Multistage pH-responsive codelivery liposomal platform for synergistic cancer therapy.

BACKGROUND: Small interfering RNA (siRNA) is utilized as a potent agent for cancer therapy through regulating the expression of genes associated with tumors. While the widely application of siRNAs in cancer treatment is severely limited by their insufficient biological stability and its poor ability to penetrate cell membranes. Targeted delivery systems hold great promise to selectively deliver loaded drug to tumor site and reduce toxic side effect. However, the elevated tumor interstitial fluid pressure and efficient cytoplasmic release are still two significant obstacles to siRNA delivery. Co-delivery of chemotherapeutic drugs and siRNA represents a potential strategy which may achieve synergistic anticancer effect. Herein, we designed and synthesized a dual pH-responsive peptide (DPRP), which includes three units, a cell-penetrating domain (polyarginine), a polyanionic shielding domain (ehG)n, and an imine linkage between them. Based on the DPRP surface modification, we developed a pH-responsive liposomal system for co-delivering polo-like kinase-1 (PLK-1) specific siRNA and anticancer agent docetaxel (DTX), D-Lsi/DTX, to synergistically exhibit anti-tumor effect. RESULTS: In contrast to the results at the physiological pH (7.4), D-Lsi/DTX lead to the enhanced penetration into tumor spheroid, the facilitated cellular uptake, the promoted escape from endosomes/lysosomes, the improved distribution into cytoplasm, and the increased cellular apoptosis under mildly acidic condition (pH 6.5). Moreover, both in vitro and in vivo study indicated that D-Lsi/DTX had a therapeutic advantage over other control liposomes. We provided clear evidence that liposomal system co-delivering siPLK-1 and DTX could significantly downregulate expression of PLK-1 and inhibit tumor growth without detectable toxic side effect, compared with siPLK-1-loaded liposomes, DTX-loaded liposomes, and the combinatorial administration. CONCLUSION: These results demonstrate great potential of the combined chemo/gene therapy based on the multistage pH-responsive codelivery liposomal platform for synergistic tumor treatment.

Molecular Mechanisms and Structural Basis of Retigabine Analogues in Regulating KCNQ2 Channel.

KCNQ2 channel is one of the important members of potassium voltage-gated channel. KCNQ2 is closely related to neuronal excitatory diseases including epilepsy and neuropathic pain, and also acts as a drug target of the anti-epileptic drug, retigabine (RTG). In the past few decades, RTG has shown strong efficacy in the treatment of refractory epilepsy but has been withdrawn from clinical use due to its multiple adverse effects in clinical phase III trials. To overcome the drawbacks of RTG, several RTG analogues have been developed with different activation potency to KCNQ2. However, the detailed molecular mechanism by which these RTG analogues regulate KCNQ2 channel remains obscure. In this study, we used molecular simulations to analyse the interaction mode between the RTG analogues and KCNQ2, and to determine their molecular mechanism of action. Our data show that the van der Waals interactions, hydrophobic interactions, hydrogen bond, halogen bond, and π-π stacking work together to maintain the binding stability of the drugs in the binding pocket. On an atomic scale, the amide group in the carbamate and the amino group in the 2-aminophenyl moiety of RTG and RL648_81 are identified as key interaction sites. Our finding provides insight into the molecular mechanism by which KCNQ2 channels are regulated by RTG analogues. It also provides direct theoretical support for optimizing design of the KCNQ2 channel openers in the future, which will help treat refractory epilepsy caused by nerve excitability.

Entering the spotlight: Chitosan oligosaccharides as novel activators of CaCCs/TMEM16A.

Calcium-activated chloride channels (CaCCs)/TMEM16A control diverse fundamental physiological functions, and abnormal function of TMEM16A will lead to various diseases including asthma, hypertension, gastrointestinal hypomotility and cancers. Therefore, TMEM16A as drug targets for related diseases has been increasingly concerned by researchers. In this work, COS were reported as novel natural activators of TMEM16A. It was demonstrated that COS can activate TMEM16A in a concentration dependent manner, with an EC50 of 74.5 µg/mL. Then, fluorescence experiments and inside-out patch clamp experiments were combined to confirm that COS can directly activate TMEM16A. Further, we compared the activation effects of COS monomers DP2 to DP6, with DP3 the best activator. Molecular simulation was performed to find that the binding sites between DP3 and TMEM16A are E143 and E146 in TMEM16A, and it was speculated that COS and TMEM16A may be combined by electrostatic interaction. Finally, we verified that guinea pig ileum contraction was promoted by COS and the monomers through activating TMEM16A. Collectively, COS are novel efficient natural activators of TMEM16A, with potential to be developed to treatment diseases caused by down-regulation of TMEM16A including gastrointestinal hypomotility.

