CYP3A probes can quantitatively predict the in vivo kinetics of other CYP3A substrates and can accurately assess CYP3A induction and inhibition

Because the rates at which therapeutics are cleared from the body can affect their effectiveness, knowing and accounting for the variables that contribute to drug clearance is of utmost importance when designing a drug dosage regimen for patients. The activities of the manifold cytochrome P450 enzymes [(CYPs), the most clinically important of which is often CYP3A] must be considered as they are essential for modification and metabolism of compounds (i.e., therapeutic, xenobiotic, etc.) prior to their excretion. To this end, much research has been expended on trying to identify and develop drug probes that accurately predict the metabolism of CYP3A substrates in individuals. Recently, Benet has written on the futility of such an enterprise; however, other researchers believe the identification of valuable predictive probes is not only possible but crucial.     

There is no evidence that mitochondria are the main source of reactive oxygen species in mammalian cells

It is often assumed that mitochondria are the main source of reactive oxygen species (ROS) in mammalian cells, but there is no convincing experimental evidence for this in the literature. What evidence there is suggests mitochondria are a significant source for ROS, which may have physiological and pathological effects. But quantitatively, endoplasmic reticulum and peroxisomes have a greater capacity to produce ROS than mitochondria, at least in liver. In most cells and physiological or pathological conditions there is a lack of evidence for or against mitochondria being the main source of cellular ROS. Mitochondria can rapidly degrade ROS and thus are potential sinks for ROS, but whether mitochondria act as net sources or sinks within cells in particular conditions is unknown.     

Cloning and characterization of a novel CYP3A1 allelic variant: analysis of CYP3A1 and CYP3A2 sex-hormone-dependent expression reveals that the CYP3A2 gene is regulated by testosterone.

A clone was isolated from a cDNA library constructed from phenobarbital-treated Wistar rat liver and proven to correspond to the full-length mRNA of a polymorphic variant of Sprague-Dawley CYP3A1. Eight nucleotide differences were detected in a single 76-nucleotide stretch and confirmed to be present in the genomic clone. They are seated in a region implicated in the definition of a substrate binding domain of the native P450. Three out of the eight nucleotide changes are nonconservative, implicating the replacement of Thr/Ala 207, Phe/Ile 213, and Ile/Val 232. This is the first report of an allelic variant of CYP3A1, a new example of interstrain P450 variability. The CYP3A subfamily is composed of several genes coding for active testosterone 6 beta-hydroxylases which are expressed in the liver. CYP3A genes are under strong and distinct developmental regulation. Conversely to CYP3A1, transiently expressed in immature animals, CYP3A2 is constitutively expressed in the liver early after birth and characterized by an extinction in the adult females. Castration of 90-day-old male rats causes a drastic reduction (80%) of CYP3A2 mRNA relative abundance. Administration of testosterone propionate restores the physiological levels of CYP3A2 mRNA characteristic of the male rat liver. Our results demonstrate the existence of a direct relationship between the male hormonal status and the constitutive expression of rat liver CYP3A2.     

Functional characterization of medaka CYP3A38 and CYP3A40: kinetics and catalysis by expression in a recombinant baculovirus system

