Proteomics analysis of histone deacetylase inhibitor-resistant solid tumors reveals resistant signatures and potential drug combinations.

Histone deacetylase inhibitors (HDACis) are important drugs for cancer therapy, but the indistinct resistant mechanisms of solid tumor therapy greatly limit their clinical application. In this study we conducted HDACi-perturbated proteomics and phosphoproteomics analyses in HDACi-sensitive and -resistant cell lines using a tandem mass tag (TMT)-based quantitative proteomic strategy. We found that the ribosome biogenesis proteins MRTO4, PES1, WDR74 and NOP16 vital to tumorigenesis might regulate the tumor sensitivity to HDACi. By integrating HDACi-perturbated protein signature with previously reported proteomics and drug sensitivity data, we predicted and validated a series of drug combination pairs potentially to enhance the sensitivity of HDACi in diverse solid tumor. Functional phosphoproteomic analysis further identified the kinase PDK1 and ROCK as potential HDACi-resistant signatures. Overall, this study reveals the potential HDACi-resistant signatures and may provide promising drug combination strategies to attenuate the resistance of solid tumor to HDACi.

Investigation of targets and anticancer mechanisms of covalently acting natural products by functional proteomics.

Eriocalyxin B (EB), 17-hydroxy-jolkinolide B (HJB), parthenolide (PN), xanthatin (XT) and andrographolide (AG) are terpenoid natural products with a variety of promising antitumor activities, which commonly bear electrophilic groups (α,ß-unsaturated carbonyl groups and/or epoxides) capable of covalently modifying protein cysteine residues. However, their direct targets and underlying molecular mechanisms are still largely unclear, which limits the development of these compounds. In this study, we integrated activity-based protein profiling (ABPP) and quantitative proteomics approach to systematically characterize the covalent targets of these natural products and their involved cellular pathways. We first demonstrated the anti-proliferation activities of these five compounds in triple-negative breast cancer cell MDA-MB-231. Tandem mass tag (TMT)-based quantitative proteomics showed all five compounds commonly affected the ubiquitin mediated proteolysis pathways. ABPP platform identified the preferentially modified targets of EB and PN, two natural products with high anti-proliferation activity. Biochemical experiments showed that PN inhibited the cell proliferation through targeting ubiquitin carboxyl-terminal hydrolase 10 (USP10). Together, this study uncovered the covalently modified targets of these natural products and potential molecular mechanisms of their antitumor activities.

Proteomic characterization of post-translational modifications in drug discovery.

Protein post-translational modifications (PTMs), which are usually enzymatically catalyzed, are major regulators of protein activity and involved in almost all celluar processes. Dysregulation of PTMs is associated with various types of diseases. Therefore, PTM regulatory enzymes represent as an attractive and important class of targets in drug research and development. Inhibitors against kinases, methyltransferases, deacetyltransferases, ubiquitin ligases have achieved remarkable success in clinical application. Mass spectrometry-based proteomics technologies serve as a powerful approach for system-wide characterization of PTMs, which facilitates the identification of drug targets, elucidation of the mechanisms of action of drugs, and discovery of biomakers in personalized therapy. In this review, we summarize recent advances of proteomics-based studies on PTM targeting drugs and discuss how proteomics strategies facilicate drug target identification, mechanism elucidation, and new therapy development in precision medicine.

The novel cereblon modulator CC-885 inhibits mitophagy via selective degradation of BNIP3L.

Mitophagy is a degradative pathway that mediates the degradation of the entire mitochondria, and defects in this process are implicated in many diseases including cancer. In mammals, mitophagy is mediated by BNIP3L (also known as NIX) that is a dual regulator of mitochondrial turnover and programmed cell death pathways. Acute myeloid leukemia (AML) cells with deficiency of BNIP3L are more sensitive to mitochondria-targeting drugs. But small molecular inhibitors for BNIP3L are currently not available. Some immunomodulatory drugs (IMiDs) have been proved by FDA for hematologic malignancies, however, the underlining molecular mechanisms are still elusive, which hindered the applications of BNIP3L inhibition for AML treatment. In this study we carried out MS-based quantitative proteomics analysis to identify the potential neosubstrates of a novel thalidomide derivative CC-885 in A549 cells. In total, we quantified 5029 proteins with 36 downregulated in CRBN+/+ cell after CC-885 administration. Bioinformatic analysis showed that macromitophagy pathway was enriched in the negative pathway after CC-885 treatment. We further found that CC-885 caused both dose- and time-dependent degradation of BNIP3L in CRBN+/+, but not CRBN-/- cell. Thus, our data uncover a novel role of CC-885 in the regulation of mitophagy by targeting BNIP3L for CRL4CRBN E3 ligase-dependent ubiquitination and degradation, suggesting that CC-885 could be used as a selective BNIP3L degradator for the further investigation. Furthermore, we demonstrated that CC-885 could enhance AML cell sensitivity to the mitochondria-targeting drug rotenone, suggesting that combining CC-885 and mitochondria-targeting drugs may be a therapeutic strategy for AML patients.

[Flame atomic absorption spectrometric determination of copper and lead in beer after preconcentration using a rapid coprecipitation technique with 8-oxyquinoline].

A method was proposed for the determination of trace copper and lead in beer with flame atomic absorption spectrometry after preconcentration of copper and lead by rapid coprecipitation technique with 8-oxyquinoline-Mg(II) using manganese as an internal standard at pH 9. The standard addition recovery of lead is between 97.6%-103.0%. The detection limit is 6.28 x 10(-3) microg x mL(-1) for copper and 2.26 x 10(-2) microg x mL(-1) for lead when the sample volume is 100 mL. The effect of matrix can be overcome by the method and the results are satisfying. The method proposed here is rapid and has good reproducibility.