Treatment with tumour infiltrating lymphocytes and interleukin-2 in patients with metastatic melanoma: A pilot study

Tumour infiltrating lymphocytes (TIL) were isolated and expanded from biopsy samples of 4 patients with metastatic melanoma. The patients were treated with autologous expanded TIL and continuous or bolus infusion of Interleukin 2 (IL-2) at a dose of 18 × 10⁶ International Units/m²/day for 5 days starting 36–48 hours after administration of cyclophosphamide at a dose of 1 g/m². The number of TIL infused ranged from 10¹⁰ to 5,56 × 10¹⁰ cells. Two patients had stable disease (SD) lasting for 2 1/2 and 4 months respectively and they died 24 and 13 months after therapy. One patient died during therapy due to a pseudomonas septicaemia and another patient developed progressive disease (PD). He died 3 months after the start of therapy. The side effects were substantial but most of them were reversible upon cessation of the treatment.The majority of the expanded TIL of all patients were of the CD8+ phenotype. Cutaneous metastases from two patients, removed after treatment with IL-2 and TIL, showed moderate lymphocytic infiltration also mainly of CD8+ T cells.The treatment with IL-2 and TIL is feasible, but further investigations should continue in an attempt to improve the efficacy of the therapy, to reduce toxicity and to diminish the costs and labour of the culture methods.     

Progesterone induction of the uterine milk proteins: major secretory proteins of sheep endometrium

Progesterone induction of the uterine milk proteins (UTMP), the major secretory products of the ovine uterus during pregnancy, was studied in ovariectomized ewes given physiological levels of progesterone for 0, 2, 6, 14, or 30 days. Western blotting of uterine flushes and of endometrial explant culture medium, endometrial RNA analyses on dot and Northern blots, and immunocytochemistry performed on uterine tissue sections demonstrated the presence of low levels of UTMP mRNA and UTMP protein after 6 days of progesterone therapy, and increasing levels of UTMP production and secretion after 14 days. Highest activity was observed at Day 30. The induction of the UTMP progressed from small amounts of antigen present in the supranuclear region of a few epithelial cells in deep and middle-depth regions of uterine glands in the Day 6 progesterone-treatment group to large amounts detected in epithelial cells spread throughout the length of the glands in later groups. UTMP production was also identified in the uteri of intact ewes at Day 16 (but not earlier) of the estrous cycle and during early pregnancy (Days 14 to 22). Production of a protein similar to the UTMP was also noted in the uterus of a pregnant cow. The UTMP provide a good model of a progesterone-responsive secretory protein in a mammal whose synthesis is increased gradually over a period of weeks.     

In vivo and in vitro characterization of novel microparticulates based on hyaluronan and chitosan hydroglutamate

This study examined the application of previously characterized microparticles composed of hyaluronan (HA) and chitosan hydroglutamate (CH) as well as novel microparticles consisting of both polymers (HA/CH) to improve the nasal delivery of a model drug. The rabbit bioavailabilities of gentamicin incorporated in HA, CH, and HA/CH microparticles were increased 23-, 31-, and 42-fold, respectively, compared with the control intranasal solution of gentamicin, indicating that all test microparticles were retained for longer periods on the nasal mucosa of the rabbits as supported by previous in vitro dissolution as well as frog palate mucoadhesion studies, thereby improving drug absorption. The higher bioavailabilities of CH-based formulations (CH and HA/CH) suggest the penetration-enhancing effects of CH may also be partially responsible for the improvement. A model was developed, based on a glass impinger device, to deliver dry powder formulations reproducibly onto the surface of cultured cell monolayers. In vitro permeability and fluorescence microscopy studies on the tight junctions of the 16HBE14o- cell lines further confirmed the ability of CH-based formulations to enhance penetration. Furthermore, the in vitro absorption profile from cell culture studies was consistent with those determined from in vivo studies. The complementary effect from the mucoadhesive nature of HA coupled with the penetration-enhancing effects of CH makes the novel HA/CH formulation a promising nasal delivery system.     

Regression models for linkage heterogeneity applied to familial prostate cancer

Linkage heterogeneity frequently occurs for complex genetic diseases, and statistical methods must account for it to avoid severe loss in power to discover susceptibility genes. A common method to allow for only a fraction of linked pedigrees is to fit a mixture likelihood and then to test for linkage homogeneity, given linkage (admixture test), or to test for linkage while allowing for heterogeneity, using the heterogeneity LOD (HLOD) score. Furthermore, features of the families, such as mean age at diagnosis, may help to discriminate families that demonstrate linkage from those that do not. Pedigree features are often used to create homogeneous subsets, and LOD or HLOD scores are then computed within the subsets. However, this practice introduces several problems, including reduced power (which results from multiple testing and small sample sizes within subsets) and difficulty in interpretation of results. To address some of these limitations, we present a regression-based extension of the mixture likelihood for which pedigree features are used as covariates that determine the probability that a family is the linked type. Some advantages of this approach are that multiple covariates can be used (including quantitative covariates), covariates can be adjusted for each other, and interactions among covariates can be assessed. This new regression method is applied to linkage data for familial prostate cancer and provides new insights into the understanding of prostate cancer linkage heterogeneity.     