Phylogenic analysis of the teleost genomic lineages has demonstrated the precedent for multiple genome duplications. Among many of the genes duplicated, cytochrome P450 genes have undergone independent diversification, which can be traced to a single ancestral gene. In teleosts, cytochrome P450s, from all major families, have been identified. Among these, the CYP3A family has been cloned in several teleost species and demonstrated to contain multiple paralogs differing in gene expression patterns and tissue distribution. Herein we characterized the catalytic and kinetic activities of two medaka CYP3A paralogs (CYP3A38 and CYP3A40) with benzyloxyresorufin (BFC), a fluorescent 3A-selective substrate, and testosterone, a known metabolic substrate for CYP3A enzymes. Recombinant CYP3A was produced using the baculovirus expression vector system in Spodoptera frugiperda (Sf9) and Trichoplusia ni (Tn5) insect cells and accounted for up to 24% of total cellular protein. Following addition of a heme-albumin conjugate to log phase cells, spectral P450 content reached a maximum of 560 and 2350 pmol/mg microsomal protein for CYP3A38 and CYP3A40, respectively. Incubations containing recombinant CYP3A, human NADPH-cytochrome P-450 oxidoreductase reductase, human cytochrome b5, and a NADPH generation system catalyzed the dealkylation of BFC and hydroxylation of testosterone with a high degree of stereoselectivity. However, efficiencies and specificities were significantly different between the two isoforms. Km and Vmax activities based on BFC-catalysis were 0.116 and 0.363 muM, and 7.95 and 7.77 nmol/min/nmol P450 for CYP3A38 and CYP3A40, respectively. CYP3A38 preferentially catalyzed testosterone hydroxylation at the 6beta-, 2beta- and 16beta-positions with minor hydroxylation at other positions within the steroid nucleus. Testosterone catalysis with CYP3A40 was limited predominantly to the 6beta- and 2beta-positions. Putative identification of CYP3A substrate recognition sites (SRS) 1-6 indicates that 12 of the 49 amino acid differences between CYP3A38 and CYP3A40 OFRs occur in SRS regions previously known to be associated with steroid hydroxylation. We suggest that differences in kinetics and catalytic activities are a result of amino acid substitutions in SRS regions 1, 3 and 5 within the CYP3A38 and CYP3A40 protein sequence.     

Dextromethorphan as an in vivo probe for the simultaneous determination of CYP2D6 and CYP3A activity

Dextromethorphan (DM) is O-demethylated into dextrorphan (DEX) in humans by the cytochrome P450 designated as CYP2D6 and N-demethylated into 3-methoxymorphinan (3MM) via CYP3As. Clinically, DM has been successfully used as an index of CYP2D6 and this paper describes analytical and clinical data that will help evaluate the use of DM hydrobromide as a probe of CYP3A activity. DM and its three demethylated metabolites were measured in a 4-h spot urine sample using a HPLC method employing solid-phase extraction (C₁₈), analysis on a phenyl column [mobile phase, methanol-acetonitrile-phosphate buffer (10 mM, pH 3.5, 20:25:55, v/v)] and fluorescence detection (excitation at λ=228 nm, no emmission cut-off filter). The urinary molar ratio DM-DEX was used to assess CYP2D6 activity while DM-3MM was used for CYP3As. The DM-3MM ratios were sensitive to the co-administration of selective CYP3A inhibitors grapefruit juice and erythromycin. In addition, in healthy volunteers and cancer patients, the N-demethylation of DM correlated with the CYP3A-mediated metabolism of verapamil and tamoxifen. DM appears to be a promising way to simultaneously phenotype patients for CYP2D6 and CYP3As.     

Temporal kinetics and concentration-response relationships for induction of CYP1A, CYP2B, and CYP3A in primary cultures of beagle dog hepatocytes

