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The water extract of killed Penicillium chrysogenum(Pen) was used to induce resistance against Phytophthora infestansin tomato plants. After two treatments with Pen as foliar spray, a protection of over 90% was achieved compared to control plants. The protective effect was shown under controlled conditions with two assays, whole plants and on leaf disk. The achieved resistance was proportional to the content of total carbohydrate, assuming sugars to be involved as inducing compounds. The extract had no direct antifungal activity on in vitrogrowth of P. i n f e s t a n s, suggesting that disease control resulted from the induction of natural defense mechanisms in the tomato plants. The induction of resistance was accompanied by an activity increase of peroxidase enzymes. After separation of soluble, ionically cell wall-bound extracellular and cytosolic fractions, only the extracellular soluble fraction contained at least two isoenzymes specifically inducible by Pen and corresponding to the achieved resistance. Furthermore, application of Pen led to an acidification of the intercellular fluid in the leaf tissue. The intercellular acidification was closely associated with the achieved resistance indicating a participation in the resistance inducing process. The acidification of the intercellular space introduces a possibly new mode of action of resistance induction. Der wässrige Extrakt von abgetötetem Penicillium chrysoge-num(Pen) wurde zur Resistenzinduktion bei Tomaten gegen Phytophthora infestansbenutzt. Nach zwei Sprühapplikationen des Pen-Extrakts auf die Blätter wurde ein Schutz von über 90% im Vergleich zu den Kontrollpflanzen erreicht. Die Schutzwirkung wurde an Ganzpflanzen und Blattscheiben unter Gewächshausbedingungen getestet. Da der Effekt proportional zur Gesamtkohlenhydratkonzentration des Extrakts war, werden Zucker als aktive Komponenten vermutet. Der Extrakt zeigte keine direkte antifungale Aktivität auf das Wachstum von P. infestans. Das lässt auf eine Induktion pflanzlicher Abwehrmechanismen schließen. Die Resistenzinduktion wurde von einer Steigerung der Peroxidaseaktivität begleitet. Nach einer Trennung in extrazellulär lösliche, extrazellulär zellwandgebundene und cytoplasmatische Isoenzyme zeigte sich, dass ausschließlich in der extrazellulär löslichen Fraktion zwei Isoenzyme spezifisch durch die Behandlung mit Pen induziert wurden und ihre Aktivität proportional zum erreichten Resistenzgrad war. Weiterhin führte die Applikation von Pen zu einer starken Ansäuerung der Interzellularflüssigkeit des Blattgewebes. Die Ansäuerung des Interzellularraumes war eng mit dem erreichten Resistenzgrad verknüpft und scheint ein neuer Wirkmechanismus der Resistenzinduktion zu sein.

We have recently proposed a macromolecular prodrug strategy for improved cancer chemotherapy based on two features (Kratz, F.; et al. J. Med. Chem 2000, 43, 1253−1256.):  (a) rapid and selective binding of thiol-reactive prodrugs to the cysteine-34 position of endogenous albumin after intravenous administration and (b) release of the albumin-bound drug in the acidic environment at the tumor site due to the incorporation of an acid-sensitive bond between the drug and the carrier. To investigate this therapeutic strategy in greater depth, four (maleinimidoalkanoyl)hydrazone derivatives of doxorubicin were synthesized differing in the length of the aliphatic spacer (1, −(CH2)2−; 2, −(CH2)3−; 3, −(CH2)5−; 4, −(CH2)7−). The albumin-binding doxorubicin prodrugs, especially the (6-maleimidocaproyl)hydrazone derivative of doxorubicin (3), are rapidly and selectively bound to the cysteine-34 position of endogenous albumin. 3 was distinctly superior to the parent compound doxorubicin in three animal tumor models (RENCA, MDA-MB 435, and MCF-7) with respect to antitumor efficacy and toxicity.

