNA-binding domain of hERR2 (residues 96-194) consisting of the two putative zinc fingers and the requisite 26-amino acid CTE was analyzed by multidimensional heteronuclear magnetic resonance spectroscopy. The highly conserved zinc-finger region (residues 103-168) has a fold similar to those reported for steroid and retinoid receptors, with two helices that originate from the carboxyl-terminal ends of the two zinc fingers and that pack together orthogonally, forming a hydrophobic core. The CTE element of hERR2 is unstructured and highly flexible, exhibiting nearly random coil chemical shifts, extreme sensitivity of the backbone amide protons to solvent presaturation, and reduced heteronuclear (1H-15N) nuclear Overhauser effect values. This is in contrast to the dimer-binding retinoid X and thyroid hormone receptors, where, in each case, a helix has been observed within the CTE. The implications of this property of the hERR2 CTE are discussed.     

ACE inhibitors and proteinuria

This review discusses the clinical consequences of urinary protein loss and the effects of inhibitors of the angiotensin converting enzyme (ACE) on this clinical finding. Proteinuria appears to be an important risk factor for renal function deterioration and for cardiovascular mortality. ACE inhibitors have been shown to reduce proteinuria more effectively than other antihypertensives. Their antiproteinuric effect seems to be independent of the underlying renal disease, and is mediated by a specific, not yet fully elucidated mechanism. Urinary protein loss related phenomena, such as hypoalbuminemia and aberrant lipoprotein profile, tend to improve also during ACE inhibitor treatment. Furthermore, ACE inhibition has been shown to prevent the renal function deterioration that is frequently observed in patients with renal disease. Interestingly, it has recently been shown that in proteinuric patients with renal disease the initial proteinuria lowering response to ACE inhibition predicts long-term renal function outcome during this treatment the more proteinuna is lowered during the first months, the better renal function will be preserved over the following years. Because of these favorable effects ACE inhibitors have become a widely used class of agents in nephrology. They are not only prescribed for lowering blood pressure in the hypertensive renal patient, but also as symptomatic treatment of patients with proteinuria, and to prevent renal function loss in patients with both diabetic and non-diabetic renal disease.     

1,25-Dihydroxyvitamin D3 pretreatment limits prostaglandin biosynthesis by cytokine-stimulated adult human osteoblast-like cells

The addition of fibronectin fragments to cultured cartilage causes an initial suppression of proteoglycan synthesis, induction of matrix metalloproteinases, and resultant decrease in proteoglycan content by about 50% during the first few days in culture. Because the proteoglycan loss appears to be limited, we investigated whether the fibronectin fragments induce anabolic responses that might counter the damage. The effects of various lengths of exposure of cultured cartilage to the fibronectin fragment on proteoglycan content, proteoglycan synthesis rates, stromelysin-1 release, and tumor necrosis factor-alpha, interleukin-1alpha, and interleukin-6 release were investigated. The results showed that about 7 days of exposure of cultured cartilage to the fibronectin fragment was required for maximal cytokine release, proteoglycan depletion, and stromelysin-1 release. However, nearly maximal suppression of proteoglycan synthesis occurred within 1 day of the addition of the fibronectin fragment and, after its removal, the rates increased to supernormal levels. Decreasing exposure to 3 days caused only a small decrease in cartilage proteoglycan content, although stromelysin-1 release still occurred. Decreasing exposure to 1 day caused an immediate increase in proteoglycan synthesis and an increase to supernormal proteoglycan contents. The effect of first treating cartilage with the fibronectin fragment for various periods and then allowing a recovery was to make the cartilage more resistant to secondary exposures. This study shows that cartilage damage can be caused by short exposures to the fibronectin fragment and that exposures either optimal or suboptimal for damage additionally amplify anabolic processes to make the cartilage resistant to further damage and, thus, condition it against pending amplification of damage.     

Immunoepidemiology of Dracunculus medinensis infections I. Antibody responses in relation to infection status

OBJECTIVES: The sites of recurrent carcinoma of the prostate were localized with radiolabeled monoclonal antibody, and these sites were correlated with the response of patients treated with pelvic radiation after prostatectomy. METHODS: Radionuclide scans were performed with indium 111-labeled CYT-356, a monoclonal antibody that binds to prostate epithelial cells, in 48 men diagnosed with recurrent carcinoma detected by prostate-specific antigen (PSA) screening after radical retropubic prostatectomy. RESULTS: In 48 patients with recurrent carcinoma detected by PSA screening following radical retropubic prostatectomy, 73% had monoclonal antibody activity beyond the prostatic fossa, and only 3 patients (6%) had activity in the prostatic fossa alone; 65% had monoclonal antibody activity in pelvic lymph nodes despite the fact that lymph node dissections were pathologically negative at the time of prostatectomy in 90% of the patients; and 23% of patients had monoclonal antibody activity in abdominal and extrapelvic retroperitoneal nodes. Of 48 patients, 13 underwent external beam radiation therapy after monoclonal antibody scans. Six patients had scans showing activity beyond the field of radiation, and radiation therapy failed in 4 of these patients. Seven patients had scans with no activity beyond the field of radiation therapy, and radiation therapy failed in only 2 of these patients. CONCLUSIONS: The scans frequently show monoclonal antibody uptake in pelvic, abdominal, and extrapelvic retroperitoneal sites beyond the region of limited obturator node dissections and may account for the understaging and subsequent failure of radical prostatectomy in some patients. The monoclonal antibody scan seems to be a good predictor of which patients will respond to radiation therapy after radical prostatectomy, but because these patients often have nodal activity beyond the radiated field, this initial response may not be curative.     

Intracranial hemorrhage: diagnosis and emergency management

Intracranial hemorrhages are an important cause of acute neurologic disease presenting in the emergency setting. To optimize outcome, it is important that the physician quickly recognize intracranial hemorrhages. To minimize mortality and neurologic morbidity, it is often necessary to initiate urgent therapy in the emergency rooms and to obtain neurosurgical consultation in order to pursue early surgical therapy. This article discusses the recognition and early treatment of the various types of intracranial hemorrhages.     

Regeneration of the ferrous heme of soluble guanylate cyclase from the nitric oxide complex: acceleration by thiols and oxyhemoglobin

Soluble guanylate cyclase (sGC) catalyzes the conversion of GTP to cGMP and is activated several hundred-fold by binding of nitric oxide (*NO) to the heme prosthetic group. We have examined the stability of the nitrosyl-heme complex of sGC (*NO-sGC) at 37 degreesC in order to determine whether simple dissociation of *NO from sGC could account for the observed in vivo deactivation time. Recombinant sGC was purified from Sf9 cells coinfected with baculoviruses containing the cDNAs for the alpha1 and beta1 subunits of rat lung sGC. The purified protein contained a stoichiometric equivalent of ferrous high-spin heme. Characterization of the purified protein found it to be essentially identical to that purified from bovine lung. Ferrous-nitrosyl sGC prepared anaerobically and exchanged into aerobic buffer containing no reducing agents was essentially stable on ice and had a half-life of approximately 90 min at 37 degreesC. In the presence of thiols [DTT, glutathione (GSH), or L-cysteine], *NO was rapidly lost from sGC regenerating the ferrous high-spin form of the heme. The half-life of *NO-sGC in the presence of 1 mM GSH at 37 degreesC was 6.3 min. In the presence of oxyhemoglobin, the half-life was further reduced to 2.9 min. Although these rates are not fast enough to account for that observed in vivo, and thus probably involve additional agent(s), these data do imply a role for low molecular weight thiols, such as GSH, and oxyferrohemoproteins, such as oxymyoglobin, in the deactivation of sGC.