An Integrative Proteome-Based Pharmacologic Characterization and Therapeutic Strategy Exploration of SAHA in Solid Malignancies.

Targeting histone epigenetic modification is an important strategy for anticancer therapy. Histone deacetylase inhibitors (HDACis) have been clinically approved in the treatment of diverse hematological cancers, but mechanisms of drug resistance and poor therapeutic efficacy in solid malignancies remain largely unknown. In this study, we applied a mass spectrometry-based quantitative proteomic strategy to investigate the molecular differences in HDACi vorinostat (SAHA) sensitive and resistant cell lines. The proteomic results revealed that the glycolysis pathway was highly enriched after vorinostat treatment in the resistant cell line, leading to the prediction of a new drug combination, SAHA and hexokinase inhibitor (2-deoxyglucose). The efficacy of this combination was further verified in several solid tumor cell lines. Quantitative proteomics revealed that alterations in the transcription process and protein homeostasis could play roles in the synergetic utilization of these two compounds. Our study showed the application of proteomics in elucidating the drug mechanism and predicting drug combination and the potential of expanding the utilization of HDACi.

Protein Acylation is a General Regulatory Mechanism in Biosynthetic Pathway of Acyl-CoA-Derived Natural Products.

Coenzyme A (CoA) esters of short fatty acids (acyl-CoAs) function as key precursors for the biosynthesis of various natural products and the dominant donors for lysine acylation. Herein, we investigated the functional interplay between beneficial and adverse effects of acyl-CoA supplements on the production of acyl-CoA-derived natural products in microorganisms by using erythromycin-biosynthesized Saccharopolyspora erythraea as a model: accumulation of propionyl-CoA benefited erythromycin biosynthesis, but lysine propionylation inhibited the activities of important enzymes involved in biosynthetic pathways of erythromycin. The results showed that the overexpression of NAD+-dependent deacylase could circumvent the inhibitory effects of high acyl-CoA concentrations. In addition, we demonstrated the similar lysine acylation mechanism in other acyl-CoA-derived natural product biosynthesis, such as malonyl-CoA-derived alkaloid and butyryl-CoA-derived bioalcohol. These observations systematically uncovered the important role of protein acylation on interaction between the accumulation of high concentrations of acyl-CoAs and the efficiency of their use in metabolic pathways.

Effect of Qiangjing Tablets on the MAPK signaling pathway in SD rats with asthenospermia / 中华男科学杂志

<p><b>Objective</b>To investigate the effects of Qiangjing Tablets (QJT) on sperm quality and the MAPK signaling pathway in the SD rat model of asthenospermia (AS).</p><p><b>METHODS</b>A total of 100 male SD rats were randomly divided into five groups of equal number, blank control, AS model control, high-dose QJT, medium-dose QJT, and low-dose QJT. All the rats were intragastrically administered ORN at 200 mg/kg/d for establishment of the AS model except those in the blank control group, which were given 1% CMC sodium solution at 1 ml/100 g by gavage. Meanwhile the animals of the high-, medium-, and low-dose QJT groups were gavaged with QJT at 6700, 3300 and 1700 mg/kg/d, respectively, qd 6 days a week for 20 days. Then the testis issue and the apoptosis of the testicular cells were observed under the electron microscope, the expression of vimentin in the testis was determined with the immunohistochemical SP method, that of ERK1/2 detected by Western blot, and the concentration of TGF-β1 in the semen measured by ELISA.</p><p><b>RESULTS</b>The AS model controls showed round nuclei of spermatocytes, homogeneously distributed chromatins, broken or lost mitochondria, and expanded rough endoplasmic reticulum in the testis tissue. In comparison, the rats of the high-, medium-, and low-dose QJT groups exhibited round nuclei of spermatocytes, homogeneously distributed chromatins, and well-structured mitochondria, rough endoplasmic reticulum and ribosome, which were all similar those of the blank controls. Compared with the blank controls, the AS model rats manifested significantly increased expressions of ERK1/2 (1.00 ± 0.00 vs 1.26 ± 0.10, P<0.01) and vimentin (0.16 ± 0.01 vs 0.17 ± 0.01, P<0.01) and apoptosis rate of cells in the testis tissue (［9.20 ± 3.07］ vs ［42.20 ± 9.17］ %, P<0.01), but decreased level of TGF-β1 in the semen (［627.67 ± 26.07］ vs ［566.73 ± 68.44］ ng/ml, P<0.05). In comparison with the model controls, the rats of the high- and medium- -dose QJT groups presented remarkably down-regulated expressions of ERK1/2 (1.26 ± 0.10 vs 1.14 ± 0.08, P<0.01; 1.26 ± 0.10 vs 1.18 ± 0.05, P<0.05) and vimentin (0.17 ± 0.01 vs 0.16 ± 0.01, P<0.01; 0.17 ± 0.01 vs 0.17 ± 0.09, P<0.05) and decreased rate of cell apoptosis (［42.20 ± 9.17］ vs ［21.60 ± 5.94］ %, P<0.01; ［42.20 ± 9.17］ vs ［33.95 ± 6.39］ %, P<0.05). The concentration of TGF-β1 in the semen was markedly lower in the high-dose QJT than in the AS model control group (［621.78 ± 30.80］ vs ［566.73 ± 68.44］ ng/ml, P < 0.05).</p><p><b>CONCLUSIONS</b>Qiangjing Tablets could improve semen quality in asthenospermia rats by acting against oxidative stress.</p>