Poly(ethylene glycol)s (PEGs) are potential drug carriers for improving the therapeutic index of anticancer agents. In this work, the anticancer drug methotrexate (MTX) was activated with N,N‘-dicyclohexylcarbodiimide (DCC) and coupled to amino group bearing PEGs of MW 750, 2000, 5000, 10 000, 20 000, and 40 000. First, the activation process of MTX with DCC in the presence and absence of N-hydroxysuccinimide was analyzed through HPLC. Preincubation of methotrexate with DCC alone at 0 °C proved to be favorable with respect to the amount of activated species and the formation of byproducts. MTX−PEG conjugates were synthesized according to this procedure, isolated through size-exclusion chromatography, and characterized through analytical HPLC, MALDI-TOF spectrometry, and gel permeation chromatography. In a cell-free assay, all of the drug polymer conjugates inhibited the target enzyme of MTX, dihydrofolate reductase (DHFR), to a similar extent, but were not as active as free MTX. Additionally, incubation of the MTX−PEG40000 conjugate for 6 days at 37 °C in phosphate buffered saline (pH 7.4), in cell-conditioned medium, or in human serum revealed no significant release of methotrexate. These results, taken together, indicate that release of MTX from polymer conjugates is not necessary for an effective interaction with the active site of dihydrofolate reductase. Evaluation of the in vitro cytotoxicity of the MTX−PEG conjugates in two adherent and three suspension human tumor cell lines revealed that the IC50 values of the tested compounds increased with the size of the drug−polymer conjugates. The most effective compound tested in these assays was the free drug MTX itself (IC50 value ranging from ~0.01 to 0.05 μM), while the IC50 values of the polymer conjugates were higher (IC50 value for MTX−PEG750, 2000 and 5000:  ~0.6−3 μM; for MTX-PEG10000 and 20000:  ~2−7 μM; and for MTX−PEG40000:  > 6 μM). Subsequently, MTX−PEG5000, MTX−PEG20000, and MTX−PEG40000 were evaluated in a human mesothelioma MSTO-211H xenograft model, and their antitumor effects were compared with free methotrexate and the albumin conjugate MTX−HSA, a conjugate that is currently in phase II clinical trials. In contrast to the in vitro results, the high molecular weight MTX−PEG conjugates exhibited the highest in vivo antitumor activity:  At a dose of 40 and 80 mg/kg MTX−PEG5000 was less active than MTX at its optimal dose of 100 mg/kg; MTX−PEG20000 at a dose of 40 mg/kg showed antitumor efficacy comparable to MTX, but MTX−PEG40000 at a dose of 20 mg/kg was superior to MTX and demonstrated antitumor activity of the same order as MTX−HSA (20 mg/kg).

Collagen-induced arthritis (CIA) is a widely used model for rheumatoid arthritis. Induction of CIA in rats using rat type II collagen (CII) results in a chronic arthritis in which anti-CII antibodies are believed to play a pathogenic role. In this study, we analyzed the epitope selection and V gene usage in the anti-CII response in the DA rat. A panel of CII-reactive B cell hybridomas was established from the draining lymph nodes 11 days after immunization. All of the CII-specific antibodies bound cartilage in vivo, showing that these are true autoantibodies. These antibodies were all IgG and specific for several distinct triple helical epitopes on CII. Interestingly, the major epitope, recognized by four different antibodies, was identical with the major B cell epitope in the mouse CII located at position 359-369 (denoted as C1^III). The Q52 and PC7183 V_H gene families encoded 12 out of 14 sequenced heavy chains. There was a relatively more heterogeneous usage of V_L genes as the antibodies were encoded by four different V_κ families (V_κ1, V_κ2, V_κ12/13 and V_κRF). As in the mouse, some of the V genes used showed germline characteristics. We conclude that the immune response in the rat shares epitope specificity and a constrained V gene repertoire with the mouse. However, the V genes used for recognition of the closely related collagen structures differed considerably between mouse and rat, indicating an influence of the species-specific variation in the V gene repertoire.