Ginsenoside Rb1, a novel activator of the TMEM16A chloride channel, augments the contraction of guinea pig ileum.

Calcium-activated chloride channels (CaCCs) play important roles in many physiological processes, and the molecular basis of CaCCs has been identified as TMEM16A in many cell types. It is well established that TMEM16A is a drug target in many diseases, including cystic fibrosis, hypertension, asthma, and various tumors. Therefore, identifying potent and specific modulators of the TMEM16A channel is crucial. In this study, we identified the first natural activator of TMEM16A from traditional Chinese medicine and explored its mechanism. Our data showed that Ginsenoside Rb1 (GRb1) can activate TMEM16A directly from the intracellular side in a dose-dependent manner at an EC50 of 38.4 ± 2.14 µM. GRb1 specifically activated TMEM16A/B, but not the other previously proposed CaCC mediators such as CFTR and bestrophin. Moreover, GRb1 promoted proliferation of CHO cells stably expressing TMEM16A, in a concentration-dependent manner. Finally, we showed that GRb1 increased the amplitude and frequency of contractions in an isolated guinea pig ileum assay in vivo. In summary, GRb1 can be considered a lead compound for the development of novel drugs for the treatment of diseases caused by TMEM16A dysfunction.

Identification of Resveratrol, an Herbal Compound, as an Activator of the Calcium-Activated Chloride Channel, TMEM16A.

Calcium-activated chloride channels (CaCCs) play vital roles in a variety of physiological processes. Dysfunction of the CaCCs is implicated in many diseases. Drug discovery targeting at CaCCs has recently become possible with the determination that TMEM16A is the molecular identity of CaCCs. In this study, we demonstrated that resveratrol (RES), a Chinese traditional medicine compound, is a novel activator of TMEM16A. The yellow fluorescence protein quenching assay and measurement of intracellular calcium fluorescence intensity show that RES activates TMEM16A channels in an intracellular Ca2+-independent way. The data of inside-out patch clamp revealed that RES dose-dependently activates TMEM16A (EC50 = 47.92 ± 9.35 µM). Furthermore, RES enhanced the contractions of the ileum of guinea pigs by activating the TMEM16A channel, which indicated that RES might be a promising drug for the treatment of gastrointestinal hypomotility. As RES was able to induce TMEM16A channel activation, TMEM16A can be added to the list of RES drug targets.

Activation of neuronal Kv7/KCNQ/M-channels by the opener QO58-lysine and its anti-nociceptive effects on inflammatory pain in rodents.

AIM: The aim of this study was to examine the activation of neuronal Kv7/KCNQ channels by a novel modified Kv7 opener QO58-lysine and to test the anti-nociceptive effects of QO58-lysine on inflammatory pain in rodent models. METHODS: Assays including whole-cell patch clamp recordings, HPLC, and in vivo pain behavioral evaluations were employed. RESULTS: QO58-lysine caused instant activation of Kv7.2/7.3 currents, and increasing the dose of QO58-lysine resulted in a dose-dependent activation of Kv7.2/Kv7.3 currents with an EC50 of 1.2±0.2 µmol/L. QO58-lysine caused a leftward shift of the voltage-dependent activation of Kv7.2/Kv7.3 to a hyperpolarized potential at V1/2=-54.4±2.5 mV from V1/2=-26.0±0.6 mV. The half-life in plasma (t1/2) was derived as 2.9, 2.7, and 3.0 h for doses of 12.5, 25, and 50 mg/kg, respectively. The absolute bioavailabilities for the three doses (12.5, 25, and 50 mg/kg) of QO58-lysine (po) were determined as 13.7%, 24.3%, and 39.3%, respectively. QO58-lysine caused a concentration-dependent reduction in the licking times during phase II pain induced by the injection of formalin into the mouse hindpaw. In the Complete Freund's adjuvant (CFA)-induced inflammatory pain model in rats, oral or intraperitoneal administration of QO58-lysine resulted in a dose-dependent increase in the paw withdrawal threshold, and the anti-nociceptive effect on mechanical allodynia could be reversed by the channel-specific blocker XE991 (3 mg/kg). CONCLUSION: Taken together, our findings show that a modified QO58 compound (QO58-lysine) can specifically activate Kv7.2/7.3/M-channels. Oral or intraperitoneal administration of QO58-lysine, which has improved bioavailability and a half-life of approximately 3 h in plasma, can reverse inflammatory pain in rodent animal models.