Inhibitory effect of Qilan Capsules on the expressions of vasculogenic mimicry-related proteins in prostate cancer / 中华男科学杂志

<p><b>Objective</b>To investigate the effect of Qilan Capsules (QLC) on the expressions of the related proteins HIF-1α, VEGF-α, EphA2 and MMP-1 in the formation of vasculogenic mimicry (VM) in prostate cancer.</p><p><b>METHODS</b>Prostate cancer PC-3 cells were cultured, transfected with siRNA, and divided into eight groups, blank control, HIF-1α siRNA, VEGF-α siRNA, EphA2 siRNA, QLC intervention, QLC + HIF-1α siRNA, QLC + VEGF-α siRNA, and QLC + EphA2 siRNA. The expressions of the HIF-1α, VEGF-α and EphA2 proteins in the pathway of VEGF were determined by Western blot.</p><p><b>RESULTS</b>Compared with the blank control group, the expression of HIF-1α was evidently decreased in the HIF-lα siRNA and QLC + HIF-lα siRNA groups (0.624 7 ± 0.042 8 vs 0.032 8 ± 0.002 5 and 0.036 8 ± 0.018 1, P < 0.05), so were that of VEGF-α in the VEGF-α siRNA and QLC + VEGF-α siRNA groups (0.068 9 ± 0.005 1 vs 0.016 9 ± 0.000 7 and 0.010 9 ± 0.000 8, P < 0.05), that of EphA2 in the EphA2 siRNA and QLC + EphA2 siRNA groups though with no statistically significant difference (0.1684 ± 0.0126 vs 0.134 5 ± 0.028 6 and 0.165 4 ± 0.039 8, P > 0.05), and that of MMP-1 in the HIF-lα siRNA, VEGF-α siRNA and EphA2 siRNA groups (1.696 1 ± 0.152 7 vs 0.435 9 ± 0.036 9, 0.198 7 ± 0.009 0 and 0.0218 ± 0.000 7, P < 0.05).</p><p><b>CONCLUSIONS</b>Qilan Capsules can suppress VM formation in prostate cancer by inhibiting the expressions of HIF-1α, VEGF-α and MMP-1, which plays a role in the clinical treatment of prostate cancer by checking the growth and development of the blood supply system in the tumor tissue.</p>

Qilan Capsules plus androgen-deprivation therapy for Qi-deficiency blood-stasis type of prostate cancer after castration / 中华男科学杂志

Objective@#To observe the synergistic effect of Qilan Capsules in the treatment of the patient with Qi-deficiency blood-stasis type of prostate cancer receiving androgen-deprivation therapy after castration.@*METHODS@#This randomized controlled double-blind study included 246 cases of Qi-deficiency blood-stasis type of prostate cancer after castration, which were randomly divided into an experiment and a control group of equal number to be treated with Qilan Capsules + androgen-deprivation and placebo + androgen-deprivation, respectively. After 6 months of treatment, we compared the International Prostate Symptoms Scores (IPSS), TCM Symptoms Scores (TCMSS), maximal urine flow rate (Qmax), and the level of serum prostate-specific antigen (PSA) between the two groups of patients.@*RESULTS@#Statistically significant differences were observed between the experiment and control groups in the syndrome classification-based efficacy (87.7% vs 67.9%, P 0.05).@*CONCLUSIONS@#Qilan Capsules can significantly enhance the effect of androgen-deprivation therapy in the treatment of Qi-deficiency blood-stasis type of prostate cancer after castration though cannot obviously improve the PSA level.