Compared to other species, little information is available on the xenobiotic-induced regulation of cytochrome P450 enzymes in the beagle dog. Dogs are widely used in the pharmaceutical industry for many study types, including those that will impact decisions on compound progression. The purpose of this study was (1) to determine the temporal kinetics of drug-induced changes in canine CYP1A, CYP2B, and CYP3A mRNA and enzymatic activity, and (2) to characterize concentration-response relationships for CYP1A2, CYP2B11, and CYP3A12 using primary cultures of canine hepatocytes treated with beta-naphthoflavone (BNF), phenobarbital (PB), and rifampin (RIF), respectively. CYP1A1 and CYP1A2 mRNA exhibited maximal expression (12,700-fold and 206-fold, respectively) after 36 h of treatment with BNF. PB treatment, but not RIF treatment, caused maximal induction of CYP2B11 mRNA (149-fold) after 48 h of treatment. CYP3A12 and CYP3A26 mRNA levels were increased maximally after 72 h of treatment with PB and RIF (CYP3A12, 35-fold and 18-fold, and CYP3A26, 72-fold and 22-fold with PB and RIF treatment, respectively). Concentration-response relationships for BNF induced 7-ethoxyresorufin O-dealkylation (EROD) (EC(50) = 7.8 +/- 4.2 microM), PB induced 7-benzyloxyresorufin O-dealkylation (BROD) (EC(50) = 123 +/- 30 microM), and PB and RIF induced testosterone 6beta-hydroxylation (EC(50) = 132 +/- 28 microM and 0.98 +/- 0.16 microM) resembled the relationship for human CYP induction compared to that of rodent. Interestingly, RIF had no effect on CYP2B11 expression, which represents a species difference overlooked in previous investigations. Overall, the induction of dog CYP1A, CYP2B, and CYP3A exhibits characteristics that are intermediate to those of rodent and human.     

The decreased in vivo clearance of CYP2D6 substrates by CYP2D6*10 might be caused not only by the low-expression but also by low affinity of CYP2D6

CYP2D6 exhibits genetic polymorphism with interindividual differences in metabolic activity. We have found a significant influence on the pharmacokinetics of venlafaxine by the CYP2D6*10 allele in a Japanese population. CYP2D6.10, which is translated from CYP2D6*10, has two amino acid substitutions: Pro34 --> Ser and Ser486 --> Thr. In this study, CYP2D6.10 was expressed in Saccharomyces cerevisiae and its catalytic activity for CYP2D6 substrates was investigated. The CYP2D6*10B- and *10C-associated cDNA were isolated from human lymphocyte genotyped as CYP2D6*10. In addition, three forms of CYP2D6, Pro34/Thr486 (PT), Ser34/Ser486 (SS), and Pro34/Ser486 (wild type, CYP2D6.1), were constructed by PCR-site mutagenesis to clarify the effects of the two amino-acid substitutions. The expression of CYP2D6 protein was confirmed by immunoblotting using CYP2D antibody. The absorbance at 450 nm was measured by CO-reduced difference spectra from five all microsome preparations. The CYP2D6 forms with Pro34 --> Ser amino acid substitution were at a lower expression than CYP2D6.1 from the findings of immunoblotting and spectral analysis. The apparent K(m) values of CYP2D6.1, CYP2D6.10A, and CYP2D6.10C were 1.7, 8.5, and 49.7 microM, respectively, for bufuralol 1'-hydroxylation, and 9.0, 51.9, and 117.4 microM, respectively, for venlafaxine O-demethylation, respectively. The V(max) values were not significantly different among the three variants. These findings suggest that the decreased in vivo clearance by CYP2D6*10 was caused not only by low expression of but also the increased K(m) value of CYP2D6.     

Comparison of three fluorescent CYP3A substrates in two vertebrate models: pig and Atlantic salmon

We investigated in vitro inhibitory effects of ketoconazole (KTZ) on cytochrome P450 activity in microsomes from pigs and Atlantic salmon. The following enzymatic reactions were studied: 7-benzyloxyresorufin and 7-ethoxyresorufin O-dealkylation (BROD and EROD, respectively), 7-benzyloxy-4-trifluoromethylcoumarin O-debenzylation (BFCOD) and 7-benzyloxyquinoline O-debenzylation (BQOD). KTZ was a potent non-competitive inhibitor of BROD and BQOD in the microsomes from pigs, whereas in the microsomes from Atlantic salmon, these reactions were competitively inhibited by KTZ. BFCOD activity was inhibited by KTZ in a non-competitive manner in both species. KTZ non-competitively inhibited EROD in Atlantic salmon, but not in porcine microsomes. The activity of BROD and BQOD was higher in male than that in female pigs, but the activity of BFCOD showed no sex-related differences.     