Online restricted-access material combined with high-performance liquid chromatography and tandem mass spectrometry for the simultaneous determination of vanillin and its vanillic acid metabolite in human plasma.

An automated online solid-phase extraction with restricted-access material combined with high-performance liquid chromatography and tandem mass spectrometry was developed and validated for the simultaneous quantification of vanillin and its vanillic acid metabolite in human plasma. After protein precipitation by methanol, which contained the internal standards, the supernatant of plasma samples was injected to the system, the endogenous large molecules were flushed out, and target analytes were trapped and enriched on the adsorbent, resulting in a minimization of sample complexity and ion suppression effects. Calibration curves were linear over the concentrations of 5-1000 ng/mL for vanillin and 10-5000 ng/mL for vanillic acid with a coefficient of determination >0.999 for the determined compounds. The lower limits of quantification of vanillin and vanillic acid were 5.0 and 10.0 ng/mL, respectively. The intra- and inter-run precisions expressed as the relative standard deviation were 2.6-8.6 and 3.2-10.2%, respectively, and the accuracies expressed as the relative error were in the range of -6.1 to 7.3%. Extraction recoveries of analytes were between 89.5 and 97.4%. There was no notable matrix effect for any analyte concentration. The developed method was proved to be sensitive, repeatable, and accurate for the quantification of vanillin and its vanillic acid metabolite in human plasma.

Characterization of the effects of Cl⁻ channel modulators on TMEM16A and bestrophin-1 Ca²âº activated Cl⁻ channels.

The Ca(2+) activated Cl(-) channels (CaCCs) play a multitude of important physiological functions. A number of candidate proteins have been proposed to form CaCC, but only two families, the bestrophins and the TMEM16 proteins, recapitulate the properties of native CaCC in expression systems. Studies of endogenous CaCCs are hindered by the lack of specific pharmacology as most Cl(-) channel modulators lack selectivity and a systematic comparison of the effects of these modulators on TMEM16A and bestrophin is missing. In the present study, we studied seven Cl(-) channel inhibitors: niflumic acid (NFA), NPPB, flufenamic acid (FFA), DIDS, tannic acid, CaCCinh-A01 and T16Ainh-A01 for their effects on TMEM16A and bestrophin-1 (Best1) stably expressed in CHO (Chinese hamster ovary) cells using patch clamp technique. Among seven inhibitors studied, NFA showed highest selectivity for TMEM16A (IC50 of 7.40 ± 0.95 µM) over Best1 (IC50 of 102.19 ± 15.05 µM). In contrast, DIDS displayed a reverse selectivity inhibiting Best1 with IC50 of 3.93 ± 0.73 µM and TMEM16A with IC50 of 548.86 ± 25.57 µM. CaCCinh-A01 was the most efficacious blocker for both TMEM16A and Best1 channels. T16Ainh-A01 partially inhibited TMEM16A currents but had no effect on Best1 currents. Tannic acid, NPPB and FFA had variable intermediate effects. Potentiation of channel activity by some of these modulators and the effects on TMEM16A deactivation kinetics were also described. Characterization of Cl(-) channel modulators for their effects on TMEM16A and Best1 will facilitate future studies of native CaCCs.

Development and validation of HTS assay for screening the calcium-activated chloride channel modulators in TMEM16A stably expressed CHO cells.

