Monitoring biological action of rapamycin in renal transplantation.

BACKGROUND: Inhibition of P70S6 kinase (P70(S6K)) phosphorylation in activated T cells is 1 of the major mechanisms by which rapamycin exerts its immunosuppressive action. STUDY DESIGN: Observational cohort study. SETTINGS & PARTICIPANTS: 2 different groups of kidney transplant recipients at a single center: 30 transplant recipients converted from mycophenolic acid and low-dose prednisone plus cyclosporine A to mycophenolic acid and low-dose prednisone plus rapamycin therapy for chronic allograft nephropathy (group 1) and 16 recipients of suboptimal organs converted from tacrolimus plus rapamycin to rapamycin therapy alone after 3 months (group 2). PREDICTOR: Exposure to rapamycin therapy and rapamycin trough levels. OUTCOMES & MEASUREMENTS: Basal and stimulated phosphorylation of P70(S6K) was measured by using Western blotting in patients' peripheral-blood mononuclear cells before and 6 to 11 months after conversion to rapamycin-based therapy. Kinase activation was attained in vivo by means of intravenous insulin injection. RESULTS: The potency of rapamycin inhibition of P70(S6K) phosphorylation varied among patients (RAPA blood concentration required to achieve 50% inhibition of P70(S6K) activation for mitogen-activated kinase, 3.14 to 12.14 ng/mL) and failed to correlate with drug trough levels. The combination of tacrolimus and rapamycin limited the inhibitory effect of the latter drug on P70(S6K) activation. LIMITATIONS: Need for additional studies exploring the relationship between P70(S6K) activity and kidney graft outcome. Exclusion of patients with diabetes. CONCLUSIONS: Long-term rapamycin treatment inhibits P70(S6K) phosphorylation in peripheral-blood mononuclear cells without significant correlation with rapamycin trough levels. By measuring in vivo the biological action of rapamycin, the assay may provide potentially relevant information for the clinical management of rapamycin-treated patients.

Chronic inhibition of mammalian target of rapamycin signaling downregulates insulin receptor substrates 1 and 2 and AKT activation: A crossroad between cancer and diabetes?

Overactivation of the mammalian target of rapamycin (mTOR) branch downstream of the phosphatidylinositol 3-kinase-AKT pathway critically modulates insulin and growth factor signaling by insulin receptor substrates (IRS). On the basis of in vitro studies, the mTOR inhibitor rapamycin has been reported to lead to enhanced activation of AKT by relieving this feedback inhibition on IRS function. In view of the critical role of AKT in insulin signaling and tumorigenesis, the in vivo expression and activation of this kinase and of IRS-1 and IRS-2 were explored in PBMC of 30 patients who were treated long term with rapamycin. A marked decrease of basal and insulin-stimulated AKT phosphorylation, which correlated with the increase of patients' insulin resistance, and a significant increase of IRS total protein expression, together with a lower (IRS-2) or absent (IRS-1) increase of insulin-induced tyrosine phosphorylation, were found. Therefore, contrary to the expectations, long-term exposure to rapamycin caused the impairment of IRS signaling and AKT activation, and this would help to explain the antiproliferative effect and the possible deterioration of glucose metabolism that are observed in rapamycin-treated patients. These findings may form a novel basis for improved understanding of the role of mTOR inhibition in human diseases, such as diabetes and cancer.

Drug targets in stress-related disorders.

Nervous and immune systems mutually cooperate via release of mediators of both neurological and immunological derivation. Adrenocorticotropin hormone (ACTH) is a product of the hypothalamus-pituitary adrenal axis (HPAA) which stimulates secretion of corticosteroids from adrenals. In turn, corticosteroids modulate the immune response in virtue of their anti-inflammatory activity. On the other hand, catecholamines, products of the sympathetic nervous system (SNS), regulate immune function by acting on specific beta-adrenergic receptors. Conversely, cytokines released by monocytes/macrophages and lymphocytes, upon antigenic stimulation, are able to cross the blood-brain-barrier, thus modulating nervous functions (e.g., thermoregulation, sleep, and appetite). However, cytokines are locally produced in the brain, especially in the hypothalamus, thus contributing to the development of anorexic, pyrogenic, somnogenic and behavioural effects. Besides pathogens and/or their products, the so-called stressors are able to activate both HPAA and SNS, thus influencing immune responses. In this respect, many studies conducted in medical students taking exams have evidenced an array of stress-induced immune alterations. Phobic disorders and migraine without aura (MWA) represent examples of stress-related disorders in which phagocytic immune deficits, endotoxemia and exaggerated levels of proinflammatory cytokines [Tumor Necrosis Factor-alpha (TNF- alpha), and interleukin- 1 beta] have been detected. Quite interestingly, administration of a thymic hormone could ameliorate clinical symptoms in phobic patients. In MWA patients, a beta-blocker, propranolol, could mitigate migraine, whose cessation coincided with a drop of TNF-alpha serum concentration. In phobic disorders and in MWA, benzodiazepines are very often administered and, in this respect, some of them, such as diazepam, inhibit immune functions, while others, e.g., alprazolam, enhance immune responses. Alprazolam could improve clinical symptoms in MWA patients. Chronic Fatigue Syndrome (CFS) is a disorder whose etiology and pathogenesis are still unknown. In this syndrome both abnormalities of nervous and immune systems have been reported. Despite many immune parameters evaluated in CFS no specific biomarkers of disease have been found. Our own data are in agreement with current literature in that we found decreased levels of serum (IFN)-gamma in these patients, thus indicating a predominance of T helper (h)1 response in CFS. Also leptin, a hormone which regulates food intake, fluctuates within normal ranges in CFS individuals. Quite interestingly, in depressed patients, used as controls, leptinaemia was more elevated than in CFS. Finally, in a series of recent therapeutic trials several immunomodulating agents have been used, such as staphypan Berna, lactic acid bacteria, kuibitang and intravenous immunoglobulin. In conclusion, it seems that major drug targets in stress-related disorders are immune cells in terms of inhibition of proinflammatory cytokines and modulation of Th responses. In particular, according to recent evidences, antidepressants seem to exert beneficial effects in experimental autoimmune neuritis in rats by decreasing IFN- beta release or augmenting NK activity in depressed patients.