The aim of the study was to evaluate the efficacy, tolerability and quality of life in 5‐fluorouracil (5‐FU) pretreated colorectal cancer patients after combined 5‐FU and Ginkgo bilobaextract GBE 761 ONC (i.e. the Ginkgo bilobaspecial extract EGb 761®) therapy. Following conventional 5‐FU therapy, 44 patients (32 evaluable for response) with advanced progressive colorectal cancer were treated every 3 weeks with courses of 350 mg GBE 761 ONC as a 30 min i.v. infusion on days 1–6 followed by 500 mg/m2/d 5‐FU as a 30 min i.v. infusion on days 2–6. The response to therapy was evaluated after the second and fourth course of treatment. The data of 32 patients could be evaluated for efficacy. We observed a progression of disease in 22 patients, no change in 8 patients and a partial response in 2 patients (overall response = 6.3%). Seventeen of 22 patients with observed progressive disease showed further progression after two cycles, two after three cycles and three after four cycles. The median survival time was 9.5 months (7.7–11.5 months). GBE 761 ONC was well tolerated. Adverse events that occurred during the study were mainly myelosuppression and gastrointestinal symptoms and were judged to be 5‐FU related or consistent with liver toxicity and thus tumour related. Our results suggest a good benefit–risk ratio of the combined 5‐FU and GBE 761 ONC therapy as second line treatment in metastatic colorectal cancer. The survival time was similar to that known from second line treatment according to the Ardalan scheme. Since an improvement was observed in some patients despite the failure of the conventional 5‐FU pretreatment, it would be interesting to evaluate whether the application of 5‐FU plus GBE 761 ONC as a first line treatment is of benefit. Copyright © 2001 John Wiley & Sons, Ltd.

The aim of the study was to evaluate the efficacy, tolerability and quality of life in 5-fluorouracil (5-FU) pretreated colorectal cancer patients after combined 5-FU and Ginkgo biloba extract GBE 761 ONC (i.e. the Ginkgo biloba special extract EGb 761®) therapy. Following conventional 5-FU therapy, 44 patients (32 evaluable for response) with advanced progressive colorectal cancer were treated every 3 weeks with courses of 350 mg GBE 761 ONC as a 30 min i.v. infusion on days 1–6 followed by 500 mg/m²/d 5-FU as a 30 min i.v. infusion on days 2–6. The response to therapy was evaluated after the second and fourth course of treatment. The data of 32 patients could be evaluated for efficacy. We observed a progression of disease in 22 patients, no change in 8 patients and a partial response in 2 patients (overall response = 6.3%). Seventeen of 22 patients with observed progressive disease showed further progression after two cycles, two after three cycles and three after four cycles. The median survival time was 9.5 months (7.7–11.5 months). GBE 761 ONC was well tolerated. Adverse events that occurred during the study were mainly myelosuppression and gastrointestinal symptoms and were judged to be 5-FU related or consistent with liver toxicity and thus tumour related. Our results suggest a good benefit–risk ratio of the combined 5-FU and GBE 761 ONC therapy as second line treatment in metastatic colorectal cancer. The survival time was similar to that known from second line treatment according to the Ardalan scheme. Since an improvement was observed in some patients despite the failure of the conventional 5-FU pretreatment, it would be interesting to evaluate whether the application of 5-FU plus GBE 761 ONC as a first line treatment is of benefit. Copyright © 2001 John Wiley & Sons, Ltd.

Profiles William Calley, 46, the former Army Lieutenant convicted of killing 22 people during the 1969 massacre in a Vietnamese village called My Lai. Calley now is a respected family man and figure in Columbus, Ga., where residents are sympathetic to him. Personal life and background; My Lai incident; Other soldiers of Charlie Company and their present lives.

Visits No. 19 Gramercy Park in New York City, a 37-room house with a $13-million price tag. For nearly a century, the great house was `the' place to party-with Astors, Rockefellers, and Dylan dropping by. A historical look at both owners and famous guests.

Comments that the Republican party, fearful that President Jimmy Carter might pull on `October surprise' by managing to get the American hostages in Iran home, made certain he couldn't. So charges former Carter aide Gary Sick; Engineered by CIA chief William Casey; The claim that Casey stopped Iran from releasing the hostages until after the election; The charge that then vice-presidential nominee George Bush was involved.

Profile of Sacramento, Calif., Assemblyman Tom Hayden. Breakup of his marriage with actress Jane Fonda; Extramarital affair; Career in politics; Personal history; His political future without Jane.