Lysine Malonylome May Affect the Central Metabolism and Erythromycin Biosynthesis Pathway in Saccharopolyspora erythraea.

Lysine acylation is a dynamic, reversible post-translational modification that can regulate cellular and organismal metabolism in bacteria. Acetylome has been studied well in bacteria. However, to our knowledge, there are no proteomic data on the lysine malonylation in prokaryotes, especially in actinomycetes, which are the major producers of therapeutic antibiotics. In our study, the first malonylome of the erythromycin-producing Saccharopolyspora erythraea was described by using a high-resolution mass spectrometry-based proteomics approach and high-affinity antimalonyllysine antibodies. We identified 192 malonylated sites on 132 substrates. Malonylated proteins are enriched in many biological processes such as protein synthesis, glycolysis and gluconeogenesis, the TCA cycle, and the feeder metabolic pathways of erythromycin synthesis according to GO analysis and KEGG pathway analysis. A total of 238 S/T/Y/H-phosphorylated sites on 158 proteins were also identified in our study, which aimed to explore the potential cross-talk between acylation and phosphorylation. After that, site-specific mutations showed that malonylation is a negative regulatory modification on the enzymatic activity of the acetyl-CoA synthetase (Acs) and glutamine synthetase (Gs). Furthermore, we compared the malonylation levels of the two-growth state to explore the potential effect of malonylation on the erythromycin biosynthesis. These findings expand our current knowledge of the actinomycetes malonylome and supplement the acylproteome databases of the whole bacteria.

Qilin Pills for obstructive azoospermia after ejaculatory duct reconstruction / 中华男科学杂志

<p><b>Objective</b>To observe the clinical effect of Qilin Pills in the treatment of severe oligozoospermia after microsurgical ejaculatory duct reconstruction for obstructive azoospermia.</p><p><b>METHODS</b>We retrospectively analyzed 75 cases of obstructive azoospermia treated by ejaculatory duct reconstruction followed by administration of Qilin Pills. The patients were divided into a Qilin group (n=42) and a control group (n=33) postoperatively, treated with Qilin Pills and placebo, respectively. After 3 months of medication, we compared the sperm quality between the two groups of patients.</p><p><b>RESULTS</b>After 3 months' treatment, all the patients experienced remarkable improvement in sperm quality (P<0.05). Compared with the controls, the patients in the Qilin group showed dramatic increases in sperm concentration, from (0.57±0.25) and (0.60±0.18) ×10⁶/ml before medication to (2.83±0.59) and (1.72 ±0.52) ×10⁶/ml after medication, significantly higher in the Qilin than in the control group (P<0.05). The percentage of grade a sperm was increased from (5.52±5.97) and (5.30±6.26)% to (11.56±9.96) and (10.27±6.52)%, that of grade a+b sperm from (9.68±8.63) and (8.64±10.10)% to (23.42 ±14.10) and (20.81±14.70)%, and that of morphologically normal sperm from (2.00±1.27) and (2.31±0.94)% to (3.54±2.47) and (3.47±1.33)%, respectively. No statistically significant differences were observed in sperm motility and normal sperm morphology between the two groups after treatment (P>0.05). The total effectiveness rate was higher in the Qilin group than in the controls (88.1% vs 72.7%), but with no significant difference between the two groups (P>0.05).</p><p><b>CONCLUSIONS</b>Qilin Pills are fairly effective in improving the quantity of sperm in obstructive azoospermia patients after ejaculatory duct reconstruction.</p>

Longbishu Capsule combined with mesylate doxazosin: an efficacious therapy for benign prostatic hyperplasia / 中华男科学杂志

