Phase I clinical and pharmacokinetic trial of dextran conjugated doxorubicin (AD-70, DOX-OXD)

Coupling of anthracyclines to high-molecular-weight carriers may alter drug disposition and improve antitumor effects. We have performed a clinical phase I trial of doxorubicin coupled to dextran (70000 m.w.). The drug was administered as single dose i.v. every 21–28 days. Thirteen patients have received a total of 24 courses (median 2; range 1–3). At the starting dose of 40 mg/m² doxorubicin equivalent (DOXeq), WHO grade IV thrombocytopenia was noted in 2/2 patients. WHO grade IV hepatotoxicity and WHO grade III cardiotoxicity were noted in a patient with preexisting heart disease. Five patients were treated with 12.5 mg/m² DOXeq. Maximal toxicity at this dose level was WHO grade III thrombocytopenia and local phlebitis (WHO grade II) in 1/5 patients, elevation of alkaline phosphatase (WHO grade III) and WHO grade III vomiting in another patient. Subsequently, five patients received 20 mg/m² DOXeq. Hepatotoxicity was noted in 5/5 patients (1 × WHO grade IV, 1 × WHO grade III). Thrombocytopenia was noted in 3/5 patients (1 × WHO grade IV, 2 × WHO grade III). At 12.5 mg/m² DOXeq, a patient diagnosed with a malignant fibrous histiocytoma had stable disease for 4 months. Pharmacokinetic analyses of total and free doxorubicin were performed in plasma and urine. The maximum peak plasma concentration (ppc) for total DOX was 12.3 g/ml at 40 mg/m² DOXeq. The area under the plasma concentration time curve (AUC) ranged from 28.83–80.21 g/ml^*h with dose-dependent elimination half lives (t_1/2: 0.02–0.87 h;_1/2: 2.69–11.58 h;_1/2: 41.44–136.58 h). We conclude that the maximal tolerated dose (MTD) of AD-70 using this schedule is 40 mg/m² DOXeq. The recommended dose for clinical phase II studies is 12.5 mg/m² DOXeq.Abbreviations ALT Alanine Aminotransferase - AST Aspartate Aminotransferase - DOX Doxorubicin - DOXeq Doxorubicin Equivalent - ECG Electrocardiogram - HPLC High Pressure Liquid Chromatography - LD₁₀ Lethal Dose for 10% of individuals - MTD Maximal Tolerated Dose - ppc Peak Plasma Concentration - WHO World Health Organisation     

Immunization with excretory-secretory molecules of intestinal nematodes induces antigen-specific protective memory Th2 cell responses

Parasitic nematodes infect more than 1 billion people in the global south. The development of effective antihelminthic vaccines is a crucial tool for their future elimination. Protective immune responses to nematodes depend on Gata3⁺ Th2 cells, which can also be induced by nematode-released products. Whether these nematode products induce antigen-specific long-lived memory T cells and thereby confer protection against a challenge infection is not known yet. Hence, we set out to characterize the formation of memory Th2 cells induced by immunization with Heligmosomoides polygyrus excretory-secretory (HES) products, infection-induced versus immunization-induced recall responses to a challenge infection, and whether HES-induced memory T cells show protective properties following adoptive transfer. Our results show that 8 weeks postimmunization, HES induces long-lived functional memory Th2 cells at the site of immunization in the peritoneal cavity. Following a H. polygyrus challenge infection, HES-immunized mice display MHC-II-dependent antigen-specific Th2 cytokine responses in the gut-draining lymph nodes, comparable to those induced by a prior natural infection. Moreover, adoptive transfer of sorted memory CD4⁺ T cells from HES-immunized donors reduces female worm fecundity following a challenge H. polygyrus infection in recipient mice, highlighting a protective role for immunization-induced memory T cells.     

