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F M Chen

Publications and source records attributed to F M Chen.

At least 91 records · Page 5Linked to original sources
PubMed1989

Binding of mithramycin to DNA in the presence of second drugs.

Comparative DNA equilibrium binding studies with mithramycin (MTR) and ethidium bromide in the presence and in the absence of second drugs were investigated by spectral titrations. Unusual curvatures (in contrast to those due to neighbor exclusion or anticooperativity) are found in the Scatchard plots of MTR-DNA titrations in the presence of netropsin, a minor-groove binder. Parallel studies with ethidium bromide indicate that although the presence of netropsin significantly reduces the binding ability of ethidium, no unusually curved Scatchard plots are obtained. The unusual curvature exhibited by the Scatchard plots of MTR titrations in the presence of netropsin indicates that the binding of netropsin greatly affects the MTR binding to DNA and can be simulated by an explicit incorporation of the second drug-DNA interaction in the binding formalism. Since netropsin is a minor-groove binder, its interference with the binding of MTR is in accord with the notion that MTR also binds at this groove. The observation of negligible effects on the DNA binding ability of MTR in the presence of either a major-groove or a phosphate group binder lends further support to this conclusion. Consistent with its guanine specificity, studies with synthetic polynucleotides suggest that MTR exhibits negligible affinity for poly(dA-dT).poly(dA-dT) or poly(dA).poly(dT).(ABSTRACT TRUNCATED AT 250 WORDS)

Binding Sites↗
PubMed1989

Hairpin formation of d(CGCG-TA-CGCG), d(CGCG-TG-CGCG) and their cytosine methylated analogs.

Hairpin formations of decamers d(CGCG-TA-CGCG), d(CGCG-TG-CGCG), and their m5dC analogs are evidenced by the existence of biphasic absorbance melting profiles in which the lower transition temperature increases with increasing oligomer concentration, whereas the higher melting temperature is concentration independent. The corresponding temperature dependent CD intensity at 285 nm exhibits a maximum around 55 degrees C. These observations are consistent with the interpretation that the lower temperature transition corresponds to the duplex to hairpin transformation while the melting of hairpins into single strands constitutes the higher temperature transition. The CD spectrum of the hairpin conformation appears to be characterized by a couplet with nearly equal positive and negative intensities at 285 and 255 nm, respectively, while a significantly smaller intensity at 285 nm is apparent for the duplex form. The hairpin conformation is suspected to contain a two-nucleotide loop. Titrations with NaCl further suggest that, in contrast to the TA sequence, the TG sequence with wobble base pairing favors Z formation under high salt conditions.

Base Sequence↗
PubMed1989

Studies on racemization associated with the use of benzotriazol-1-yl-tris (dimethylamino)phosphonium hexafluorophosphate (BOP).

The influence of the amount and nature of the necessary tertiary amine on racemization attending the BOP-mediated coupling of N-benzyloxycarbonylglycyl-dipeptides has been examined by determining the epimeric products by high-performance liquid chromatography. Racemization was encountered in all cases examined that included dichloromethane as solvent. The least racemization occurred when the base was diisopropylethylamine, the most when the base was N-methylmorpholine. Excess base promoted racemization. Racemization was significantly diminished but not eliminated when 1-hydroxybenzotriazole was added to the reaction mixtures. The above results pertain to the coupling of peptide segments; they do not pertain to the coupling of N-alkoxycarbonylamino acids.

Amines↗
PubMed1988

A novel method for the detection of necrotic lesions in human cancers.