<p><b>OBJECTIVE</b>To assess the clinical effect and safety of the Chinese medicine Longbishu Capsule combined with mesylate doxazosin in the treatment of benign prostatic hyperplasia (BPH) of the kidney deficiency and blood stagnation type.</p><p><b>METHODS</b>This was a randomized, double-blind, double-simulation control study. We equally assigned 60 men diagnosed with BPH of the kidney deficiency and blood stagnation type to an experimental and a control group, the former treated with mesylate doxazosin plus Longbishu Capsule and the latter with mesylate doxazosin plus placebo. We compared the International Prostate Symptom Score (IPSS), quality of life (QOL), Chinese symptom score (CSS), maximal urinary flow rate (Qmax), and prostate volume between the two groups of patients before and after 6 months of medication.</p><p><b>RESULTS</b>After treatment, there were 5 cured cases, 13 markedly effective cases, 9 effective cases, 1 ineffective case, and 2 eliminated cases in the experimental group, as compared with 2 cured cases, 8 markedly effective cases, 10 effective cases, 7 ineffective cases, and 3 eliminated cases in the control group. The total effectiveness rate was obviously higher in the former (96.4%) than in the latter (74.1%). IPSS, Qmax, and CSS were improved in both of the groups after medication, even more significantly in the experimental than in the control group (IPSS: 15.22 ± 2.98 vs 18.15 ± 5.88, P <0.05; Qmax: [13.56 ± 2.26] ml/s vs [11.78 ± 2.97] ml/s, P <0.05; CSS: 6.18 ± 2.13 vs 9.52 ± 3.15, P <0.05). Because of the difference in the QOL score between the two groups at the baseline (P = 0.038 <0.05), no more comparison was made in this aspect after treatment.</p><p><b>CONCLUSION</b>The combination of Longbishu Capsule with mesylate doxazosin is safe and effective for the treatment of BPH.</p>

[Studies on solid phase extraction method of aristolochic acids and aristololactams in rat urine].

OBJECTIVE: To develop a urine pretreatment method of Solid Phase Extraction (SPE) for the quantitative determination of a number of aristolochic acids (AAs) and aristololactams (ALs) in rat urine. METHOD: The HPLC peak area of AA-I , AA-II, AL-I and AL-II, and other sixteen AAs and ALs was chosen as evaluating index to study the extract results of five Solid Phase Extraction columns (Agilent C18/100 mg, Alltech HG18/100 mg, Alltech C18/100 mg, Alltech C18/300 mg and Agilent Phenyl/200 mg) comparatively. The influences of two washing solvents (water and 1% acetic acid-0.02% triethylamine solution) and seven eluting solvents (ether, acetone, chloroform, ethyl acetate, dichloromethane, methanol and acetonitrile) on extract results of AAs and ALs are comparatively studied with the extracting recoveries of AA-I , AA-II, AL-I and AL-II as indicators. The HPLC peak area of AA-I , AA-II, AL-I and AL-II, and other seven AAs and ALs with good separation being targets, several factors which affect extracting efficiency of analytes, including activating volume, cleansing volume, washing volume and eluting volume, are optimized by orthogonal design experiments with four factors at three levels. RESULT: The established method of SPE is as follows: Agilent Phenyl SPE column of 200 mg, activating with 1.0 mL methanol, cleansing with 1 mL water, adding 1.0 mL rat urine sample, washing with 0.8 mL 1% acetic acid 0.02% triethylamine solution, and eluting with 3.0 mL methanol. CONCLUSION: The established method of SPE is efficient, selective, simple and fast, and can be used as urine pretreatment method to analyze a variety of aristolochic acids and aristololactams in rat urine.

[Affection of different preparations on the content of toxic constituents in traditional Chinese medicines, examplified with Caulis Aristolochiae Manshuriensis].

OBJECTIVE: Taking Caulis Aristolochiae Manshuriensis (Guanmutong in Chinese, derived from the stem of Aristolochia manshuriensis) as an example, to study the affection of different preparations on the content of toxic constituents in traditional Chinese medicines. METHOD: The separation was performed on a zorbax SB-C18 column with mobile phase of acetonitrile-3.7 mmol x L(-1) phosphoric acid buffer, detected at 260 nm. RESULT: The extraction percentage of aristolochic acids I, II and IV a in water extraction (1 h x 2) of Guanmutong were 53.4%, 75.5% and 61.9%, respectively; the remaining quantity of aristolochic acids I, II and IVa in the dregs of the decoction were 22.3%, 15.7% and 30.3%, respectively; Aristolochic acid I was still main substance among these aristolohic acids in the decoction of Guanmutong. CONCLUSION: The content of toxic constituents of the traditional Chinese medicines varies evidently with different preparations of Guanmutong. So the preparation methods of traditional Chinese medicines should be suitably selected according to characteristics of the toxic constituents so as to lessen the body damages of human.