Over the last decade, combination of drugs in all stages of pharmaceutical development has accelerated availability of promising new therapies for difficult to treat diseases. Safety assessment of combined drugs to be tested in humans can occur at a critical path prior to proceeding in clinical testing. A recent survey by The International Consortium for Innovation and Quality in Pharmaceutical Development (IQ DruSafe) summarized member companies’ approaches to combination safety strategies. In addition, feedback from Health Authorities (HAs) support a case-by-case scientific approach in assessing combination products’ safety in accordance with the International Council on Harmonization (ICH) guidelines. Here, we present Pfizer’s drug combination safety approach for various therapeutic areas (TA) including inflammation and immunology, metabolic, and anti-cancer products. There is no one-size-fits-all approach; rather, our main considerations include: strength of the existing clinical safety data for the individual compounds, common target organs, the potential for a synergistic effect, potential drug-drug interaction, routes of administration of each product and disease indications. No formal toxicity studies are considered necessary for anti-cancer drugs, while safety endpoints may be collected in preclinical pharmacology studies especially when the combined drugs present a novel mechanism. Combination safety studies when conducted for non-cancer indications can range from 2 to 13-weeks in duration, conducted usually in rodents, with dosages of individual molecules within clinical pharmacologic ranges. A case-by-case strategy guided by scientific rationale and in close collaboration with HAs remains the best approach to decide on the design and conduct of combination safety studies.
DEHP (di-2-ethylhexyl phthalate), an environmental endocrine disruptor, is widely used in industrial products, particularly as plasticizers and softeners which could disrupt the function of the hypothalamic-pituitary-thyroid (HPT) axis. Rosmarinic acid (RA) possesses potential antioxidant and anti-inflammatory capacities in disease models. Nevertheless, evidence on the association between DEHP-induced thyroid dysfunction and inflammation, as well as the molecular mechanism underlying the protective effects of RA-mitigated DEHP-induced thyroid injury remains inconclusive. Male Sprague Dawley (SD) rats were intragastrically administered DEHP (150 mg/kg, 300 mg/kg, 600 mg/kg) once a day for 90 consecutive days. Also, FRTL-5 cells were treated with a wide range of DEHP concentrations (10-8, 10-7, 10-6, 10-5, 10-4, 10-3, 10-2 M) for 24 hr. Subsequently, RA (50 μM) was administered for 24 hr before 10-4 M DEHP challenge. We found that DEHP induced thyroid damage and inflammatory infiltration in vivo. In addition, we showed that DEHP triggered inflammatory cell death, which is mediated by multiple inflammasomes. Moreover, RA, pyroptosis inhibitor (Ac-YVAD-cmk) and antioxidant inhibitor (NAC) treatment significantly alleviated DEHP-induced thyrocyte death, suppressing pro-inflammatory cytokine production, inhibiting multiple inflammasomes activation and attenuating thyrocyte death, respectively. Collectively, our results reveal that a critical role of inflammasomes activation in DEHP-induced thyroid injury, and suggest that RA confers protection against DEHP-induced thyroid inflammation, and facilitating control of the effects of DEHP after given pyroptosis inhibitor or antioxidant inhibitor. These results indicate that it should be possible to provide novel insights into toxicologically and pharmacologically targeting this molecule to DEHP-induced inflammation.
This study was aimed at examining propofol- (a known anesthetic) induced emotion-related behavioral disorders in mice, and exploring the possible molecular mechanisms. A total of 60 mice were divided into two groups: control and propofol group. Mice were injected with propofol (150 mg/kg, ip) at 8:00 a.m. (once a day, lasting for 30 days). During the 30 days, loss of righting reflex (LORR) and return of righting reflex (RORR) of mice were recorded every day. At the 1st (T1) and 30th (T2) day of drug discontinuance (T2), 15 mice of each group were selected to perform the open field test; then the mice underwent perfusion fixation, and the midbrain and corpus striatum were separated for immunofluorescence assay with anti-tyrosine hydroxylase (Th) and anti- dopamine transporter (DAT) antibodies. Results showed that after propofol injection, LORR and RORR increased and decreased, respectively. Long-term use of propofol resulted in decreased activities of mice (activity trajectory, line crossing, rearing time, scratching times and defecating frequency). Immunofluorescence assay showed long-term use of propofol induced decrease of Th and DAT. Collectively, our present work suggested long-term abuse of propofol induces neuropsychiatric function impairments, and the possible mechanisms are related to dopamine dyssynthesis via down-regulating tyrosine hydroxylase and dopamine transporter.
Dihydropyrazines (DHPs), including 3-hydro-2,2,5,6-tetramethylpyrazine (DHP-3), are glycation products that are spontaneously generated in vivo and ingested via food. DHPs generate various radicals and reactive oxygen species (ROS), which can induce the expression of several antioxidant genes in HepG2 cells. However, detailed information on DHP-response pathways remains elusive. To address this issue, we investigated the effects of DHP-3 on the nuclear factor-κB (NF-κB) pathway, a ROS-sensitive signaling pathway. In lipopolysaccharide-stimulated (LPS-stimulated) HepG2 cells, DHP-3 decreased phosphorylation levels of inhibitor of NF-κB (IκB) and NF-κB p65, and nuclear translocation of NF-κB p65. In addition, DHP-3 reduced the expression of Toll-like receptor 4 (TLR4) and the adaptor protein myeloid differentiation primary response gene 88 (MyD88). Moreover, DHP-3 suppressed the mRNA expression of tumor necrosis factor-alpha (TNFα), and interleukin-1 beta (IL-1β). Taken together, these results suggest that DHP-3 acts as a negative regulator of the TLR4-MyD88-mediated NF-κB signaling pathway.