Effects of human cytochrome b5 on CYP3A4 activity and stability in vivo

Cytochrome P450s (P450) form a superfamily of membrane-bound proteins that play a key role in the primary metabolism of both xenobiotics and endogenous compounds such as drugs and hormones, respectively. To be enzymically active, they require the presence of a second membrane-bound protein, NADPH P450 reductase, which transfers electrons from NADPH to the P450. Because of the diversity of P450 enzymes, much of the work on individual forms has been carried out on purified proteins, in vitro, which requires the use of complex reconstitution mixtures to allow the P450 to associate correctly with the NADPH P450 reductase. There is strong evidence from such reconstitution experiments that, when cytochrome b5 is included, the turnover of some substrates with certain P450s is increased. Here we demonstrate that allowing human P450 reductase, CYP3A4, and cytochrome b5 to associate in an in vivo-like system, by coexpressing all three proteins together in Escherichia coli for the first time, the turnover of both nifedipine and testosterone by CYP3A4 is increased in the presence of cytochrome b5. The turnover of testosterone was increased by 166% in whole cells and by 167% in preparations of bacterial membranes. The coexpression of cytochrome b5 also resulted in the stabilization of the P450 during substrate turnover in whole E. coli, with 109% of spectrally active CYP3A4 remaining in cells after 30 min in the presence of cytochrome b5 compared with 43% of the original P450 remaining in cells in the absence of cytochrome b5.     

Members of a mammalian SNARE complex interact in the endoplasmic reticulum in vivo and are found in COPI vesicles

Retrograde traffic between the Golgi apparatus and the endoplasmic reticulum (ER) is largely mediated by COPI-coated transport vesicles. In mammalian cells, retrograde traffic can pass through an intermediate compartment. Here, we report that the mammalian soluble N-ethylmaleimide-sensitive factor (NSF) attachment receptor (SNARE) proteins mSec22b, mUse1/D12, mSec20/BNIP1, and syntaxin 18 form a quaternary SNARE complex. Fluorescence resonance energy transfer (FRET) experiments prove that these interactions occur in the ER of living cells. In addition, mUse1/D12 and mSec20/BNIP1 form homo-oligomers in vivo. Furthermore, we show that mSec22b, mUse1/D12, mSec20/BNIP1, and syntaxin 18 are recruited into COPI-coated vesicles formed in vitro. Immunogold electron microscopy confirmed that these SNARE proteins colocalize with the KDEL receptor ERD2 in COPI-coated vesicles. Moreover, both FRET and immunoprecipitation experiments reveal interactions of these SNAREs with both ERD2 and COPI subunits. We conclude that the SNAREs described here are sorted via interaction with components of the COPI-dependent budding complex into Golgi-to-ER retrograde COPI vesicles and function in retrograde transport from the Golgi to the ER Golgi intermediate compartment (ERGIC) or the ER.     

Regulation of the expression of two female-predominant CYP3A mRNAs (CYP3A41 and CYP3A44) in mouse liver by sex and growth hormones

A second female-predominant murine CYP3A, CYP3A44, was isolated from liver and its mRNA expression was compared with that of the previously described CYP3A41. The expression of CYP3A44 was relatively constant after birth in females, whereas it gradually declined in males after 5 weeks of age. The expression of CYP3A41 increased with age in females after 3 weeks of age, whereas it gradually declined in males after 5 weeks of age. Hypophysectomy and growth hormone replacement indicated that expression of both CYP3A mRNAs in females was dependent on the feminine plasma growth hormone profile. Estradiol induced the expression of both mRNAs and the effect was dependent on the presence of the pituitary gland. These observations suggest that endocrine control of expression might be similar, but not identical, for two female-predominant CYP3A mRNAs.     

Protein disulfide isomerase and glutathione are alternative substrates in the one Cys catalytic cycle of glutathione peroxidase 7

Background

Mammalian GPx7 is a monomeric glutathione peroxidase of the endoplasmic reticulum (ER), containing a Cys redox center (CysGPx). Although containing a peroxidatic Cys (C_P) it lacks the resolving Cys (C_R), that confers fast reactivity with thioredoxin (Trx) or related proteins to most other CysGPxs.