The present study was undertaken to investigate a possible involvement of phospholipid methyltransferases in the coupling of receptor-mediated stimulation to secretion. Phospholipid methyltransferases were assayed in isolated parotid acini in the presence of carbamoylcholine or isoprenaline. Carbamoylcholine reduced the incorporation of methyl groups into phospholipids, whereas isoprenaline showed no effect. Amylase secretion stimulated either by carbamoylcholine or by isoprenaline could not be affected by inhibitors of methyltransferases (3-deaza-adenosine alone or plus homocysteine thiolactone) under conditions where phospholipid methylation was strongly inhibited. The activity of adenylate cyclase in isolated parotid microsomal membranes was not inhibited or stimulated by S-adenosyl-homocysteine or -methionine respectively. These results indicate that phospholipid methylation does not play an essential role in stimulus-secretion coupling in the parotid gland.

One strategy for improving the selectivity and toxicity profile of antitumor agents is to design drug carrier systems employing soluble macromolecules or carrier proteins. Thus, five maleimide derivatives of chlorambucil were bound to thiolated human serum transferrin which differ in the stability of the chemical link between drug and spacer. The maleimide ester derivatives 1 and 2 were prepared by reacting 2-hydroxyethylmaleimide or 3-maleimidophenol with the carboxyl group of chlorambucil, and the carboxylic hydrazone derivatives 5−7 were obtained through reaction of 2-maleimidoacetaldehyde, 3-maleimidoacetophenone, or 3-maleimidobenzaldehyde with the carboxylic acid hydrazide derivative of chlorambucil. The alkylating activity of transferrin-bound chlorambucil was determined with the aid of 4-(4-nitrobenzyl)pyridine (NBP) demonstrating that on average 3 equivalents were protein-bound. Evaluation of the cytotoxicity of free chlorambucil and the respective transferrin conjugates in the MCF7 mammary carcinoma and MOLT4 leukemia cell line employing a propidium iodide fluorescence assay demonstrated that the conjugates in which chlorambucil was bound to transferrin through non-acid-sensitive linkers, i.e., an ester or benzaldehyde carboxylic hydrazone bond, were not, on the whole, as active as chlorambucil. In contrast, the two conjugates in which chlorambucil was bound to transferrin through acid-sensitive carboxylic hydrazone bonds were as active as or more active than chlorambucil in both cell lines. Especially, the conjugate in which chlorambucil was bound to transferrin through an acetaldehyde carboxylic hydrazone bond exhibited IC50 values which were approximately 3−18-fold lower than those of chlorambucil. Preliminary toxicity studies in mice showed that this conjugate can be administered at higher doses in comparison to unbound chlorambucil. The structure−activity relationships of the transferrin conjugates are discussed with respect to their pH-dependent acid sensitivity, their serum stability, and their cytotoxicity.

The uptake of 14C-labeled 1-(2-chloroethyl)-1-nitroso-3-(2-hydroxyethyl) urea (HECNU) into the brain was investigated in the rat after intracarotid injection according to the method of OLDENDORF, as well as in cisternal cerebrospinal fluid obtained by suboccipital puncture after i.v. injection of the drug.

In collagen-induced arthritis, a murine autoimmune model for rheumatoid arthritis, immunization with native but not heat-denatured cartilage-specific collagen type II (CII) induces a B cell response that largely contributes to arthritogenicity. Previously, we have shown that monoclonal antibodies established from arthritis prone DBA/1 mice require the triple-helical conformation of their epitopes for antigen recognition. Here, we present a novel approach to characterize arthritis-related conformational epitopes by preparing a panel of 130 chimeric collagen X/CII molecules. The insertion of a series of CII cassettes into the triple-helical recombinant collagen X allowed for the first time the identification of five triple-helical immunodominant domains of 5-11 amino acid length, to which 75% of 36 monoclonal antibodies bound. A consensus motif, "R G hydrophobic," was found in all immunodominant epitopes. The antibodies were encoded by a certain combination of V-genes in germline configuration, indicating a role of the consensus motif in V-gene selection. The immunodominant domains are spread over the entire monomeric CII molecule with no apparent order; however, a highly organized arrangement became apparent when the CII molecules were displayed in the quarter-staggered assembly within a fibril. This discrete epitope organization most likely reflects structural constraints that restrict the exposure of CII epitopes on the surface of heterotypically assembled cartilage fibrils. Thus, our data suggest a preimmune B cell selection process that is biased by the accessibility of CII determinants in the intact cartilage tissue.