Collagenase in the Walker 256 Carcinoma. A Study of the Latent and Active Enzyme in vivo and in vitro

Chemoproteomic methods can report directly on endogenous, active enzyme populations, which can differ greatly from measures of transcripts or protein abundance alone. Detection and quantification of family-wide probe engagement generally requires LC-MS/MS or gel-based detection methods, which suffer from low resolution, significant input proteome requirements, laborious sample preparation, and expensive equipment. Therefore, methods that can capitalize on the broad target profiling capacity of family-wide chemical probes but that enable specific, rapid, and ultrasensitive quantitation of protein activity in native samples would be useful for basic, translational, and clinical proteomic applications. Here we develop and apply a method that we call soluble activity-dependent proximity ligation (sADPL), which harnesses family-wide chemical probes to convert active enzyme levels into amplifiable barcoded oligonucleotide signals. We demonstrate that sADPL coupled to quantitative PCR signal detection enables multiplexed “writing” and “reading” of active enzyme levels across multiple protein families directly at picogram levels of whole, unfractionated proteome. sADPL profiling in a competitive format allows for highly sensitive detection of drug–protein interaction profiling, which allows for direct quantitative measurements of in vitro and in vivo on- and off-target drug engagement. Finally, we demonstrate that comparative sADPL profiling can be applied for high-throughput molecular phenotyping of primary human tumor samples, leading to the discovery of new connections between metabolic and proteolytic enzyme activity in specific tumor compartments and patient outcomes. We expect that this modular and multiplexed chemoproteomic platform will be a general approach for drug target engagement, as well as comparative enzyme activity profiling for basic and clinical applications.

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Corrigendum

Parasitology ◽

10.1017/s0031182006002228 ◽

2007 ◽

Vol 134 (4) ◽

pp. 607-607

Keyword(s):

Active Site ◽

Active Enzyme ◽

Distilled Water ◽

Cysteine Proteinases ◽

Anthelmintic Efficacy ◽

Order Of Magnitude ◽

Papaya Latex ◽

Published Paper

Stepek, G., Lowe A. E., Buttle D. J., Duce I. R. and Behnke J. M. (2007). Anthelmintic action of plant cysteine proteinases against the rodent stomach nematode, Protospirura muricola, in vitro and in vivo. Parasitology134, 103–112 (Published online 11 October 2006. doi:10.1017/S0031182006001302)The authors of the above article regret that several errors appear in their published paper:On Page 105: In the methods section entitled “In vivo assessment of anthelmintic efficacy of plant cysteine proteinases”, the sentence:‘Five grams of papaya latex were mixed with 8 ml of sterile distilled water (dH2O), filtered, and the amount of active enzyme present was measured, by active-site titration, to be 331 nmol’should read:‘Five grams of papaya latex were mixed with 8 ml of sterile distilled water (dH2O), filtered, and the amount of active enzyme present was measured, by active-site titration, to be 13·24 micromol’And the sentence:‘Each group of mice received a different treatment: 0·2 ml of papaya latex alone (containing 8 nmol active enzyme), …’should read:‘Each group of mice received a different treatment: 0·2 ml of papaya latex alone (containing 331 nmol active enzyme), …’On Page 110: In column 2, line 5, the sentence:‘The amount of active enzyme administered in each dose (8 nmol) is based …’should read:‘The amount of active enzyme administered in each dose (331 nmol) is based …’And on lines 10–15, the sentence:‘Assuming the volume of a mouse stomach to be 1 ml and the enzyme to be present throughout the stomach at equal dilution, the concentration of enzyme would be in the order of 8 microM, which is somewhat lower than the in vitro concentrations.’Should read:Assuming the volume of a mouse stomach to be 1 ml and the enzyme to be present throughout the stomach at equal dilution, the concentration of enzyme would be in the order of 330 microM, which is about an order of magnitude higher than effective in vitro concentrations. A possible reason for the …’