Calcium-activated chloride channels (CaCCs), for example TMEM16A, are widely expressed in a variety of tissues and are involved in many important physiological functions. We developed and validated an atomic absorption spectroscopy (AAS)-based detection system for high-throughput screening (HTS) of CaCC modulators. With this assay, Cl(-) flux from CHO cells stably transfected with TMEM16A is assayed indirectly, by measuring excess silver ions (Ag(+)) in the supernatant of AgCl precipitates. The screening process involved four steps: (1) TMEM16A CHO cells were incubated in high-K(+) and high-Cl(-) buffer with test compounds, and with ionomycin as Ca(2+) ionophore, for 12 min; (2) cells were washed with a low-K(+), Cl(-)-free and Ca(2+)-free buffer; (3) CaCC/TMEM16A were activated in high-K(+), Cl(-)-free buffer with ionomycin (10 µmol L(-1)) for 12 min; and (4) excess Ag(+) concentration was measured using an ion channel reader (ICR, an AAS system). The assay can be used to screen CaCC activators and inhibitors at the same time. With this assay, positive control drugs, including NPPB, CaCCinh-A01, flufenamic acid (Flu) and Eact, all had good concentration-dependent effects on CaCC/TMEM16A. NPPB and CaCCinh-A01 inhibited the CaCC/TMEM16A currents completely at 300 µmol L(-1), with IC50 values of 39.35 ± 4.72 µmol L(-1) and 6.35 ± 0.27 µmol L(-1), respectively; and Eact, activated CaCC/TMEM16A, with an EC50 value of 3.92 ± 0.87 µmol L(-1).

A Review on the Role of TRP Channels and Their Potential as Drug Targets_An Insight Into the TRP Channel Drug Discovery Methodologies.

Transient receptor potential (TRP) proteins are a large group of ion channels that control many physiological functions in our body. These channels are considered potential therapeutic drug targets for various diseases such as neurological disorders, cancers, cardiovascular disease, and many more. The Nobel Prize in Physiology/Medicine in the year 2021 was awarded to two scientists for the discovery of TRP and PIEZO ion channels. Improving our knowledge of technologies for their study is essential. In the present study, we reviewed the role of TRP channel types in the control of normal physiological functions as well as disease conditions. Also, we discussed the current and novel technologies that can be used to study these channels successfully. As such, Flux assays for detecting ionic flux through ion channels are among the core and widely used tools for screening drug compounds. Technologies based on these assays are available in fully automated high throughput set-ups and help detect changes in radiolabeled or non-radiolabeled ionic flux. Aurora's Ion Channel Reader (ICR), which works based on label-free technology of flux assay, offers sensitive, accurate, and reproducible measurements to perform drug ranking matching with patch-clamp (gold standard) data. The non-radiolabeled trace-based flux assay coupled with the ICR detects changes in various ion types, including potassium, calcium, sodium, and chloride channels, by using appropriate tracer ions. This technology is now considered one of the very successful approaches for analyzing ion channel activity in modern drug discovery. It could be a successful approach for studying various ion channels and transporters, including the different members of the TRP family of ion channels.

Activation of KCNQ2/3 potassium channels by novel pyrazolo[1,5-a]pyrimidin-7(4H)-one derivatives.

The voltage-gated M-type potassium channel, encoded mainly by the KCNQ2/3 genes, plays an important role in the control of neuronal excitability. Mutations in the KCNQ2 gene lead to a form of neonatal epilepsy in humans termed 'benign familial neonatal convulsions', which is characterized by hyperexcitability of neurons. KCNQ openers or activators are expected to decrease the firing of overactive neurons and are thus conducive to the treatment of epilepsy and pain. Here, we report that four novel synthesized derivatives of pyrazolo[1,5-a]pyrimidin-7(4H)-one (PPO) named QO-26, QO-28, QO-40 and QO-41 potently augmented KCNQ2/3 channels expressed in Chinese hamster ovary cells and shifted the half-maximal activation voltage (V(1/2)) in the hyperpolarizing direction. The V(1/2) was negatively shifted in a concentration-dependent manner. The compounds markedly slowed both KCNQ2/3 channel activation and deactivation kinetics. Structure-activity relationship studies suggest that trifluoromethyl at the C-2 position, phenyl or naphthyl at the C-3 position, and trifluoromethyl or chloromethyl at the C-5 position are essential for the activity. These results suggest the four PPO derivatives act as KCNQ2/3 channel openers, providing a new dimension for the design and development of more potent channel openers.

