High-dose chemotherapy with autologous hematopoietic stem-cell support compared with standard-dose chemotherapy in breast cancer patients with 10 or more positive lymph nodes: first results of a randomized trial.

PURPOSE: Investigation of high-dose chemotherapy (HD-CT) followed by autologous hematopoietic stem-cell support compared with standard-dose chemotherapy (SD-CT) as adjuvant treatment in patients with primary breast cancer and 10 or more positive axillary lymph nodes. PATIENTS AND METHODS: Between November 1993 and September 2000, 307 patients were randomized to receive (following four cycles of epirubicin 90 mg/m(2) and cyclophosphamide 600 mg/m(2), intravenously every 21 days) either HD-CT of cyclophosphamide 1500 mg/m(2), thiotepa 150 mg/m(2), and mitoxantrone 10 mg/m(2), intravenously for 4 consecutive days followed by stem-cell support; or SD-CT in three cycles of cyclophosphamide 500 mg/m(2), methotrexate 40 mg/m(2), and fluorouracil 600 mg/m(2) intravenously on days 1 and 8, every 28 days. The primary end point was event-free survival. RESULTS: After a median follow-up of 3.8 years, 144 events with respect to event-free survival have been observed (HD-CT: 63 events; SD-CT: 81 events). The first event of failure (HD-CT v SD-CT) was an isolated locoregional recurrence (nine v 11), a distant failure (52 v 68), and death without recurrence (two v two). The estimated relative risk of HD-CT versus SD-CT was 0.75 (95% CI, 0.54 to 1.06; P =.095). Overall survival showed no difference (HD-CT: 40 deaths; SD-CT: 49 deaths). CONCLUSION: There was a trend in favor of HD-CT with respect to event-free survival, but without statistical significance. Further follow-up and a meta-analysis of all randomized studies will reveal the effect of HD-CT as compared with SD-CT as adjuvant treatment in high-risk primary breast cancer.

The moss Physcomitrella patens releases a tetracyclic diterpene.

The presence of the tetracyclic diterpene 16alpha-hydroxykaurane (16alpha-hydroxy-ent-kaurane, C20H34O, CAS 5524-17-4) was detected in sterile cell cultures of the moss Physcomitrella patens (Hedw.) B.S.G. using gas chromatography and mass spectrometry. 16alpha-hydroxykaurane was found to be a major lipid compound in P. patens, with an estimated intracellular concentration of up to 0.84 mmol/l and an extracellular concentration of up to 9.3 micromol/l. The overall content of 16alpha-hydroxykaurane (in milligrams) produced per culture reached 0.37-fold that of chlorophyll a+b. In agar cultures with low air exchange, 16alpha-hydroxykaurane forms needle-like crystals on tissue and on the inner surface of the culture vessels, indicating that it is being released into the atmosphere. Solid phase microextraction confirmed the air-bound release of 16alpha-hydroxykaurane. To our knowledge this is the first report on the release of a plant-derived tetracyclic diterpene into the air.

Chemotherapy for mobilisation of Ph-negative progenitor cells from patients with CML: impact of different mobilisation regimens.

Mobilised peripheral blood stem cells are widely used for autografting in patients with chronic myeloid leukaemia (CML) and it is generally thought that a high proportion of Ph-negative progenitor cells in the graft is desirable. We report here the results of 91 stem cell mobilisations performed with various chemotherapy regimens followed by G-CSF. We show that mobilisation of Ph-negative cells is possible after diagnosis as well as in advanced stages of the disease. The yield of Ph-negative cells is highly dependent on the chemotherapy regimen: while the combination of idarubicin and cytarabin for 3-5 days (IC3-5) mobilised Ph-negative cells in most patients, high-dose cyclophosphamide was ineffective. Mobilisation of Ph-negative progenitor cells after IC3 was at least as effective as after IC5; however, less apheresis sessions were required, and toxicity was much reduced after IC3. Compared to historical controls, IC was equally effective as the widely used ICE/miniICE (idarubicin, cytarabin, etoposide) protocol. No correlation was found between graft quality and the cytogenetic response to subsequent treatment with interferon-alpha. We conclude that IC3 is an effective and well-tolerated regimen for mobilising Ph-negative cells that compares well with more aggressive approaches such as IC5 and ICE/miniICE.