Data are presented in support of the hypothesis that malignant tumors, containing abnormally permeable, degenerating cells, can be selectively detected using monoclonal antibodies to intracellular antigens. Biodistribution, imaging, and autoradiographic studies were performed in nude mice transplanted with four different human tumor cell lines to demonstrate the binding of radiolabeled antinuclear monoclonal antibodies within bulky tumors containing necrotic lesions. For these studies, two monoclonal antibodies, designated TNT-1 (IgG2a) and TNT-2 (IgM) were chosen since they were found to bind to abundant nuclear antigens which are retained in permeable, dying cells. F(ab')2 fragments prepared by pepsin digestion were radiolabeled with iodine-125 or iodine-131 by the iodogen method for i.v. administration. Biodistribution studies in nontumor-bearing BALB/c mice at various time intervals revealed normal patterns of antibody excretion with no accumulation of antibody in healthy organs. In contrast, biodistribution studies performed on Day 3 in tumor-bearing nude mice showed high tumor to organ ratios in those animals bearing necrotic tumors. Necrotic regions dissected at necropsy gave tumor to blood ratios as high as 131:1. Transplants having little demonstrable necrosis were found to have low tumor to blood ratios (0.4:1). Sequential imaging studies confirmed the high tumor-to-organ ratios and showed positive tumor imaging as early as 4 h. Autoradiographic studies of excised tumors showed the presence of label selectively in necrotic areas with preferential labeling over the nuclei of degenerating cells. Because of the universal presence of these nuclear antigens and the known prevalence of necrosis in tumors, this approach may be of value for the imaging and treatment of a wide variety of cancers in humans.

Animals↗
PubMed1988

Binding specificities of actinomycin D to self-complementary tetranucleotide sequences -XGCY-.

Binding of actinomycin D (ACTD) to self-complementary decamers d(ATA-XGCY-TAT), where XGCY = TGCA, AGCT, CGCG, and GGCC, has been investigated by equilibrium, kinetic, and thermal denaturation studies. The results indicate that despite the presence of a GC dinucleotide sequence, -GGCC- exhibits a much weaker binding affinity toward ACTD than the other three tetranucleotide sequences. Binding constants estimated from Scatchard plots indicate that binding to the -GGCC- site is at least an order of magnitude weaker than binding to -CGCG- and -AGCT-, which in turn is only slightly weaker than binding to the -TGCA- sequence. At 18.5 degrees C and 1% SDS, ACTD dissociates from d-(ATA-TGCA-TAT) with a slow characteristic time of 3300 s, roughly 4 times slower than dissociation from those containing -CGCG- and -AGCT- sequences and more than 2 orders of magnitude slower than that from -GGCC-. An 18.2 degrees C increase in the melting temperature is observed for the -TGCA-containing decamer upon binding of the ACTD, whereas increases of 10.3, 6.7, and 2.0 degrees C are observed for the -CGCG-, -AGCT-, and -GGCC-containing decamers, respectively. The effects observed by changing the adjacent base pair (sequence) may occur as a result of differential stacking and/or peptide ring-DNA groove interactions. Base sequence alterations adjacent to the ACTD binding site may result in differences in the minor groove environment and/or subtle conformational alterations at the intercalation site.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1988

Comparison of mono Q, superose-6, and ABx fast protein liquid chromatography for the purification of IgM monoclonal antibodies.

Nine immunoglobulin M (IgM) monoclonal antibodies (MAbs) produced in ascites fluids or in cell culture supernatants, have been purified on a fast protein liquid chromatography (FPLC) system using anion-exchange, size-exclusion, or mixed-mode chromatography matrices. The use of a mixed-mode ABx column provided an IgM that had a purity of greater than 99% after a single purification step. Anion-exchange chromatography using a Mono Q column, provided a partial purification of the IgM which could subsequently be purified to a product of ca. 90% purity (determined from sodium dodecyl sulfate polyacrylamide gel electrophoresis) by size-exclusion chromatography on a Superose-6 column. Alternatively, the ascites containing the IgM was ammonium sulfate precipitated and chromatographed on the Superose-6 column under normal- as well as high-ionic strength conditions, which also yielded a product of ca. 90% purity. The purification of IgM from concentrated cell culture supernatants was evaluated using the Superose-6 or the ABx column. IgM purified from this source was greater than 99% pure when chromatographed on the mixed-mode column and ca. 60% pure on the size-exclusion column. MAbs from each of the procedures retained their immunoreactivity, as shown by indirect immunofluorescence staining of fixed cell preparations. A comparison of these methods revealed that mixed mode chromatography was simple, efficient, and yielded a product of high purity. The optimization of these methods facilitates the large-scale purification of mouse IgM MAbs and provides practical procedures for generating IgMs for use as diagnostic and therapeutic reagents.

Anion Exchange Resins↗
PubMed1988

Effects of A:T base pairs on the B-Z conformational transitions of DNA.

