Alexza Pharmaceuticals (United States)

The antipsychotic drugs risperidone, paliperidone, olanzapine, quetiapine, aripiprazole, clozapine, haloperidol, and chlorpromazine have been reported to have various degrees of interaction (substrate or inhibitor) with the multidrug resistance transporter, P-glycoprotein (P-gp). An interaction of the antipsychotic drug loxapine with P-gp was recently reported, but an IC50 value was not determined. Loxapine (as the succinate salt) was evaluated as a P-gp substrate, and inhibitor of P-gp mediated transport of digoxin in vitro in Caco-2 cells. Loxapine was not a substrate for P-gp but did exhibit weak-to-moderate inhibition (IC50 = 9.1 μM). Since the typical steady state maximal plasma concentrations of loxapine in clinical use have been reported to be in the nanomolar range, pharmacokinetic interactions due to the inhibition of P-gp activity are not expected.

Prochlorperazine (PCZ) is a phenothiazine drug that was approved by the FDA in 1956. The U.S. Pharmacopeia is currently seeking an improved method for the PCZ drug substance monograph. A new reverse phase HPLC method was developed to separate the impurities and degradants in PCZ and to quantitatively determine the content of PCZ. The developed method applied an Agilent Zorbax SB-C18 column and a gradient mobile phase flow with UV detection for the separation. The method was applied to the separation of U.S. Pharmacopeia (USP) grade PCZ from its impurities. Impurity identification was performed with LC-MS and fraction isolation followed by NMR. Dimeric chlorophenothiazine (CPT) impurities were newly identified in the USP grade PCZ. Forced degradation studies were performed on PCZ under acid, base, oxidation, heat, and photolytic stress conditions. Peak purity assessments using HPLC-photodiode array (PDA) and HPLC-mass spectrometer (MS) detectors demonstrated the developed method to be stability-indicating. The method was validated and was demonstrated to be specific, accurate (recovery > 98.5%), linear (r > 0.999) and sensitive (limit of detection = 0.025 µg/mL or 0.017% of nominal concentration), and suitable for impurity testing and assay of PCZ drug substance.

Listeria monocytogenes is a facultative intracellular pathogen capable of inducing a robust cell-mediated immune response to sub-lethal infection. The capacity of L. monocytogenes to escape from the phagosome and enter the host cell cytosol is paramount for the induction of long-lived CD8 T cell–mediated protective immunity. Here, we show that the impaired T cell response to L. monocytogenes confined within a phagosome is not merely a consequence of inefficient antigen presentation, but is the result of direct suppression of the adaptive response. This suppression limited not only the adaptive response to vacuole-confined L. monocytogenes, but negated the response to bacteria within the cytosol. Co-infection with phagosome-confined and cytosolic L. monocytogenes prevented the generation of acquired immunity and limited expansion of antigen-specific T cells relative to the cytosolic L. monocytogenes strain alone. Bacteria confined to a phagosome suppressed the production of pro-inflammatory cytokines and led to the rapid MyD88-dependent production of IL-10. Blockade of the IL-10 receptor or the absence of MyD88 during primary infection restored protective immunity. Our studies demonstrate that the presence of microbes within a phagosome can directly impact the innate and adaptive immune response by antagonizing the signaling pathways necessary for inflammation and the generation of protective CD8 T cells.

Active immunotherapy has shown great promise in preclinical models for the treatment of infectious and malignant disease. Yet, these promising results have not translated into approved therapies. One of the major deficits of active immunotherapies tested to date in advanced clinical studies has been their inability to stimulate both arms of the immune system appropriately. The interest in using recombinant bacteria as vaccine vectors for active immunotherapy derives in part from their ability to stimulate multiple innate immune pathways and, at the same time, to deliver antigen for presentation to the adaptive immune system. This review will focus on the development of live-attenuated and killed strains of the intracellular bacterium Listeria monocytogenes for treatment of chronic infections and cancer. Early clinical trials intended to demonstrate safety as well as proof of concept have recently been initiated in several indications. Advances in molecular engineering as well as successes and challenges for clinical development of L. monocytogenes-based vaccines will be discussed.