Semaphorin4F interacts with the synapse-associated protein SAP90/PSD-95.

Semaphorins are a family of secreted and membrane-associated proteins involved in growth cone guidance during development. Here, we describe the interaction of Semaphorin4F (Sema4F) with the post-synaptic density protein SAP90/PSD-95. Using the yeast two-hybrid system and coprecipitation assays we were able to show an interaction between the extreme C-terminus of Sema4F and the PDZ domains of SAP90/PSD-95. Heterologous coexpression of a chimeric EphrinB1/Semaphorin4F protein with SAP90/PSD-95 in COS cells leads to translocation of SAP90/PSD-95 from the cytosol to the membrane. Deletion analysis shows that this translocation activity of Sema4F is completely dependent on the presence of the last three C-terminal amino acids. In addition, Sema4F immunoreactivity is present in synaptosome fractions and enriched in post-synaptic density fractions. Consistently, in cultured hippocampal neurons, we demonstrate punctate colocalization of Sema4F and SAP90/PSD-95 in dendrites, furthermore we found colocalization of Sema4F with synapsin1 suggesting a synaptic localization. Our data implicate a new functional context for semaphorins at glutamatergic synapses.

[Antifungal susceptibility testing in chronically recurrent vaginal candidosis as basis for effective therapy]. / Resistenzbestimmungen bei Erregern chronisch rezidivierender Vaginalcandidosen als Voraussetzung für eine effektive Therapie.

The chronically recidivist vulvo-vaginal candidiasis is one of the most stubborn problematic diagnosis in the dermatology and gynaecology ward. Prognosis and therapy are primarily determined by the causative micro-organism and the interaction of the fungal species with the currently available antifungal agents. Objective of the study was the investigation of vaginal yeast isolates from patients with chronically recidivist vaginal candidiasis against 8 antifungal agents with the aim of optimising the standard therapy with azole antifungal agents and assessment of alternative therapy schemes. 55 clinical isolates (Dermatology, Charité) of 40 patients were tested by microdilution according to DIN 58940-84. Species differentiation and identification was performed by Fourier-Transform Infrared Spectroscopy (FTIR). In the result Candida glabrata was the predominant causative agent for the recidivist vaginal candidiasis. MIC-mode values for C. glabrata were: fluconacole 32 micrograms/ml, itraconacole 1 microgram/ml, ketoconacole 1 microgram/ml, amphotericine B, voriconacole 0.03 microgram/ml, amphotericin B 0.5 microgram/ml, terbinafine 128 micrograms/ml, cicloproxolamine 4 micrograms/ml, 5-fluorocytosine 0.03 microgram/ml. Some strains of Patients with suboptimal introductory low doses of fluconacole showed increasing of MIC in course of therapy. Parallel resistance with itraconacole was observed in all these cases. Consecutively isolated strains could be clearly and reliably identified by FTIR. In conclusion of most importance is the initial dose adapatation of the drug used, e.g. for fluconacole 800/d p.o., when C. glabrata is the causative agent. Low dose fluconacole therapy is always unsuccessful in recurrent vaginal candidiasis and induces secondary resistance. Demonstrated high susceptibility of voriconacole, amphotericine B an 5-fluorocytosine particularly for C. glabrata may indicate of an anitmycotic therapy potential unconsidered regarding to dermatological indication up to now.

[Deep venous thrombosis and neoplastic pathology: our experience in emergencies]. / Trombosi venose profonde e patologia neoplastica: nostro contributo in urgenza.

Deep vein thrombosis incidence is 1/1000 per year; it is associated with many risk factors which is considered as "thrombophilic states". Its pathogenesis is complex, caused by alterations of hemostasis system. Many studies have established the relation between cancer and subsequent venous thromboembolism, confirming the relationship of neoplastic cell interaction with coagulation system. Forty-seven patients admitted to the hospital from 1987 to 1996 with symptomatic clinically proved deep vein thrombosis were included in a retrospective study. Routine examination at the time of diagnosis of deep vein thrombosis revealed an occult cancer in 8 out of 47 patients; 9 out of 47 patients were admitted in hospital with vein thrombosis and known cancer. The aim of this study is to suggest the best, first treatment of vein thromboembolism in emergency to avoid the dangerous pulmonary embolism complication. The patients affected with deep vein thrombosis and cancer were elderly (over 70 years old, in mean); the neoplasia was of digestive system (8/17) in advanced metastatic stage there was cancer familiarity in 7 out of 47 patients. The high risk of pulmonary embolism associated to deep vein thrombosis suggests the importance of early starting the anticoagulant therapy and placing caval filter.

