There and Back Again: A Perspective on 20 Years of CYP4Z1.

CYP4Z1, a highly expressed CYP gene in breast cancer, was one of the last CYPs to be identified in the human genome, some twenty years ago. CYP4 enzymes typically catalyze w-hydroxylation and metabolize w3 and w6 polyunsaturated fatty acids (PUFAs) to bioactive lipid metabolites that can influence tumor growth and metastasis. These attributes of CYP4Z1 make it an attractive target for new chemotherapeutic drug design, as a potential biomarker for selection of patients that might respond favorably to drugs and for developing enzyme inhibitors as potential therapeutic agents. This review summarizes the current state of knowledge regarding the advancing biochemistry of CYP4Z1, its role in breast cancer and the recent synthesis of selective chemical inhibitors of the enzyme. We identify gaps that need to be filled to further advance this field and present new experimental data on recombinant CYP4Z1 expression and purification of the active catalytic form. Significance Statement In breast cancer, an unmet need is the availability of highly effective therapeutic agents, especially for triple negative breast cancer. The relevance of the work summarized in this mini-review is that it identifies a new potential drug target, CYP4Z1, and discusses ways in which the gene product's catalytic activity might be modulated in order to combat this malignancy and limit its spread.

Cytochrome P450 Family 4F2 and 4F11 Haplotype Mapping and Association with Hepatic Gene Expression and Vitamin K Hydroxylation Activity.

This study evaluated the underlying mechanistic links between genetic variability in vitamin K metabolic pathway genes (CYP4F2 and CYP4F11) and phylloquinone hydroxylation activity using genotype- and haplotype-based approaches. Specifically, we characterized genetic variability in the CYP4F2/CYP4F11 locus and compared common single allele genotypes and common haplotypes as predictors of hepatic gene expression, enzyme abundance, and phylloquinone (VK1) ω-hydroxylation kinetics. We measured CYP4F2 and CYP4F11 mRNA levels, CYP4F2 and CYP4F11 protein abundances, and the VK1 concentration-dependent ω-hydroxylation rate in matched human liver nucleic acid and microsome samples, utilizing a novel in vitro population modeling approach. Results indicate that accounting for the CYP4F2*3 allele alone is sufficient to capture most of the genetic-derived variability in the observed phenotypes. Additionally, our findings highlight the important contribution that CYP4F11 makes toward vitamin K metabolism in the human liver.

Improved methods for the detection of heme and protoporphyrin IX adducts and quantification of heme B from cytochrome P450 containing systems.

Heme B is a critical prosthetic group for the function of numerous proteins including the cytochrome P450 (CYP) family of enzymes. CYP enzymes are involved in the metabolism of endogenous and xenobiotic molecules that are of central interest in drug development. Formation of reactive metabolites by CYPs can lead to heme modification and destruction of the enzyme. The structure of the adducted heme can provide key information on the mechanism of inactivation, which is of great interest during preclinical drug discovery. Historically, techniques to extract the modified heme or protoporphyrin IX species involved harsh extraction conditions and esterification of propionate groups to aid chromatography. We have developed a simplified extraction method and LC/MS chromatography system that does not require derivatization to quantify heme B and identify modified heme B species from multiple CYP-containing matrices. The method uses mass defect filter triggered data dependent MS2 scans to rapidly identify heme and protoporphyrin IX adducts. These methods may also be useful for the analysis of other heme variants and hemoproteins.

Characterization of Gla proteoforms and non-Gla peptides of gamma carboxylated proteins: Application to quantification of prothrombin proteoforms in human plasma.

