A comprehensive platform for quality control of botanical drugs (PhytomicsQC): a case study of Huangqin Tang (HQT) and PHY906

<p>Abstract</p> <p>Background</p> <p>Establishing botanical extracts as globally-accepted polychemical medicines and a new paradigm for disease treatment, requires the development of high-level quality control metrics. Based on comprehensive chemical and biological fingerprints correlated with pharmacology, we propose a general approach called PhytomicsQC to botanical quality control.</p> <p>Methods</p> <p>Incorporating the state-of-the-art analytical methodologies, PhytomicsQC was employed in this study and included the use of liquid chromatography/mass spectrometry (LC/MS) for chemical characterization and chemical fingerprinting, differential cellular gene expression for bioresponse fingerprinting and animal pharmacology for <it>in vivo </it>validation. A statistical pattern comparison method, Phytomics Similarity Index (PSI), based on intensities and intensity ratios, was used to determine the similarity of the chemical and bioresponse fingerprints among different manufactured batches.</p> <p>Results</p> <p>Eighteen batch samples of Huangqin Tang (HQT) and its pharmaceutical grade version (PHY906) were analyzed using the PhytomicsQC platform analysis. Comparative analysis of the batch samples with a clinically tested standardized batch obtained values of PSI similarity between 0.67 and 0.99.</p> <p>Conclusion</p> <p>With rigorous quality control using analytically sensitive and comprehensive chemical and biological fingerprinting, botanical formulations manufactured under standardized manufacturing protocols can produce highly consistent batches of products.</p

Interaction of a traditional Chinese Medicine (PHY906) and CPT-11 on the inflammatory process in the tumor microenvironment

<p>Abstract</p> <p>Background -</p> <p>Traditional Chinese Medicine (TCM) has been used for thousands of years to treat or prevent diseases, including cancer. Good manufacturing practices (GMP) and sophisticated product analysis (PhytomicsQC) to ensure consistency are now available allowing the assessment of its utility. Polychemical Medicines, like TCM, include chemicals with distinct tissue-dependent pharmacodynamic properties that result in tissue-specific bioactivity. Determining the mode of action of these mixtures was previously unsatisfactory; however, information rich RNA microarray technologies now allow for thorough mechanistic studies of the effects complex mixtures. PHY906 is a long used four herb TCM formula employed as an adjuvant to relieve the side effects associated with chemotherapy. Animal studies documented a decrease in global toxicity and an increase in therapeutic effectiveness of chemotherapy when combined with PHY906.</p> <p>Methods -</p> <p>Using a systems biology approach, we studied tumor tissue to identify reasons for the enhancement of the antitumor effect of CPT-11 (CPT-11) by PHY906 in a well-characterized pre-clinical model; the administration of PHY906 and CPT-11 to female BDF-1 mice bearing subcutaneous Colon 38 tumors.</p> <p>Results -</p> <p>We observed that 1) individually PHY906 and CPT-11 induce distinct alterations in tumor, liver and spleen; 2) PHY906 alone predominantly induces repression of transcription and immune-suppression in tumors; 3) these effects are reverted in the presence of CPT-11, with prevalent induction of pro-apoptotic and pro-inflammatory pathways that may favor tumor rejection.</p> <p>Conclusions -</p> <p>PHY906 together with CPT-11 triggers unique changes not activated by each one alone suggesting that the combination creates a unique tissue-specific response.</p

Mechanism Based Quality Control (MBQC) of Herbal Products: A Case Study YIV-906 (PHY906)

YIV-906 (PHY906), a four-herb Chinese medicine formulation, is inspired by an 1800 year-old Chinese formulation called Huang Qin Tang which is traditionally used to treat gastrointestinal (GI) symptoms. In animal studies, it could enhance anti-tumor activity of different classes of anticancer agents and promote faster recovery of the damaged intestines following irinotecan or radiation treatment. Several clinical studies have shown that YIV-906 had the potential to increase the therapeutic index of cancer treatments (chemotherapy, radiation) by prolonging life and improving patient quality of life. Results of animal studies demonstrated five clinical batches of YIV-906 had very similar in vivo activities (protection of body weight loss induced by CPT11 and enhancement of anti-tumor activity of CPT11) while four batches of commercial–made Huang Qin Tang, HQT had no or lower in vivo activities. Two quality control platforms were used to correlate the biological activity between YIV906 and HQT. Chemical profiles (using analysis of 77 peaks intensities) obtained from LC-MS could not be used to differentiate YIV-906 from commercial Huang Qin Tang. A mechanism based quality control (MBQC) platform, comprising 18 luciferase reporter cell lines and two enzymatic assays based on the mechanism action of YIV-906, could be used to differentiate YIV-906 from commercial Huang Qin Tang. Results of MBQC could be matched to their in vivo activities on irinotecan. In conclusion, the quality control of an herbal product should be dependent on its pharmacological usage. For its specific usage appropriate biological assays based on its mechanism action should be developed for QC. Chemical fingerprints comparison approach has limitations unless irrelevant chemicals have been filtered out. Additionally, using a similarity index is only useful when relevant information is used. A MBQC platform should also be applied on other herbal products

Study of Malformin C, a Fungal Source Cyclic Pentapeptide, as an Anti-Cancer Drug.