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Synthese, in vivo- und in vitro-Untersuchungen zur tumorhemmenden Wirkung von cis-Dichloro-dipeptidester-platin(II)-Komplexen / Synthesis, in vivo and in vitro Studies on the Antineoplastic Effect of cis-Dichloro-dipeptide Ester Platinum(II) Complexes

Zeitschrift für Naturforschung B ◽

10.1515/znb-1976-0623 ◽

1976 ◽

Vol 31 (6) ◽

pp. 832-845 ◽

Cited By ~ 28

Author(s):

Wolfgang Beck ◽

Bernhard Purucker ◽

Michael Girnth ◽

Helmut Schönenberger ◽

Horst Seidenberger ◽

...

Keyword(s):

Protein Biosynthesis ◽

Sarcoma 180 ◽

Protecting Group ◽

Yoshida Sarcoma ◽

Walker 256 Carcinosarcoma ◽

E Coli ◽

Antineoplastic Effect ◽

Walker 256

cis-Dichlorodipeptide esterplatinum complexesCl2Pt(MetGlyOEt),Cl2Pt(EthionylGlyOEt), Cl2Pt(GlyGlyOEt)2 and Cl2Pt(GlySerOEt)2 are prepared from the α-amino acid complexes by peptide synthesis using platinum as an amino protecting group. cis-Cl2Pt(GlyGlyOEt)2 and cis-Cl2Pt(GlySerOEt)2 have been prepared also directly from K2PtCl4 and the dipeptidesters. cis-Cl2Pt(GlyGlyOEt)2 (2 a) and cis-Cl2Pt(NH3)2 (5) lead to a prefered inhibition of the DNA-synthesis of sarcoma 180, Yoshida-sarcoma and Walker-256-carcinosarcoma in vitro; RNA- and protein biosynthesis are influenced to a much lower degree. 2a and 5 cause filamentous growth in Escherichia coli B. The DNA polymerase deficient strain of E. coli, p 3478 pol A-, is more inhibited by 2 a and 5 than the non deficient strain W 3110 pol A+. Tumor growth of di-2-chloro-ethylmethylamine (HN2) resistant sarcoma 180 and of Yoshida sarcoma is weakly inhibited, whereas Walker-256-carcinosarcoma is markedly inhibited; however 2a and 5 show similar inhibition of the same tumor.

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Allium subhirsutum L. as a Potential Source of Antioxidant and Anticancer Bioactive Molecules: HR-LCMS Phytochemical Profiling, In Vitro and In Vivo Pharmacological Study

Antioxidants ◽

10.3390/antiox9101003 ◽

2020 ◽

Vol 9 (10) ◽

pp. 1003

Author(s):

Riadh Badraoui ◽

Tarek Rebai ◽

Salem Elkahoui ◽

Mousa Alreshidi ◽

Vajid N. Veettil ◽

...

Keyword(s):

Pharmacological Study ◽

Antioxidant Properties ◽

Reducing Power ◽

Bioactive Molecules ◽

Skeletal Metastases ◽

Total Phenolic ◽

Walker 256 ◽

Antioxidant Effects

This study investigated Allium subhirsutum L. (AS) anticancer and antioxidant effects and inhibition of tumor angiogenesis in a murine model of skeletal metastases due to inoculation of Walker 256/B cells. Phytochemical composition of AS extract (ASE) was studied by High Resolution-Liquid Chromatography Mass Spectroscopy (HR-LCMS). Total phenolic and flavonoid contents (TPC and TFC) were determined. In vitro, the antioxidant properties were evaluated by reducing power and antiradical activity against DPPH. Cancer cells’ proliferation, apoptosis, metastatic development and angiogenesis were evaluated using Walker 256/B and MatLyLu cells. The p-coumaric acid was the major phenolic acid (1700 µg/g extract). ASE showed high levels of TPC and TFC and proved potent antioxidant effects. ASE inhibited Walker 256/B and MatLyLu cells’ proliferation (Half-maximal inhibitory concentration: IC50 ≃ 150 µg/mL) and induced apoptosis. In silico and in vivo assays confirmed these findings. ASE effectively acts as a chemo-preventive compound, induces apoptosis and attenuates angiogenesis and osteolytic metastases due to Walker 256/B malignant cells.

