Discovery of an orally active small-molecule irreversible inhibitor of protein disulfide isomerase for ovarian cancer treatment

Protein disulfide isomerase (PDI), an endoplasmic reticulum chaperone protein, catalyzes disulfide bond breakage, formation, and rearrangement. The effect of PDI inhibition on ovarian cancer progression is not yet clear, and there is a need for potent, selective, and safe small-molecule inhibitors of PDI. Here, we report a class of propynoic acid carbamoyl methyl amides (PACMAs) that are active against a panel of human ovarian cancer cell lines. Using fluorescent derivatives, 2D gel electrophoresis, and MS, we established that PACMA 31, one of the most active analogs, acts as an irreversible small-molecule inhibitor of PDI, forming a covalent bond with the active site cysteines of PDI. We also showed that PDI activity is essential for the survival and proliferation of human ovarian cancer cells. In vivo, PACMA 31 showed tumor targeting ability and significantly suppressed ovarian tumor growth without causing toxicity to normal tissues. These irreversible small-molecule PDI inhibitors represent an important approach for the development of targeted anticancer agents for ovarian cancer therapy, and they can also serve as useful probes for investigating the biology of PDI-implicated pathways.     

Phosphorylation-regulated axonal dependent transport of syntaxin 1 is mediated by a Kinesin-1 adapter

Presynaptic nerve terminals are formed from preassembled vesicles that are delivered to the prospective synapse by kinesin-mediated axonal transport. However, precisely how the various cargoes are linked to the motor proteins remains unclear. Here, we report a transport complex linking syntaxin 1a (Stx) and Munc18, two proteins functioning in synaptic vesicle exocytosis at the presynaptic plasma membrane, to the motor protein Kinesin-1 via the kinesin adaptor FEZ1. Mutation of the FEZ1 ortholog UNC-76 in Caenorhabditis elegans causes defects in the axonal transport of Stx. We also show that binding of FEZ1 to Kinesin-1 and Munc18 is regulated by phosphorylation, with a conserved site (serine 58) being essential for binding. When expressed in C. elegans, wild-type but not phosphorylation-deficient FEZ1 (S58A) restored axonal transport of Stx. We conclude that FEZ1 operates as a kinesin adaptor for the transport of Stx, with cargo loading and unloading being regulated by protein kinases.     

Combination of Stressors in the Critical Periods of Development Increases Resistance to Inflammatory Pain Stress in Adult Rats

Neuroscience and Behavioral Physiology - We report here studies of the long-term effects of combined stress in the prenatal and prepubertal periods of development on measures of tonic inflammatory...     

Maternal stress differently alters nociceptive behaviors in the formalin test in adult female and male rats

The long-term effects of restraint stress in Wistar rats during the last week of gestation were investigated on the acute and tonic phases of the specific biphasic nociceptive behavioral response in the formalin test in offspring, females and males, at 90 days. Prenatal stress produced significant changes in formalin-induced pain, which was more pronounced in females as compared to males. The distorted response in females was more at the supraspinal level with an increased intensity of the licking response in both phases as well as with an increased their duration. Results concerning changes of the interphase length indicate the impairments of inhibitory mechanisms in the central nervous system. Furthermore, profound difference in the effects of prenatal stress on the first phase but similarity in these effects on the second phase in females and males are indirect strong support of the view that the second phase in the formalin test can not be mediated by central sensitization alone but greatly depends on signals ongoing from nociceptive primary afferents. Finally, the results obtained in males are important argument in favor of assumption about different mechanisms of acute and tonic pain. Taken together, these studies show that prenatal stress alters nociceptive behaviors in the formalin test in rats at 90 days in a sex-specific manner.     

Prenatal stress alters time characteristics and intensity of formalin-induced pain responses in juvenile rats

Previous studies found that stressful events during pregnancy can alter the offspring's pain sensitivity to the phasic nociceptive stimuli. The present data constitute the first demonstration of the consequences of prenatal stress to formalin-induced pain in juvenile rats. Injection of formalin into a hind paw of a 25-day-old rat that had not been stressed prenatally produced the typical biphasic specific nociceptive behavioral response consisting of an early short phase lasting 1-4 min followed by a second prolonged phase (12-24 min). Between them there was an interphase that lasted 6-9 min during which the specific behaviors were not shown. This period is generally considered to be a period of inactivity. Prenatally stressed rat pups showed significant increase in flexing+shaking behaviors and in the duration of the second phase of formalin-induced pain in flexing+shaking and licking behaviors and decrease of the duration of the interphase. Disinhibition of the pain behaviors during the interphase was greatly more pronounced in female than in male rats. Sex differences indicate increased vulnerability of inhibitory processes to prenatal stress in females compared with males. These data also underline the importance of understanding the nature of the interphase and provide data on the mechanisms that underlie that component of the formalin test.     