Effects of A:T base pairs on the propensity of B to Z conformational transitions have been investigated by the CD salt titrations on d(CG)5' d(GC)5' terminal or central A:T replaced decamers, and terminal A:T appended dodecamers. The presence of A:T at the center greatly inhibits the B to Z transition of both G:C decamers. Moderate Z inhibitions are shown by terminal A:T replacements and additions to d(CG)5' with the former exhibiting a stronger effect. In contrast, the addition and replacement with A:T at the terminals of d(GC)5 facilitate the B to Z conversion, with the replacement exhibiting a somewhat more pronounced effect. These results may be rationalized in terms of the number of contigous CG sequences present in an oligomer and the relative inhibitory effects of other dinucleotide sequences. Our results also suggest that some short oligomers with purine at the 5'-end, such as d[A(CG)nT] with n greater than or equal to 2, may likely crystallize as Z conformations.

DNA↗
PubMed1988

Kinetic and equilibrium binding studies of actinomycin D with some d(TGCA)-containing dodecamers.

Comparative kinetic, melting, and equilibrium binding studies of actinomycin D (ACTD) with d(ATATACGTATAT), four d(TGCA)-containing dodecamers, and poly(dG-dC).poly(dG-dC) revealed that (1) the affinity of ACTD for the dC-dG sequence is much less than for the dG-dC sequence; (2) ACTD forms 1:1 and 2:1 drug-duplex complexes with d(TATATGCATATA) and d(TATGCATGCATA), respectively, and their SDS driven dissociations exhibit single-exponential characteristics with rates (approximately 5 X 10(-4)s-1 at 20 degrees C) slightly slower than that of poly(dG-dC).poly(dG-dC); (3) although the melting temperature of d(CATGCATGCATG) is 8-9 deg higher than that of d(TATGCATGCATA), the rates of ACTD dissociation from these two oligomers are not greatly different and binding constants of (1-5) X 10(7) M-1 have been estimated for both; (4) a 3:1 stoichiometry is exhibited by ACTD binding to duplex d(TGCATGCATGCA) and the complex dissociates with two characteristic times, the fast component (1/k = approximately 100 s) comprising 2/3 of the contribution and the slow process (approximately 2000 s) contributing the other 1/3; and (5) the slow dissociation kinetics of an oligomer appears to be correlated to the higher percentage of slow association kinetics detectable by non-stop-flow techniques. These results indicate that the d(TGCA) sequence is a stronger binding and a slower dissociation site than the d(CGCG) sequence and suggest that base pairs flanking the dG-dC intercalative site may modulate interactions of the pentapeptide rings of ACTD with the DNA minor groove.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1988

Translaryngeal guided intubation solved a critical airway problem.

Airway obstruction during the induction of general anesthesia remains a persistent problem in modern anesthesia practice, particularly in obstetric patients. Generally, a careful preoperative airway evaluation uncovers most abnormalities that might make intubation difficult. The planning and preparation for additional intubation equipment facilitate an anticipated difficult intubation. However, situations may arise in which unanticipated airway obstruction occurs requiring prompt management. Although every anesthesia provider is trained to manage such acute airway problems, the provision of a patent airway is not always possible, particularly when repeated attempts at endoscopic or blind intubation have failed, leaving a bloody field that prevents optimal visualization, or when time does not allow to wake up the patient. In this article a difficult airway problem is reported in which translaryngeal guided intubation was lifesaving.

Abruptio Placentae↗
PubMed1987

Denaturation and renaturation studies of benzo[a]pyrene metabolite modified DNAs.