Volatile constituents of the dried leaves of Cassia angustifolia and C. acutifolia (Sennae folium).

The official drug Sennae folium (obtained from Cassia angustifolia and/or C. acutifolia) was found to contain small amounts of volatiles (0.047% based on dry weight) which were analyzed in detail by GC and GC/MS. More than 200 compounds could be detected, 122 of them were identified, representing about 90.7% of the peak area of the total mixture. The volatile constituents can be classified into monoterpenoids (8.8% for sample A and 34.6% for B), sesquiterpenoids (4.2% and 4.0%, respectively), phenylpropanoids (4.2%/15.2%), fatty acids and esters (54.3%/14.2%), and miscellaneous compounds (19.3%/22.7%). Apart from hexadecanoic acid which was strong in both samples (36.8%/9.7%), the occurrence of menthol, geranylacetone, and (E)-anethole is of interest.

Composition of the essential oil of the flower heads of Matricaria perforata.

The essential oil of the flower heads of scentless mayweed (M. perforata Mérat) was investigated. It consists of more than 50 compounds; 26 of them, together representing about 95.2% of the total oil, were identified by GC, GC/MS, and partly by 1H-NMR and UV. They can be classified into polyacetylenes (88.8%), sesquiterpenoids (5.8%), monoterpenoids (0.1%), fatty acids (0.3%), and miscellaneous compounds (5.0%). The oil was dominated by 2Z,8Z-matricaria ester (ME; 75.2%), which was accompanied by several other characteristic polyines, occurring in much lower concentrations: e.g. 8Z-2,3-dihydro-ME, 2E- and 2Z-lachnophyllum ester, 2E-dehydro-ME, and 5E,9Z-matricaria lactone.

The severe combined immunodeficient-human peripheral blood stem cell (SCID-huPBSC) mouse: a xenotransplant model for huPBSC-initiated hematopoiesis.

Mononuclear cells (MNCs) containing peripheral blood stem cells (PBSCs) were obtained from solid-tumor patients undergoing mobilizing chemotherapy followed by granulocyte colony-stimulating factor for PBSC transplantation-supported dose-intensified anticancer chemotherapy and were transplanted into unconditioned "nonleaky" young severe combined immunodeficient mice. Multilineage engraftment was shown by flow cytometry and immunocytochemistry using monoclonal antibodies to various human cell surface antigens as well as identification of human immunoglobulin in murine sera. Within a dose range of MNCs suitable for transplantation (10 to 36 x 10(6) cells/graft) the number of CD34+ cells injected (optimal at > 0.7 x 10(6)/graft) determined the yield of human cells produced in recipient animals. Engraftment of hu PBSC preparations resulted in prolonged generation of physiologic levels of human cytokines including interleukin-3 (IL-3), IL-6, and granulocyte-macrophage colony-stimulating factor, which were detectable in the murine blood over a period of at least 4 months. In vivo survival of immature human progenitor cells was preserved even 9 months after transplantation. Because human IL-3 is known to stimulate early hematopoiesis, a rat fibroblast cell line was stably transfected with a retroviral vector carrying the human IL-3 gene and cotransplanted subcutaneously as additional source of growth factor. Cotransplants of this cell line producing sustained in vivo levels of circulating human IL-3 for at least 12 weeks significantly accelerated the process of engraftment of huPBSC and spurred the spread of mature human cells to the murine spleen, liver, thymus, and peripheral blood. Cotransplants of allogeneic human bone marrow stromal cells derived from long-term cultures resulted in a comparable--though less prominent--support of engraftment.

[Large volume lymphocytapheresis for the collection of peripheral stem cells]. / Grossvolumige Lymphozytapherese zur Sammlung peripherer Stammzellen.