Gamma (γ) carboxylation is an essential post-translational modification in vitamin K-dependent proteins (VKDPs), involved in maintaining critical biological homeostasis. Alterations in the abundance or activity of these proteins have pharmacological and pathological consequences. Importantly, low levels of fully γ-carboxylated clotting factors increase plasma des-γ-carboxy precursors resulting in little or no biological activity. Therefore, it is important to characterize the levels of γ-carboxylation that reflect the active state of these proteins. The conventional enzyme-linked immunosorbent assay for protein induced by vitamin K absence or antagonist II (PIVKA-II) quantification uses an antibody that is not applicable to distinguish different γ-carboxylation states. Liquid chromatography-mass spectrometry (LC-MS) approaches have been utilized to distinguish different γ-carboxylated proteoforms, however, these attempts were impeded by poor sensitivity due to spontaneous neutral loss of CO2 and simultaneous cleavage of the backbone bond in the collision cell. In this study, we utilized an alkaline mobile phase in combination with polarity switching (positive and negative ionization modes) to simultaneously identify and quantify γ-carboxylated VKDPs. The method was applied to compare Gla proteomics of prothrombin (FII) in 10 µL plasma samples of healthy control and warfarin-treated adults. We also identified surrogate non-Gla peptides for seven other VKDPs to quantify total (active plus inactive) protein levels. The total protein approach (TPA) was used to quantify absolute levels of the VKDPs in human plasma.

Peanut Glycerides.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 2001, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Peanut Glycerides is safe as a cosmetic ingredient in the practices of use and concentration as described in this report.

Erythorbic Acid and Sodium Erythorbate.

The Expert Panel for Cosmetic Ingredient Safety reviewed newly available studies since their original assessment in 1999, along with updated information regarding product types and concentrations of use, and confirmed that Erythorbic Acid and Sodium Erythorbate are safe as cosmetic ingredients in the practices of use and concentration as described in this report.

Polyamino Sugar Condensate.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 1982, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Polyamino Sugar Condensate is safe for topical application to humans in the practices of use and concentration as described in this report.

Choleth-24.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 1982, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Choleth-24 is safe for topical applications to humans in the practices of use and concentration as described in this report.

Mink Oil.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 1998, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Mink Oil is safe as a cosmetic ingredient in the practices of use and concentration as described in this report.

Glyceryl Diesters.

The Expert Panel for Cosmetic Ingredient Safety reviewed newly available studies since their original assessment in 2002, along with updated information regarding product types and concentrations of use, and confirmed that these 17 glyceryl diesters are safe as cosmetic ingredients in the practices of use and concentration as described in this report.

Dioscorea Villosa (Wild Yam) Root Extract.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 2004, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Dioscorea Villosa (Wild Yam) Root Extract is safe as a cosmetic ingredient in the practices of use and concentration as described in this report.

PEG Soy Sterols.

The Expert Panel for Cosmetic Ingredient Safety reviewed newly available studies since their original assessment in year 2000, along with updated information regarding product types and concentrations of use, and confirmed that PEG-5, -10, -16, -25, -30, and -40 Soy Sterol are safe as cosmetic ingredients in the practices of use and concentration as described in this report.

Phytantriol.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 2007, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Phytantriol is safe as a cosmetic ingredient in the practices of use and concentration as described in this report.

Acid Orange 3.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from year 2000, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Acid Orange 3 is safe for use in hair dye formulations at concentrations <.2%.

Methyl Alcohol.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 2001, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Methyl Alcohol is safe as used to denature alcohol in the practices of use and concentration as described in this report.

HC Yellow 5.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 2007, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that HC Yellow 5 is safe as a hair dye ingredient in the practices of use and concentration as described in this report.

Polyacrylamide.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 1991, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Polyacrylamide is safe as a cosmetic ingredient in the practices of use and concentration as described in this report.

Prunus Amygdalus Dulcis (Sweet Almond) Seed Meal.

The Expert Panel for Cosmetic Ingredient Safety reviewed updated information that has become available since their original assessment from 1983, along with updated information regarding product types, and frequency and concentrations of use, and reaffirmed their original conclusion that Prunus Amygdalus Dulcis (Sweet Almond) Seed Meal is safe for topical application to humans in the practices of use and concentration as described in this report.

Amyl Acetate and Isoamyl Acetate.

The Expert Panel for Cosmetic Ingredient Safety reviewed newly available studies since their original assessment in 1988, along with updated information regarding product types and concentrations of use, and confirmed that Amyl Acetate and Isoamyl Acetate are safe as cosmetic ingredients in the practices of use and concentration as described in this report.