Evidence from absorbance, fluorescence, and circular dichroism (CD) measurements strongly suggests that adduct conformations at the binding sites are grossly different before and after thermal denaturation of (+)-trans-7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]py ren e [(+)-anti-BPDE] modified DNAs. This conclusion is reached through the following observations: (1) upon melting and cooling, the (+)-anti-BPDE-modified DNA exhibits pronounced hypochromic effects with concomitant spectral red shifts for the pyrenyl absorbance; (2) the pyrenyl CD spectrum reverses sign upon thermal denaturation-renaturation; (3) the fluorescence emission spectra resulting from excitations at 353 nm (10 nm to the red of hydrolyzed and unbound anti-BPDE) exhibit enhanced intensities and spectral red shifts for the thermally denatured and cooled adducts; and (4) in contrast to the absence of a shoulder prior to melting, the postmelt adducts exhibit a prominent 355-nm maximum (evidence of stacking interactions) in the excitation spectrum when 384-387-nm emission is monitored. Studies with synthetic polynucleotides further reveal that (+)-anti-BPDE-modified poly(dG).poly(dC) exhibits the greatest nonreversible renaturation at the binding sites, possibly as a consequence of pyrenyl self-stacking. This, coupled with the previous findings that this polymer suffers the most extensive (+)-anti-BPDE modification, appears to suggest that (dG)n . (dC)n regions may be responsible for such observed effects in native DNA.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed1987

Two new monoclonal antibodies, Lym-1 and Lym-2, reactive with human B-lymphocytes and derived tumors, with immunodiagnostic and immunotherapeutic potential.

Two new monoclonal antibodies (Lym-1 and Lym-2), reactive with the cell surface of B-lymphocytes and derived tumors, have been produced using tumor cell nuclei preparations as immunogens. Specificity screens using live cell radioimmunoassay techniques with 52 well-characterized human lymphoma and leukemia cell lines showed that both Lym-1 and Lym-2 bound to cell lines of B-cell lineage but were unreactive with those of T-cell, myeloid, or erythroid derivation. The B-cell specificity of these reagents was confirmed on 36 lymphoma and 15 leukemia biopsy specimens by using immunoperoxidase or immunofluorescence techniques. Additionally, flow cytometric analysis of 22 lymphoma biopsies showed that the majority of B-cell tumors were Lym-1 and/or Lym-2 positive and that within a given biopsy, a high percentage of the malignant cell population was stained. In both the immunoperoxidase and flow cytometric studies, reactive T-cells or T-cell lymphomas were consistently negative with the exception of Hodgkin's disease tissues which, in some instances, showed a higher than expected positivity with Lym-1 and Lym-2. Approximately 40% of B-cell chronic lymphocytic leukemias were found to be positive with Lym-1 while 80% were positive with Lym-2. Immunoperoxidase staining of frozen sections of human lymphoid tissues showed that both Lym-1 and Lym-2 stained germinal center and mantle zone B-lymphocytes as well as interfollicular histiocytes. Flow cytometric analysis of normal peripheral blood demonstrated specific staining of B-cells which comprised approximately 8% of circulating lymphocytes. Immunoperoxidase staining of nonlymphoid human organs and tissues revealed weak reactivity of Lym-1 with surface colonic epithelium only. Consistent with these findings, 35 solid tumor cell lines of diverse nature were found unreactive with both Lym-1 and Lym-2. Although standard techniques have thus far failed to identify the antigen recognized by Lym-2, the membrane antigen which binds Lym-1 has been shown by immunoprecipitation and competitive radioimmunoassay studies to be a polymorphic variant of the HLA-Dr antigen. Solid-phase radioimmunoassay techniques have shown that the antigens recognized by Lym-1 and Lym-2 are not significantly modulated after antibody exposure nor shed into the circulation of lymphoma patients. Finally, using iodine-125 labeled preparations of purified Lym-1 and Lym-2, we have determined that both reagents have a relatively large number of antibody binding sites per tumor cell and increased avidity for lymphoma cells when compared to normal and reactive lymph node B-cells.(ABSTRACT TRUNCATED AT 400 WORDS)

Antibodies, Monoclonal↗
PubMed1987

Blood gas studies in dogs with severe steam inhalation injury.

The dynamic changes in blood gases in 34 dogs suffering severe steam inhalation injury were studied during the first 14 days post-burn to determine the effects of inhalation injury on lung function. Nineteen dogs received only severe steam inhalation injury, while 15 other dogs received both 30 per cent total body surface area (TBSA) deep burns and steam inhalation injury. The results showed marked reductions of PaO2, PaCO2, AB, SBE, T-CO2, SB, BBp, pHNR and SatO2 (P less than 0.05 approximately 0.01), whereas the P(A-a)O2 increased (P less than 0.05 approximately 0.01) within week 1 post-burn in both groups. These data suggest that the injured animals were suffering from hypoxaemia and respiratory alkalosis induced by disturbance of the gas-exchange function of the lungs. There was also metabolic acidosis, which was closely related to high fever, excessive catabolism, and particularly to hypoxaemia in week 1 post-burn. If the animals survived through the critical period (1-7 days post-burn), most of the abnormal gas parameters then gradually returned to the pre-burn levels. It is clear, therefore, that blood gas monitoring in patients with inhalation injury is not only of diagnostic importance but also of prognostic value.