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Corydalis Saxicola Bunting Total Alkaloids Attenuate Walker 256-Induced Bone Pain and Osteoclastogenesis by Suppressing RANKL-Induced NF-κB and c-Fos/NFATc1 Pathways in Rats

Frontiers in Pharmacology ◽

10.3389/fphar.2020.609119 ◽

2021 ◽

Vol 11 ◽

Author(s):

Linjie Ju ◽

Peipei Hu ◽

Ping Chen ◽

Jiejie Wu ◽

Zhuoqun Li ◽

...

Keyword(s):

Bone Pain ◽

Bone Structure ◽

Osteolytic Lesion ◽

Bone Cancer ◽

Total Alkaloids ◽

Candidate Drug ◽

Metastatic Bone Pain ◽

Walker 256

Metastatic bone pain is characterized by insufferable bone pain and abnormal bone structure. A major goal of bone cancer treatment is to ameliorate osteolytic lesion induced by tumor cells. Corydalis saxicola Bunting total alkaloids (CSBTA), the alkaloid compounds extracted from the root of C. saxicola Bunting, have been shown to possess anticancer and analgesic properties. In this study, we aimed to verify whether CSBTA could relieve cancer induced bone pain and inhibit osteoclastogenesis. The in vivo results showed that CSBTA ameliorated Walker 256 induced bone pain and osteoporosis in rats. Histopathological changes also supported that CSBTA inhibited Walker 256 cell-mediated osteolysis. Further in vitro analysis confirmed that CSBTA reduced the expression of RANKL and downregulate the level of RANKL/OPG ratio in breast cancer cells. Moreover, CSBTA could inhibit osteoclastogenesis by suppressing RANKL-induced NF-κB and c-Fos/NFATc1 pathways. Collectively, this study demonstrated that CSBTA could attenuate cancer induced bone pain via a novel mechanism. Therefore, CSBTA might be a promising candidate drug for metastatic bone pain patients.

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Effects of 2-Hydroxypropil-Β-Cyclodextrin-Lidocaine on Tumor Growth and Inflammatory Response

Current Drug Delivery ◽

10.2174/1567201817666200512101448 ◽

2020 ◽

Vol 17 (7) ◽

pp. 588-598

Author(s):

Luiz Eduardo Nunes Ferreira ◽

Henrique Ballassini Abdalla ◽

Jéssica Pereira da Costa ◽

Juliana Souza de Freitas Domingues ◽

Jonny Burga-Sánchez ◽

...

Keyword(s):

Inflammatory Response ◽

Positive Control ◽

Control Group ◽

Prostaglandin E ◽

Antitumor Effects ◽

Proliferation And Apoptosis ◽

Plantar Region ◽

Walker 256

Background: Antiproliferative and cytotoxic effects of lidocaine have been reported in tumor cells. However, the use of these drugs is restricted due to their short action with rapid dispersion from the injected site. The complexation of local anesthetics in 2-hydroxypropyl-β-cyclodextrin (HP-β- CD) is able to improve pharmacological features. Objective: This study evaluated the antitumor effects of lidocaine and the complex HP-β-CD-lidocaine (HP-β-CD-lido). Methods: In vitro, human adenocarcinoma (HeLa) and keratinocytes (HaCaT) were exposed to lidocaine formulations and cell viability, proliferation and apoptosis induction were measured. In vivo, Walker 256 carcinoma cells were subcutaneously injected into the plantar region of the rat right hind paw. The animals were treated with a local application of 5% lidocaine or 5% HP-β-CD-lido. Doxorubicin (3 mg/Kg/day, intraperitoneal) was used as a positive control. Edema sizes were measured daily and the release of cytokines (TNF-α, IL-1α and CXCL-1) and prostaglandin E2 was evaluated. Histological analysis was also performed. Results: HaCaT IG50 values were 846 μM and 2253 μM for lido and HP-β-CD-lido, respectively. In HeLa cells, the IG50 was 1765 μM for lido and 2044 μM for HP-β-CD-lido. Lidocaine formulations significantly reduced the paw edema on day 6 after Walker 256 cells inoculation. However, there were no differences in the release of inflammatory mediators in comparison to the control group. Conclusion: Lidocaine formulations were able to reduce the edema in vivo, without affecting the tumor- induced inflammatory response. The antiproliferative effects of lidocaine formulations may have contributed to tumor reduction.