Direct involvement of the CDR3-like domain of CD4 in T helper cell activation.

The CD4 glycoprotein, a member of the Ig super-family, has long been known to play an important role in the immunologic activation of Th cells. The precise manner in which CD4 participates in this activation process is not yet understood. In an attempt to further define its role in Th cell activation, we modeled the D1 domain of the murine CD4 protein (L3T4) based on the experimentally determined high resolution structure of the human CD4 protein. Because the D1 domain of CD4 strongly resembles the V kappa chain of an antibody, we addressed the question of whether the CDR-like regions of CD4 are also involved in mediating protein-protein interactions. Consequently, we used the modeled L3T4 structure as a template in the design of conformational mimics of the CDR3-like region (residues 86-94). Only the analog designed to mimic both the sequence and conformation of this region exhibited highly specific inhibition of CD4-dependent responses. Because the inhibitory activity could be localized to the Th cell itself, it appears that this analog acts by uncoupling a CD4 association (independent of an APC) critical to generating a proliferative response.     

Effect of an intervention to improve the cardiovascular health of family members of patients with coronary artery disease: a randomized trial

Background:

Family members of patients with coronary artery disease (CAD) have higher risk of vascular events. We conducted a trial to determine if a family heart-health intervention could reduce their risk of CAD.

Methods:

We assessed coronary risk factors and randomized 426 family members of patients with CAD to a family heart-health intervention (n = 211) or control (n = 215). The intervention included feedback about risk factors, assistance with goal setting and counselling from health educators for 12 months. Reports were sent to the primary care physicians of patients whose lipid levels and blood pressure exceeded threshold values. All participants received printed materials about smoking cessation, healthy eating, weight management and physical activity; the control group received only these materials. The main outcomes (ratio of total cholesterol to high-density lipoprotein [HDL] cholesterol; physical activity; fruit and vegetable consumption) were assessed at 3 and 12 months. We examined group and time effects using mixed models analyses with the baseline values as covariates. The secondary outcomes were plasma lipid levels (total cholesterol, low-density lipoprotein cholesterol, HDL cholesterol and triglycerides); glucose level; blood pressure; smoking status; waist circumference; body mass index; and the use of blood pressure, lipid-lowering and smoking cessation medications.

Results:

We found no effect of the intervention on the ratio of total cholesterol to HDL cholesterol. However, participants in the intervention group reported consuming more fruit and vegetables (1.2 servings per day more after 3 mo and 0.8 servings at 12 mo; p < 0.001). There was a significant group by time interaction for physical activity (p = 0.03). At 3 months, those in the intervention group reported 65.8 more minutes of physical activity per week (95% confidence interval [CI] 47.0–84.7 min). At 12 months, participants in the intervention group reported 23.9 more minutes each week (95% CI 3.9–44.0 min).

Interpretation:

A health educator–led heart-health intervention did not improve the ratio of total cholesterol to HDL cholesterol but did increase reported physical activity and fruit and vegetable consumption among family members of patients with CAD. Hospitalization of a spouse, sibling or parent is an opportunity to improve cardiovascular health among other family members. Trial registration: clinicaltrials.gov, no {"type":"clinical-trial","attrs":{"text":"NCT00552591","term_id":"NCT00552591"}}NCT00552591.People with a family history of coronary artery disease (CAD) among their first-degree relatives have an increased risk of vascular events.^1–5 This risk is greater if more than 1 relative has heart disease, or if the onset of disease in relatives occurred before age 55.^1–3,5 A concordance of coronary risk factors between patients with heart disease and their spouses has also been reported.^6–10 Although genetic factors are important, familial aggregation of coronary risk factors also plays a role.^9,11,12 Guidelines recommend screening individuals with a positive family history,^13–15 but screening rates are low (< 15%), and interventions are rarely initiated.^16,17 Family members of patients with heart disease may benefit from focused interventions to improve cardiovascular health. Counselling that address physical inactivity and dietary behaviours is central to these interventions; clinical management of risk factors such as tobacco addiction, dyslipidemia, hypertension and dysglycemia are also important. We developed and evaluated a year-long family-based heart-health intervention for the siblings, offspring and spouses of patients admitted to hospital with an acute coronary event.