We performed 29 large-volume leukaphereses of 20 patients for collection of peripheral blood stem cells. All patients have been pretreated with cytokines after chemotherapy. In 9 patients with precounts of > or = 3 x 10(9) mononuclear cells/l we achieved a sufficient transplantation doses with one LVL. If the MNC precount was < or = 3 x 10(9)/l we had to perform more than one LVL. On 16 patients we compared a standard apheresis procedure with the LVL procedure. It seems that especially patients with a lower MNC precount can profit from LVL. From patients with higher MNC precounts we harvested the double amount of MNCs, from patients with lower MNC precounts the triple amount.

[Stem cell pheresis and deep temperature preservation--a problem of transfusion medicine?]. / Stammzellapherese und -tieftemperaturkonservierung--eine Aufgabe der Transfusionsmedizin?

In Germany, Transfusion Medicine belongs to various medical disciplines. We would like to demonstrate our model of interdisciplinary collaboration in stem cell transplantation. Since 1989 we performed 339 leukaphereses in 53 patients. On average we got 1.4 x 10(8) MNC/kg body weight from one single apheresis procedure. To observe graft quality we measured the content of CD34-positive cells and the amount of CFU-GM. Despite diagnosis and therapeutical regimen the content of CD34+ cells was found to be 3.3% (0-20). The proliferation was 21.7 GM-CFU per 1 x 10(5) MNC seeded (0-393). The results of the first 17 transplantations showed a fast haematological recovery (WBS 1.0 Gpt/l after 9 days Plt 20 Gpt/l after 11 days). Close collaboration of Transfusion Medicine and Haematology leads to optimization of stem cell transplantation.

Radiotelemetry temperature responses of mammary gland and body to intramammary injection of Escherichia coli endotoxin or Streptococcus agalactiae in lactating dairy cows.

To investigate the feasibility of using changes in body or mammary temperature to detect mastitis, radiotransmitters were implanted midway between rear udder quarters and in the peritoneal cavity of 5 Holstein cows (1 to 3 months in lactation) housed in an environmental chamber (16 +/- 2 C; lights on 7:00 AM to 11:00 PM). After a 6-week control period, Escherichia coli endotoxin (0.5 mg) was injected after the morning milking into left rear teat cisterns via the teat canal. Wisconsin mastitis test score and somatic cell count in all quarters increased significantly (P < 0.01) by the next milking. Effects were greatest in the endotoxin-exposed quarters. Milk yields for all quarters decreased significantly (P < 0.01) by the first milking after endotoxin injection. Udder and body temperatures at milkings were similar and were not affected by treatment. When temperatures were averaged for the 5 cows for each of 120 time points/d, average temperatures, relative to time of injection of endotoxin, were increased by 0.5 C above baseline at 2.75 hours, peaked at +2.9 C at 6.50 hours, and remained high through 9.25 hours after injection. Power spectra calculated for individual cows on a daily basis universally indicated an increase in power at low frequencies on the day of injection. Subsequently, Streptococcus agalactiae (200 colony-forming units) was injected into right rear teat cisterns. Wisconsin mastitis test score increased at the second milking after injection. Cell count and quarter milk yield decreased by the third milking. As with endotoxin, injection of S agalactiae could not be detected via a change in temperature at milkings.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of milking without pulsation on teat duct colonization with Streptococcus agalactiae and penetrability to endotoxin.

Forty quarters of 10 cows were milked for a 9-d period with one of four treatments: 1) no liner pulsation, 2) conventional milking, 3) no pulsation for 4 d followed by conventional milking for 5 d, and 4) conventional milking for 4 d followed by milking without pulsation for 5 d. All teat orifices were inoculated with approximately .5 million cfu of Streptococcus agalactiae and Streptococcus dysgalactiae on d 1 and 5. Recoveries of Strep. agalactiae from the teat end were increased for teats milked without pulsation. Recoveries of Strep. dysgalactiae were lower than those of Strep. agalactiae and not increased by milking without pulsation. In a second experiment, teats of 20 cows were milked for a 15-d period with or without liner pulsation. For 10 successive milkings, all teats were inoculated with 1.0 micrograms of Escherichia coli endotoxin either immediately or 2.5 h after each milking. The frequency of endotoxin penetration, measured by the Wisconsin Mastitis Test, in pulsated quarters and in nonpulsated quarters was similar. For quarters milked without pulsation but not for pulsated quarters, inoculation of endotoxin immediately after milking led to greater incidence of teat duct penetration than for inoculation 2.5 h after milking.

Volatile Constituents of Peucedanum palustre.