The pharmacokinetics of high-dose epirubicin and of the cardioprotector ADR-529 given together with cyclophosphamide, 5-fluorouracil,and tamoxifen in metastatic breast-cancer patients

A high-pressure liquid chromatographic method for determination of the bisdioxopiperazine derivative ADR-529 (ICRF-187), a compound proven effective in protection against anthracycline-induced cardiotoxicity, has been developed. The limit of quantitation was 5 ng/ml using a narrow-bore 5-m silica column and UV detection. The method was used for determination of pharmacokinetic profiles of ADR-529 after a 3-weekly i.v. administration of different doses of ADR-529 (600–1000 mg/m²) together with different doses of epirubicin (E, 60–100 mg/m²), fixed-dose cyclophosphamide (C, 600 mg/m²), fixed-dose 5-fluorouracil (F, 600 mg/m²), and daily administration of tamoxifen (T, 30 mg; CEF-T) in the treatment of patients with metastatic breast cancer. Pharmacokinetic parameters for epirubicin were also determined. The aim of the study was to determine (1) whether the pharmacokinetics of ADR-529 as part of a combination with CEF-T changes with increasing doses of ADR-529 and increasing doses of epirubicin and (2) whether the pharmacokinetics of epirubicin in the same combinations is altered with the administration of increasing doses of ADR-529. A total of 82 patients were included. A crossover study including 16 of the patients showed no significant difference in epirubicin pharmacokinetic parameters when epirubicin was given with or without concomitant administration of ADR-529. Apart from minor changes in the distributional half-lives, the pharmacokinetic parameters of epirubicin were not altered with increasing doses of ADR-529, nor were the pharmacokinetic parameters of ADR-529 itself. Escalating doses of epirubicin did not significantly alter the pharmacokinetic parameters of ADR-529 with the exception of a 30% increase in the terminal half-life and a decrease in total body clearance when the epirubicin dose was raised from 60 to 100 mg/m². We conclude that concomitant administration of ADR-529 does not alter the distribution and elimination of epirubicin in doses suitable for preventing the anthracycline-induced cardiotoxicity.     

Service utilization among homeless and runaway youth in Los Angeles, California: rates and reasons

Purpose: To describe the service utilization patterns of homeless and runaway youth in a “service-rich” area of Los Angeles, California; identify demographic and other correlates of utilization; and contextualize the findings with qualitative data.

Method: During Phase 1 of this study, survey data were collected from an ethnically diverse sample of 296 youth aged 13–23 years, recruited from both service and natural “hang-out” sites using systematic sampling methods. During Phase 2, qualitative data were collected from 46 youth of varying ethnicities and lengths of time homeless.

Results: Drop-in centers and shelters were the most commonly used services (reported by 78% and 40%, respectively). Other services were used less frequently [e.g., medical services (28%), substance abuse treatment (10%) and mental health services (9%)]. Utilization rates differed by ethnicity, length of time in Los Angeles, and city of first homeless episode (Los Angeles versus all others). Shelter use was strongly associated with use of all other services. Despite youths’ generally positive reactions to services, barriers were described including rules perceived to be restrictive, and concerns youth had about confidentiality and mandated reporting. Youth suggested improvements including more targeted services, more long-term services, revised age restrictions, and more and/or better job training and transitional services to get them off the streets.

Conclusions: Because shelters and drop-in centers act as gateways to other services and offer intervention potential for these hard-to-reach youth, it is vital that we understand the perceived barriers to service utilization.     

Concomitant radiochemotherapy with 5-FU and cisplatin for invasive bladder cancer

Purpose

To evaluate acute toxicity and efficacy of simultaneous radiochemotherapy for invasive urothelial cancer of the bladder.

Patients and Methods

From September 1993 to July 1997,61 patients with invasive bladder cancer were treated with a transurethral resection (TURB) followed by radiochemotherapy (RCT). Twenty-five received a combination of 5-FU and cisplatin. The prescribed doses were 600 mg/m² 5-FU daily as continuous infusion over 5 days each in the 1st and 5th treatment week and 20 mg/m² cisplatin daily at the same days as a short infusion. The pelvis was irradiated with 54 Gy, the bladder with 59.4 Gy and the paraortic nodes in 7 cases with 45 Gy, respectively. Six to 8 weeks after RCT a second TURB was performed for reasons of restaging.