Reduced serotonin synthesis during early embryogeny changes effect of subsequent prenatal stress on persistent pain in the formalin test in adult male and female rats

The considerable evidence supporting a role for serotonin (5-HT) in the embryonic formation of CNS, mediation of prenatal stress, and pain processing is reviewed. Long-term influences of prenatal 5-HT depletion as well as its combination with prenatal stress effects on tonic nociceptive system in 90-day-old Wistar rats were studied in the formalin test. Pregnant dams were injected with para-chlorophenylalanine (pCPA, 400 mg/kg/2 ml, ip), producing 5-HT depletion during the early period of fetal serotonergic system development. The adult offspring from pCPA-treated dams revealed changes in behavioral indices of persistent pain (flexing + shaking and licking) in the formalin test (2.5%, 50 microl) that were accompanied by irreversible morphological alterations in the dorsal raphe nuclei. In the other series of experiments, the role of 5-HT in the mediation of prenatal stress on the behavioral indices of persistent pain was investigated in the adult offspring from dams with 5-HT depletion followed by restraint stress. Stress during the last embryonic week caused much more increase in flexing + shaking and licking in the second tonic phase of the response to formalin in offspring from pCPA- than saline-treated (control) dams. The former was characterized by alterations in the durations of the interphase, the second phase, and the whole behavioral response too. In offspring from pCPA-treated dams, sex dimorphism was revealed in tonic pain evaluated by licking. Together with our previous results in juvenile rats demonstrating the necessity of definite level of prenatal 5-HT for normal development of tonic nociceptive system, the present pioneering findings obtained in adult rats indicate that prenatal 5-HT depletion causes long-term morphological abnormalities in the dorsal raphe nuclei accompanied by alterations in behavioral indices of tonic pain. Early prenatal 5-HT depletion increases vulnerability of tonic nociceptive circuits to the following prenatal stress.     

Effect of prenatal stress on tonic pain in rats

The effect of prenatal stress on specific biphasic behavioral response in 25-day rat pups was studied using the experimental model of formalin-induced tonic pain. Prenatally stressed rats showed hypersensitivity to tonic nociceptive stimulation manifested in increased amplitude and duration of the response. Behavioral responses observed during the interphase period attest to impairment of inhibitory processes (more pronounced in females). These findings suggest that the interphase interval is the period of active inhibitory process rather than rest period.     

Some problems of the current therapy of infective endocarditis

AIM: To analyse clinical characteristics of endocarditis for the last 10 years, treatment difficulties and how to overcome them. MATERIAL AND METHODS: 135 patients with infectious endocarditis (IE) were examined according to the routine scheme using modern methods of diagnosis and therapy control: transthoracic and transesophageal echo-CG, test for antibiotics sensitivity of the microflora, etc. Immediate results were assessed in all the patients, some of them were followed up for maximum 5 years. RESULTS: Last decade was marked for growing difficulties in the treatment of IE related to its polyetiology. It can be caused by such therapy-resistant microbes as Staphylococcus aureus, Pseudomonas aeruginosa, anaerobic infection, nosocomial infection, injections of narcotic drugs, etc. CONCLUSION: Current course of IE dictates the necessity of fighting resistant microflora especially in case of nosocomial disease. Recurrences become more frequent. Indications to surgery did not change for the last decade. The best treatment results are achieved after antibacterial treatment of the valve.     

Persistent pain responses in inflammation and corticosterone levels in juvenile rats born to adrenalectomized dams

Studies in juvenile (Wistar) rats born to adrenalectomized dams (surgery performed 3–4 weeks before mating) addressed the intensity of behavioral pain responses (numbers of flexion + shaking patterns and durations of licking patterns) induced by foci of inflammation in the formalin test and plasma corticosterone levels on the background of the pain response. Maternal adrenalectomy had no effect on basal corticosterone levels or measures of the intensity of the pain response. In conditions of persistent pain (25 min after injection of formalin), corticosterone levels significantly (p < 0.05) increased in the offspring of both intact and operated dams. Females born to adrenalectomized dams, as compared with females from shamoperated dams, showed higher (p = 0.008) hormone levels in response to persistent pain. There were no differences in measures of the pain responses of the female offspring of adrenalectomized and sham-operated dams. There were no gender differences in measures of the pain response and hormone levels. Thus, pain evoked by acute foci of inflammation in the formalin test activated the hypothalamo-hypophysealadrenal system in 25-day-old rats, though the corticosterone released did not decrease pain intensity, which is consistent with data obtained from the offspring of adrenalectomized dams. Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 94, No. 3, pp. 312–317, March, 2008.