[Simultaneous determination of three sesquiterpene lactones in Inula hupehensis by RP-HPLC].

OBJECTIVE: A RP-HPLC method was developed for simultaneous determination of bigelovin, ergolide and tomentosin in Inula hupehensis. METHOD: An Agilent C18 column (4.6 mm x 250 mm, 5 microm) was used for separation at 40 degrees C. The mobile phase was acetonitrile-water, and the flow rate was 1.2 mL x min(-1). The detection wavelength was set at 210 nm. RESULT: The method has good linearity in the ranges of 0.01792-0.1792 g x L(-1) (r =0.9999) for bigelovin, 0.0424-0.4240 g x L(-1) (r =0.9996) for ergolide, and 0.044 8-0.4480 g x L(-1) (r = 0.9996) for tomentosin. The average recoveries of bigelovin, ergolide, and tomentosin were 98.5%, 98.2%, 98.4%, with the RSD of 1. 3%, 1.3%, 1.7%, respectively. The results demonstrated that there was a significant difference in the contents of three sequterpene lactones among the tested Inulae Flos. CONCLUSION: The results indicated that the present RP-HPLC method is simple, quick and accurate, and can be used for the quality control of I. hupehensis, especially for the authentication of Inulae Flos.

Gender differences in acute toxicity, toxicokinetic and tissue distribution of amphotericin B liposomes in rats.

Amphotericin B (AmB), an effective polyene drug with broad spectrum antifungal activity, is used for serious fungal infections. Liposomal amphotericin B (LAmB) is a lipid dosage form, which has a significantly improved toxicity profile compared with conventional amphotericin B deoxycholate (DAmB). This study focused on verifying the gender differences in the acute toxicity of LAmB and further exploring its causes. Acute toxicity study of LAmB and DAmB were performed in rats, and toxicity responses and mortality of different sexes were observed and recorded. Concentrations of AmB in rat plasma and tissues were determined by a fully validated UPLC-MS/MS assay. The results demonstrated that LAmB showed significant gender differences in acute toxicity, with more severe toxic symptoms and higher mortality for female rats at different doses, but the same differences were not observed for DAmB under the same condition. To explore the cause of differences, toxicokinetic and tissue distribution studies were performed and the results showed that female animals had higher drug exposure, longer half-life and lower plasma clearance compared to male rats, and the drug was mostly distributed in the liver and kidneys, in which female rats displayed a significant higher concentration than that of male rats.

Rab7b Overexpression-Ameliorated Ischemic Brain Damage Following tMCAO Involves Suppression of TLR4 and NF-κB p65.

Cerebral stroke is one of the leading causes of death and permanent disability worldwide. Toll-like receptor 4 (TLR4) and nuclear factor-kappa B (NF-κB) p65 play a critical role in brain damage following ischemia-induced stroke. Rab7b, a lysosome-associated small Rab GTPase, has been implicated in TLR4 regulation; however, its role in cerebral stroke is poorly understood. In this study, by investigating a rat model with cerebral stroke, we found that Rab7b was upregulated in the rat brain following the transient middle cerebral artery occlusion (tMCAO). Functionally, overexpression of Rab7b in the brain by DNA transfection reduced cerebral infarction and improved neurological outcome following tMCAO, suggesting that Rab7b alleviates ischemic brain damage. Mechanistically, Rab7b overexpression suppressed the expression of TLR4 and NF-κB p65 and also inhibited the activation of NF-κB p65. Furthermore, Rab7b overexpression suppressed the production of proinflammatory mediators including TNF-α, IFN-γ, IL-1ß, and IL-6 in the brain following tMCAO. In summary, these results suggest that Rab7b protects against ischemic brain damage following tMCAO and that this protection may relate to the suppressed inflammatory response mediated by TLR4 and NF-κB p65. Our study might offer Rab7b as a novel therapeutic target in the treatment of cerebral stroke.