Malformin C, a fungal cyclic pentapeptide, has been claimed to have anti-cancer potential, but no in vivo study was available to substantiate this property. Therefore, we conducted in vitro and in vivo experiments to investigate its anti-cancer effects and toxicity. Our studies showed Malformin C inhibited Colon 38 and HCT 116 cell growth dose-dependently with an IC50 of 0.27±0.07μM and 0.18±0.023μM respectively. This inhibition was explicated by Malformin C's effect on G2/M arrest. Moreover, we observed up-regulated expression of phospho-histone H2A.X, p53, cleaved CASPASE 3 and LC3 after Malformin C treatment, while the apoptosis assay indicated an increased population of necrotic and late apoptotic cells. In vivo, the pathological study exhibited the acute toxicity of Malformin C at lethal dosage in BDF1 mice might be caused by an acute yet subtle inflammatory response, consistent with elevated IL-6 in the plasma cytokine assay. Further anti-tumor and toxicity experiments proved that 0.3mg/kg injected weekly was the best therapeutic dosage of Malformin C in Colon 38 xenografted BDF1 mice, whereas 0.1mg/kg every other day showed no effect with higher resistance, and 0.9mg/kg per week either led to fatal toxicity in seven-week old mice or displayed no advantage over 0.3mg/kg group in nine-week old mice. Overall, we conclude that Malformin C arrests Colon 38 cells in G2/M phase and induces multiple forms of cell death through necrosis, apoptosis and autophagy. Malformin C has potent cell growth inhibition activity, but the therapeutic index is too low to be an anti-cancer drug

YIV‑906 potentiated anti‑PD1action against hepatocellular carcinoma by enhancing adaptive and innate immunity in the tumor microenvironment

YIV-906 (PHY906) is a standardized botanical cancer drug candidate developed with a systems biology approach—inspired by a traditional Chinese herbal formulation, historically used to treat gastrointestinal symptoms including diarrhea, nausea and vomiting. In combination with chemotherapy and/or radiation therapy, preclinical and clinical results suggest that YIV-906 has the potential to prolong survival and improve quality of life for cancer patients. Here, we demonstrated that YIV-906 plus anti-PD1 could eradicate all Hepa 1–6 tumors in all tumor bearing mice. YIV-906 was found to have multiple mechanisms of action to enhance adaptive and innate immunity. In combination, YIV-906 reduced PD1 or counteracted PD-L1 induction caused by anti-PD1 which led to higher T-cell activation gene expression of the tumor. In addition, YIV-906 could reduce immune tolerance by modulating IDO activity and reducing monocytic MDSC of the tumor. The combination of anti-PD1 and YIV-906 generated acute inflammation in the tumor microenvironment with more M1-like macrophages. YIV-906 could potentiate the action of interferon gamma (IFNg) to increase M1-like macrophage polarization while inhibiting IL4 action to decrease M2 macrophage polarization. Flavonoids from YIV-906 were responsible for modulating IDO activity and potentiating IFNg action in M1-like macrophage polarization. In conclusion, YIV-906 could act as an immunomodulator and enhance the innate and adaptive immune response and potentiate anti-tumor activity for immunotherapies to treat cancer

Multidrug resistance of Malformin C and reversal effects of Verapamil.

<p>Note: Values were means ± SD from more than three independent experiments, with each data point done in duplicate, and all the cells were exposed to different drugs for 72 hours. Verapamil (VRP) was added to the media at a concentration of 5 μM. VRP’s IC₅₀ of KB cells were 34±1 μM, and no toxicity was observed at 5 μM.</p><p>Multidrug resistance of Malformin C and reversal effects of Verapamil.</p

Expression of phosphorylated and total H2A.X in Colon 38 and HCT 116 cells treated with Malformin C.

<p><b>(A, C)</b> The expression of phosphorylated and total H2A.X in Colon 38 cells treated with different concentrations of Malformin C (0μM, 0.14μM, 0.27μM, 0.54μM) and Hydroxyurea (1mM, 2mM) for 2-hour, 4 hours, 8 hours and 24 hours tested by Western blot, with β-Actin expression as an internal control. Hydroxyurea was used as positive control. H2A.X is phosphorylated at Ser139. <b>(B, D)</b> The expression of phosphorylated and total H2A.X in HCT116 cells treated with different concentrations of Malformin C (0μM, 0.14μM, 0.27μM, 0.54μM) and Hydroxyurea (1mM, 2mM) for 2 hours, 4 hours, 8 hours and 24 hours tested by Western blot, with β-Actin expression as an internal control. Hydroxyurea was used as positive control. H2A.X is phosphorylated at Ser139.</p

Cell cycle analysis of Colon 38 cells treated by Malformin C and its combinations.

<p><b>(A)</b> Chemical structure of Malformin C. Malformin C is a member of Malformins, a group of fungal cyclic pentapeptides. Its chemical formula is C₂₃H₃₉O₅N₅S₂ with a molecular weight of 529.7. <b>(B)</b> Cell cycle progression of Colon 38 cells exposed to an increasing concentration of Malformin C for 24 hours. <b>(C)</b> Cell cycle progression of Colon 38 cells exposed to an increasing concentration of Malformin C for 48 hours. <b>(D)</b> The dose-dependent accumulation of G2-M phase Colon 38 cells treated by Malformin C at concentrations of 90 nM, 270 nM and 810 nM. Compared to the control group, the symbol Δ represented <i>P</i><0.05, while * represented <i>P</i><0.01. <b>(E)</b> Cell cycle analysis of Colon 38 cells treated with combinations of Malformin C and SN38. All the cells were exposed to respective compounds indicated above the graph for 24 hours and the analysis was done in duplicate. When treated with SN38, no significant changes of cell cycle progression were observed without or with different dosage of Malformin C.</p