Results

Twenty out of 25 patients received at least 80% of the prescribed chemotherapy, in 13 cases the full dose could be given. Gastrointestinal toxicity of Grade I and II occurred in 10 cases, 1 patient developed severe diarrhea (Grade VI). After the 1st course of chemotherapy 7 patients had leucoor thrombopenia of Grade III. One patient had a leucopenia of Grade IV. After the 2nd course 4 patients developed Grade III leuko- and thrombopenia, 1 of Grade IV. Two Grade II anemia were found. All more severe toxicities and necessary dose reductions were related to radiation of the paraaortic nodes. No life threatening infections, bleedings or cardiotoxicity was found. Restaging TURBs resulted in 22 complete remissions, 1 patient had a de-novo-carcinoma (Tis) at this time, 2 were non-responders (8%). After a median follow-up of 38 months 20 patients are alive (80%).

Conclusions

1. If irradiation of paraaortic nodes is necessary, 5-FU should not be applied, because the gastrointestinal toxicity is too extensive. In all other cases side effects are tolerable and can be managed by supportive care. 2. The first results are promising and should be evaluated in a prospective study.     

A novel stopped-flow method for measuring the affinity of actin for myosin head fragments using μg quantities of protein

Summary The dissociation constant for actin binding to myosin and its subfragments (S1 & HMM) is 1 m at physiological ionic strength. Many of the methods used to measure such affinities are unreliable for a K_d below 0.1 m. We show here that the use of phalloidin to stablise F-actin and fluorescently labelled proteins allows the affinity of actin for myosin S1 to be measured in a simple transient kinetic assay. The method can be used for K_d's as low as 10 nm and we demonstrate that the K_d's can be estimated using only g quantities of material. Furthermore we suggest how this method may be adapted for ng quantities of protein. This will allow the affinity of actin for myosin fragments to be estimated for proteins which are difficult to obtain in large quantities i.e. from biopsy material or from proteins expressed in baculovirus.     

Intraocular silicone lenses in silicone oil: an experimental study

Background: To evaluate a potential effect of silicone oil on flexible silicone intraocular lenses, four lenses (STAAR AA-4203) were stored in silicone oil under sterile conditions for periods between 1 month and 3 years. Method: The edge and surface of the lenses were examined by scanning electron micrography and the findings compared with a lens of the same model which had been stored in Ringer's solution for 2 years. Results: After 1 year of silicone oil exposure, droplets of different sizes adherent to the surface of the lens were found. These changes proceeded to a wave-like appearance of the surface after 2 and 3 years of storage, so that a continuous layer of silicone oil polymers is probably covering the intraocular lens. Conclusion: Optical interference has to be considered a possibility if it turns out that the droplets cannot be removed during silicone oil evacuation. Consequently silicone intraocular lenses without hydrophilic preparation of the surface should not be implanted in eyes undergoing combined anterior and posterior segment surgery with silicone oil tamponade or in eyes with high risk for vitreoretinal complications.The authors state that they have no proprietary interest in the marketing of the products mentioned herein or competing products     

Inducible nitric oxide synthase activity of cloned murine microglial cells

Nitric oxide (NO) is a short-lived diffusable molecule now believed to participate in multiple physiologic functions in the CNS including neurotransmission and the maintenance of vascular tone. Previously, we reported that cell lines obtained by retroviral immortalization of tissue macrophages (M?;) could be induced to synthesize nitrite (NO), a stable end product of the NO synthetic pathway. We have further characterized the induction and activity of this pathway in a panel of seven microglial clones derived from primary embryonic mouse brain cultures. Like M?;, these clones were found to release high levels of NO_-² in response to recombinant interferon-γ (rIFN-γ) as a priming signal together with either bacterial lipopolysaccharide (LPS) or exogenous recombinant tumor necrosis factor-α (rTNF-α). As previously demonstrated for M?;, phagocytosis of zymosan particles during induction of enzyme activity enhanced subsequent NO production, which is of interest in light of the postulated phagocytic role of microglia within the CNS. Biochemical characterization of enzyme activity in intact microglial clones and in isolated cytosolic fractions indicates that the microglial NO synthase present in these murine cell clones represents the M?;-like isotype. These findings suggest that microglial cells could represent a major source of NO within the CNS.     