[Baculovirus expression and establishment of the indirect ELISA for the HA gene of swine influenza virus H1 subtype].

The hemagglutinin (HA) gene fragment of swine influenza virus A/Swine/Guangdong/LM/05(H1N1) was amplified with HA gene specific primers and cloned into baculovirus transfer plasmid pFASTBacGP67B. The recombinant plasmid pFastBacGP67B-H1 was identified by restriction enzyme digestion and gene sequencing. Following the transformation of DH10Bac Escherichia coli component cells by pFastBacGP67B-H1, recombinant bacmids rBacmid-H1 were identified by blue/white selection and PCR analysis. Then recombinant baculovirus rBV-H1 was rescued by lipofectant reagent Cellfectin induced rBacmid-H1 DNA transfection of long-phage sf9 insect cells. The recombinant HA protein was characterized by hemagglutination test, western-blot and immunohistochemistry. An indirect enzyme-linked immunosorbent assay (ELISA) was assessed to detect in pigs IgG against H1 subtype SIV present in Inner Mongolia, Liaoning and Heilongjiang provinces. Positive was found in 31.15% (29 of 93) serum samples tested from swine reared in commercial herds. However, all irrelevant control sera tested were negative. We conclude, therefore, that ELISA performed with recombinant HA as coating antigen was a better tool for swine influenza surveillance in China.

Intractable nausea and vomiting as an uncommon presentation in an anti-aquaporin 4-positive patient.

Autoantibodies targeting aquaporin 4 (AQP4) water channels are a sensitive and specific biomarker for neuromyelitis optica spectrum disorder (NMOSD). Presence of AQP4 antibodies distinguishes NMOSD from multiple sclerosis. We present our experience with an anti-AQP4 antibody-positive patient diagnosed with NMOSD who complained of intractable nausea and vomiting, not restricted to optic neuritis or acute myelitis during the first attack. Her symptoms partially resolved after appropriate therapy with intravenous methylprednisolone and oral prednisolone. Through this case, we hope to draw attention to an unusual neurological presentation of NMOSD which should be included in the differential diagnosis of intractable nausea and vomiting.

Exploring in vivo metabolism and excretion of QO-58L using ultra-high-performance liquid chromatography coupled with tandem mass spectrometry.

QO-58 lysine (QO-58L) as a new potassium channel opener, reported to have a potential activity to cure neuropathic pain. The aim of this research is to develop and validate a high-performance liquid chromatography with tandem spectrometry (LC-MS/MS) method for the quantification of QO-58L in rat urine, feces and bile. In addition, analyze and identify the metabolites in urine and bile. The assay for this compound in samples detected with multiple reaction monitoring mode (MRM), and take nimodipine as internal standards (IS). To better understand the biotransformation of QO-58L, metabolites in urine and bile were identified by using ultra high performance liquid chromatography tandem quadrupole/time of flight mass spectrometry (UHPLC-Q-TOF-MS) in the positive and negative ion mode. Urine, feces and bile were quantified by three new methods. The results showed that: QO-58L was mainly eliminated through fecal route (92.94%), a small amount of it via biliary excretion (2.05%), and rarely through urinary excretion (0.024%). As a result, there are 11 metabolites were identified, including 8 phase I metabolites resulting from elimination, hydroxylation and dihydroxylation, and 3 phase II metabolites originating from sulfation, N-acetylcysteine conjugation and glucuronidation. Furthermore, the newly discoveries of excretion and metabolism significantly expanded our understanding and was going to be greatly helpful for QO-58L's further pharmacokinetic study in vivo.

Efficacy of levetiracetam compared with phenytoin in prevention of seizures in brain injured patients: A meta-analysis.