The essential oils of fruits, stalks, leaves, and roots of PEUCEDANUM PALUSTRE (L.) Moench (Apiaceae) were analyzed by various chromatographic and spectroscopic methods. While the fruit and stalk oils consist nearly exclusively of monoterpene hydrocarbons, the essential leaf oil comprises monoterpene hydrocarbons (47%), sesquiter-pene hydrocarbons (38%), aliphatic alcohols and esters (7%), and traces of oxygenated sesquiterpenoids. 35 constituents of this oil could be identified. The detailed analysis of the complex root oil, mainly consisting of mono- and sesquiterpenoids, resulted in identification of 68 compounds, representing about 92% of the oil. The occurrence of some new sesquiterpenoids with unusual carbon skeletons is noteworthy.

Effect of abraded intramammary device on outcome in lactating cows after challenge exposure with Streptococcus uberis.

Intramammary devices (IMD) were abraded with medium-grade emery cloth or were left smooth. One IMD of each type was inserted into a mammary quarter of each of 5 lactating cows. The remaining 2 quarters served as controls. Quarter foremilk, bucket milk, and stripping milk samples were collected for 3 consecutive days at 2 weeks after IMD insertion, and milk somatic cell counts (SCC) were determined. Milk samples also were collected immediately after and 0.5, 1, 2, 4, 6, 8, and 11 hours after milking. All quarters were challenge exposed with 250 colony-forming units of Streptococcus uberis at 2 months after IMD insertion. Foremilk and stripping milk samples were collected for bacteriologic culture and SCC at the next 10 milkings. Mean foremilk, bucket milk, and stripping milk SCC (X 10(6) cells/ml) were 0.18, 0.07, and 0.91, respectively, for quarters with abraded IMD; 0.06, 0.05, and 0.43, respectively, for quarters with smooth IMD; and 0.03, 0.03, and 0.15, respectively, for control quarters. Mean SCC after milking (X 10(6) cells/ml) for the various intervals were 0.70, 1.29, 0.70, 0.97, 1.15, 1.17, 0.77, and 0.85 for quarters with abraded IMD; 0.43, 0.62, 0.61, 0.45, 0.64, 0.60, 0.31, and 0.26 for quarters with smooth IMD; and 0.15, 0.24, 0.15, 0.19, 0.15, 0.15, 0.14 and 0.06 for control quarters. After challenge exposure, 2 of 5 of the quarters with abraded IMD, 4 of 5 of the quarters with smooth IMD, and 8 of 9 control quarters became infected. Results indicated that abraded IMD increased SCC in stripping milk to concentrations that provided 60% protection against challenge exposure with S uberis.

Effect of intramammary devices on the outcome of induced Escherichia coli infection of bovine mammary quarters.

Eighteen Holstein cows, free of intramammary infection, were fitted with smooth (n = 9) or abraded (n = 9) intramammary devices (IMD) in 2 diagonally opposed quarters within 4 weeks after calving. The 2 other quarters of each cow were used as controls. Three to 6 weeks after IMD insertion, depending on when milk somatic cell counts returned to a base-line value of less than 4 X 10(5)/ml, all cows were subjected to bacterial challenge exposure in the front or rear quarters by intracisternal injection of about 30 colony-forming units of Escherichia coli/quarter. Challenge exposure was done immediately after milking. Three weeks after the initial bacterial exposure, the other quarter pairs were similarly challenge exposed. Quarter bacteriologic status, concentration of milk somatic cells, and clinical observations (rectal temperature, milk appearance, udder palpation, and general condition of the cow) were monitored. Infection developed in 14 of 16 (88%) quarters with smooth IMD vs 16 of 16 (100%) control quarters and in 7 of 17 (41%) quarters with abraded IMD vs 17 of 17 (100%) control quarters. The difference in infection frequency between quarters with smooth IMD and quarters with abraded IMD was significant (P less than 0.05). Protection against establishment of infection was associated with somatic cell counts greater than 8.0 X 10(5)/ml in milk collected immediately after milking (7 of 12 quarters) or 4 hours later (11 of 12 quarters). In 10 quarters (59%) of cows fitted with abraded IMD, secretory abnormalities appeared before bacterial challenge inoculation. Abnormal milk or visible blood was observed over periods varying from 2 weeks after insertion through the entire lactation.