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XTH31, Encoding an in Vitro XEH/XET-Active Enzyme, Regulates Aluminum Sensitivity by Modulating in Vivo XET Action, Cell Wall Xyloglucan Content, and Aluminum Binding Capacity in Arabidopsis

The Plant Cell ◽

10.1105/tpc.112.106039 ◽

2012 ◽

Vol 24 (11) ◽

pp. 4731-4747 ◽

Cited By ~ 112

Author(s):

Xiao Fang Zhu ◽

Yuan Zhi Shi ◽

Gui Jie Lei ◽

Stephen C. Fry ◽

Bao Cai Zhang ◽

...

Keyword(s):

Cell Wall ◽

Binding Capacity ◽

Active Enzyme

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Flow cytometric measurement of cell cycle kinetics in rat Walker-256 carcinoma following in vivo and in vitro pulse labelling with bromodeoxyuridine

Cytometry ◽

10.1002/cyto.990120106 ◽

1991 ◽

Vol 12 (1) ◽

pp. 33-41 ◽

Cited By ~ 17

Author(s):

Franz Fogt ◽

Jennifer Wan ◽

Carl O'Hara ◽

Bruce R. Bistrian ◽

George L. Blackburn ◽

...

Keyword(s):

Cell Cycle ◽

Pulse Labelling ◽

Cell Cycle Kinetics ◽

Flow Cytometric ◽

Flow Cytometric Measurement ◽

Walker 256

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Induction and Differentiation of Dental Epithelium in Vivo and in Vitro

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100053425 ◽

1982 ◽

Vol 40 ◽

pp. 142-145

Author(s):

E. J. Kollar

Keyword(s):

Connective Tissue ◽

Oral Mucosa ◽

Basal Lamina ◽

Functional Derivatives ◽

Specific Source ◽

Dental Epithelium ◽

Connective Tissue Stroma

The differentiation and maintenance of many specialized epithelial structures are dependent on the underlying connective tissue stroma and on an intact basal lamina. These requirements are especially stringent in the development and maintenance of the skin and oral mucosa. The keratinization patterns of thin or thick cornified layers as well as the appearance of specialized functional derivatives such as hair and teeth can be correlated with the specific source of stroma which supports these differentiated expressions.

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Ultrastructural Correlations between Clonal Human Tumor Colonies and in Vivo Neoplasms

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100053711 ◽

1982 ◽

Vol 40 ◽

pp. 210-211

Author(s):

M.J. Murphy ◽

R.R. Price ◽

J.C. Sloman

Keyword(s):

Cultured Cells ◽

Human Tumor ◽

Cell Type ◽

Tumor Mass ◽

Initial Cell ◽

Cloning Assay ◽

Human Tumor Cloning ◽

The Relationship

The in vitro human tumor cloning assay originally described by Salmon and Hamburger has been applied recently to the investigation of differential anti-tumor drug sensitivities over a broad range of human neoplasms. A major problem in the acceptance of this technique has been the question of the relationship between the cultured cells and the original patient tumor, i.e., whether the colonies that develop derive from the neoplasm or from some other cell type within the initial cell population. A study of the ultrastructural morphology of the cultured cells vs. patient tumor has therefore been undertaken to resolve this question. Direct correlation was assured by division of a common tumor mass at surgical resection, one biopsy being fixed for TEM studies, the second being rapidly transported to the laboratory for culture.

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