BACKGROUND: Early and/or late onset in patients with brain injury (BI) is associated with a poorer prognosis, and phenytoin (PHT) is standard of care to prevent seizures. Levetiracetam (LEV), an alternative antiepileptic drug, is associated with less cognitive disruption. The purpose of this study was to evaluate the safety and efficacy of LEV in the prevention of brain traumatic seizures with the standard drug PHT. METHODS: Search the publications on comparison the safety and efficacy of LEV against the standard agent PHT in prevention of traumatic seizures in BI to January 2018. After rigorous reviewing on quality, the data were extracted from eligible trials. All trials analyzed the summary hazard ratios of the endpoints of interest. RESULTS: LEV was found not more effective than PHT in terms of overall seizure (odds ratio [OR] = 0.73; 95% confidence interval [CI] = 0.51-1.05; P = .09), and late seizure (OR = 0.64; 95% CI = 0.34-1.19; P = .16) occurrence. However, there is significant difference in terms of early seizure (OR = 0.63; 95% CI = 0.40-0.99; P = .04). Moreover, there were no significant differences in terms of mortality (OR = 0.67; 95% CI = 0.43-1.05; P = .08), or side effects (OR = 1.31; 95% CI = 0.80-2.15; P = .29) between groups. CONCLUSION: The meta-analysis showed that LEV prevention of seizures was associated with early seizure rates that were lower than the PHT-prolonged course of treatment. There is no statistically significant difference in the efficacy and safety profile of PHT and LEV in cases of traumatic BI.

Enhancing siRNA-based cancer therapy using a new pH-responsive activatable cell-penetrating peptide-modified liposomal system.

As a potent therapeutic agent, small interfering RNA (siRNA) has been exploited to silence critical genes involved in tumor initiation and progression. However, development of a desirable delivery system is required to overcome the unfavorable properties of siRNA such as its high degradability, molecular size, and negative charge to help increase its accumulation in tumor tissues and promote efficient cellular uptake and endosomal/lysosomal escape of the nucleic acids. In this study, we developed a new activatable cell-penetrating peptide (ACPP) that is responsive to an acidic tumor microenvironment, which was then used to modify the surfaces of siRNA-loaded liposomes. The ACPP is composed of a cell-penetrating peptide (CPP), an acid-labile linker (hydrazone), and a polyanionic domain, including glutamic acid and histidine. In the systemic circulation (pH 7.4), the surface polycationic moieties of the CPP (polyarginine) are "shielded" by the intramolecular electrostatic interaction of the inhibitory domain. When exposed to a lower pH, a common property of solid tumors, the ACPP undergoes acid-catalyzed breakage at the hydrazone site, and the consequent protonation of histidine residues promotes detachment of the inhibitory peptide. Subsequently, the unshielded CPP would facilitate the cellular membrane penetration and efficient endosomal/lysosomal evasion of liposomal siRNA. A series of investigations demonstrated that once exposed to an acidic pH, the ACPP-modified liposomes showed elevated cellular uptake, downregulated expression of polo-like kinase 1, and augmented cell apoptosis. In addition, favorable siRNA avoidance of the endosome/lysosome was observed in both MCF-7 and A549 cells, followed by effective cytoplasmic release. In view of its acid sensitivity and therapeutic potency, this newly developed pH-responsive and ACPP-mediated liposome system represents a potential platform for siRNA-based cancer treatment.

Local GABAergic signaling within sensory ganglia controls peripheral nociceptive transmission.

The integration of somatosensory information is generally assumed to be a function of the central nervous system (CNS). Here we describe fully functional GABAergic communication within rodent peripheral sensory ganglia and show that it can modulate transmission of pain-related signals from the peripheral sensory nerves to the CNS. We found that sensory neurons express major proteins necessary for GABA synthesis and release and that sensory neurons released GABA in response to depolarization. In vivo focal infusion of GABA or GABA reuptake inhibitor to sensory ganglia dramatically reduced acute peripherally induced nociception and alleviated neuropathic and inflammatory pain. In addition, focal application of GABA receptor antagonists to sensory ganglia triggered or exacerbated peripherally induced nociception. We also demonstrated that chemogenetic or optogenetic depolarization of GABAergic dorsal root ganglion neurons in vivo reduced acute and chronic peripherally induced nociception. Mechanistically, GABA depolarized the majority of sensory neuron somata, yet produced a net inhibitory effect on the nociceptive transmission due to the filtering effect at nociceptive fiber T-junctions. Our findings indicate that peripheral somatosensory ganglia represent a hitherto underappreciated site of somatosensory signal integration and offer a potential target for therapeutic intervention.