Influence of selective phosphodiesterase inhibitors on human neutrophil functions and levels of cAMP and Cai

Summary Chromatographic analysis of 3,5-cyclic nucleotide phosphodiesterase (PDE) isoenzymes in the cytosol of human neutrophils shows the predominant presence of PDE IV (cAMP specific) and PDE V (cGMP specific). PDE IV is characterized by (1) cAMP selectivity, (2) a K_M for cAMP of 1.2 M and (3) a typical rank order of IC ₅₀-values for PDE inhibitors: 0.13, 0.17, 47 and 9.5 M for PDE IV selective rolipram, PDE III/IV selective zardaverine, PDE III selective motapizone and unselective 3-isobutyl-l-methylxanthine (IBMX), respectively. Functions of polymorphonuclear leukocytes (PMN) such as N-formylmethionyl-leucyl-phenylalanine (fMLP)-stimulated superoxide release and fMLP/thimerosal elicited leukotriene (LT) biosynthesis are inhibited by these PDE inhibitors with the same rank order and even lower IC₅₀-values. Measurements of changes in cytosolic Ca_i in Fura-2 loaded PMN demonstrate a transient Ca_i increase after stimulation with 0.1 M fMLP and an additional sustained elevation of Ca_i levels in the presence of thimerosal. PDE inhibitors suppress this sustained phase of Ca_i release with the same rank order of IC₅₀-values as LT biosynthesis. The correlation between fMLP/thimerosal-induced LT biosynthesis and Ca_i levels reveal a Ca_i threshold of 150 nM for arachidonic acid metabolism. cAMP levels in PMN were elevated by PDE inhibitors alone by less than 2-fold. In the presence of fMLP however, cAMP was increased up to 10-fold and the efficacy of PDE inhibitors to increase cAMP paralleled their potency to inhibit PDE IV. It is concluded that (1) suppression of PMN functions is achieved by PDE IV inhibition, (2) necessary cAMP elevations are within 50% increase, (3) superoxide release was affected by cAMP/protein kinase A (PKA) directly whereas (4) for inhibition of LT biosynthesis a cAMP related reduction of Ca-influx is involved. Send offprint requests to Ch. Schudt at the above address     

Multiple-dose pharmacokinetics of epirubicin at four different dose levels: studies in patients with metastatic breast cancer

Summary Pharmacokinetic analysis of epirubicin and its metabolites epirubicinol and 7-deoxy-13-dihydro-epirubicinol aglycone during the first and the fourth courses of treatment was performed in 78 patients with metastatic breast cancer. The patients were treated every 3 weeks with epirubicin given as 10-min i.v. infusions at four different dose levels: 40, 60, 90 and 135 mg/m². In most cases (76 of 78 cases), plasma concentration-time curves fitted to a three-compartmental pharmacokinetic model. The terminal half-life of epirubicin was independent of dose and duration of treatment. Large interindividual differences were demonstrated (meant _1/2, 21.6±7.9 h; range, 10.6–69 h;n=110). In two subjects, extremely long half-lives and high serum bilirubin concentrations indicated impaired liver function. No correlation was found between the half-life and levels of liver alanine aminotransferase (ALAT) or serum creatinine. The metabolite epirubicinol appeared quickly after epirubicin administration and its half-lives were shorter than that of the parent compound (meant _1/2, 18.1±4.8 h; range, 8.2–38.4 h;n=105).Formation of the aglycone metabolite was delayed and the half-life of this metabolite was shorter than that of epirubicin (meant _1/2, 13±4.6 h; range, 2.7–29 h;n=104). The AUC of epirubicin and the total AUC (drug and metabolites) were linearly proportional to the dose, with the former value constituting two-thirds of the latter. A correlation was found between AUC and the plasma concentration of epirubicin at two time points (2 and 24 h after administration). The proposed model was AUC=9.44×c ₂+62.5×c ₂₄+157.7 (r=0.953).This work was supported by the Lundbeck Foundation, the Michaelsen Foundation and Farmitalia Carlo Erba Ltd.