Acidosis, Respiratory↗
PubMed1987

1H n.m.r. long-range coupling in 2,4-disubstituted-5(4H)-oxazolones and 4-alkyl-5(2H)-oxazolones generated therefrom by the action of triethylamine.

Long-range couplings were observed between H-4 and 2-CCHn of 2,4-disubstituted-5(4H)-oxazolones, and H-4 and H-2 of 4-alkyl-5(4H)-oxazolones. In the presence of triethylamine, H-4 of the latter migrates to C-2 accompanied by a shift of the double bond to give 4-alkyl-5(2H)-oxazolones which show 5J coupling between H2-2 and 4-CCHn protons.

Ethylamines↗
PubMed1986

Corticotropin releasing factor receptor-mediated stimulation of adenylate cyclase activity in the rat brain.

Corticotropin releasing factor (CRF)-stimulated adenylate cyclase activity and receptor binding were examined in rat brain homogenates using a potent synthetic CRF analog--[D-Tyr3,D-Pro4,Nle18,21,alpha-helical]CRF3-41 (alpha-hel CRF3-41). Binding of alpha-hel CRF3-41 in the rat brain was saturable, reversible, of high affinity and exhibited relevant peptide specificity. This analog also stimulated adenylate cyclase activity of various brain regions; the greatest magnitude of stimulation was in the cerebral cortex followed by the septum, cerebellum and thalamus. Adenylate cyclase stimulation in the cerebral cortex was concentration-dependent with an ED50 of 2.5 +/- 0.4 nM for alpha-hel CRF3-41 and an ED50 of 16 +/- 2 nM for ovine and rat CRF. Maximal stimulation was comparable for all peptides. Agonist-stimulated adenylate cyclase activity was competitively blocked by the CRF antagonists. The inactive CRF analog, ovine CRF1-39, at concentrations less than 1 microM, did not significantly stimulate adenylate cyclase. Adrenalectomy, which has been reported to modulate CRF receptor number and CRF-stimulated adenylate cyclase activity in the anterior pituitary, had no effect on CRF receptor binding or CRF-stimulated adenylate cyclase activity in the cerebral cortex. These results suggest that, as in the anterior pituitary, at least some of the physiological responses mediated by CRF receptors in the brain utilize the cyclic nucleotide regulatory pathway as a post-receptor mechanism.

Adenylyl Cyclases↗
PubMed1986

Conformational lability of poly(dG-m5dC):poly(dG-m5dC).

The remarkable conformational lability of poly(dG-m5dC):poly(dG-m5dC) is demonstrated by the observation of an acid-mediated conformational hysteresis. An acid-mediated Z conformation that exists in solutions containing low sodium concentrations that would normally favor the B conformation is described in this report. This Z conformation is reached by an acid-base titration of a B-poly(dG-m5dC):poly(dG-m5dC) solution which is not far from the B-Z transition midpoint. The resulting Z conformation is thermally very stable, with direct melting into single strands at approximately 100 degrees C. In contrast, the B form DNA, initially in solutions of the same ionic strength but without exposure to acidic pH, exhibits a biphasic melting profile, with conversion into the Z form (with high cooperativity) prior to an eventual denaturation into single strands at around 100 degrees C. Cooling experiments reveal that such biphasic transitions are quite reversible. The transition midpoint for the thermally poised B to Z transformation depends strongly on the NaCl concentration and varies with sample batch. The acid-mediated Z form binds ethidium more weakly than its B counterpart, and the ethidium induced Z to B conversion occurs in a step-wise (non-allosteric) fashion without the requirement of a threshold concentration. The acid-mediated as well as the thermally poised Z conformations are reversed by the addition of EDTA, suggesting the involvement of trace amounts of multivalent metal ions.

DNA↗