Methods

Reducing substrate specificity and mechanism were addressed by steady-state kinetic analysis of wild type or mutated mouse GPx7. The enzymes were heterologously expressed as a synuclein fusion to overcome limited expression. Phospholipid hydroperoxide was the oxidizing substrate. Enzyme–substrate and protein–protein interaction were analyzed by molecular docking and surface plasmon resonance analysis.

Results

Oxidation of the C_P is fast (k_+ 1 > 10³ M^− 1 s^− 1), however the rate of reduction by GSH is slow (k′_+ 2 = 12.6 M^− 1 s^− 1) even though molecular docking indicates a strong GSH–GPx7 interaction. Instead, the oxidized C_P can be reduced at a fast rate by human protein disulfide isomerase (HsPDI) (k_+ 1 > 10³ M^− 1 s^− 1), but not by Trx. By surface plasmon resonance analysis, a K_D = 5.2 μM was calculated for PDI–GPx7 complex. Participation of an alternative non-canonical C_R in the peroxidatic reaction was ruled out. Specific activity measurements in the presence of physiological reducing substrate concentration, suggest substrate competition in vivo.

Conclusions

GPx7 is an unusual CysGPx catalyzing the peroxidatic cycle by a one Cys mechanism in which GSH and PDI are alternative substrates.

General significance

In the ER, the emerging physiological role of GPx7 is oxidation of PDI, modulated by the amount of GSH.     

A membrane-associated, carbohydrate-modified form of the v-abl protein that cannot be phosphorylated in vivo or in vitro.

Abelson murine leukemia virus encodes a transforming protein which contains tyrosine kinase activity and is phosphorylated in vivo and in vitro. We found that P160 and P160-derived virus strains expressed an additional, altered v-abl protein which could not be phosphorylated. The altered v-abl protein (L-v-abl) differed from the phosphorylated form (K-v-abl) in that it was glycosylated and localized exclusively to the membrane fraction. Tunicamycin inhibition of N-linked carbohydrate addition did not restore phosphorylation. It did, however, reveal that L-v-abl had additional sequences relative to K-v-abl. The coding sequences required for this region and for the expression of L-v-abl were identified by replacing sequences in the P120 virus genome, which did not express L-v-abl, with sequences from the P160 virus genome. The necessary sequences were localized to the Moloney murine leukemia virus-derived gag gene. Comparison between the in vitro altered P120 and wild-type P120 virus strains indicated that expression of L-v-abl did not increase the efficiency of lymphoid transformation. Although the biological role of L-v-abl is not clear, our analyses have revealed that a specific amino terminal gag sequence can prevent v-abl from acting as a kinase substrate and can alter the cellular localization and modification of v-abl. These properties distinguish L-v-abl from previously reported v-abl proteins.     

‘There are no chickens in suicide vests’: the decoupling of human rights and animal rights in Israel

In this article, I consider the shifting politics of animal rights activism in Israel in relation to human rights activism. I find that whereas in the past, human and animal rights activism were tightly linked, today they have become decoupled, for reasons I explore in this article. Although human and animal rights activism once shared social and ideological foundations in Israeli society, today much of the current animal rights activism is assertive and explicit in its disregard for human rights issues, such as the ongoing occupation of Palestine and the treatment of Palestinians. This decoupling has been heightened by the appropriation of animal rights politics by a right‐wing state for the purposes of ethical legitimation. This article considers the dilemmas of ethical responsibilities towards humans and animals as it plays out in one of the most vexed political environments in the world. I consider the shifting politics of human and animal rights activism, and demonstrate how they implicate and entangle each other in the context of the ongoing Israeli‐Palestinian conflict. I further consider what the decoupling of the human and animal rights movements might suggest regarding the ongoing academic critique of human rights and humanism.     

Involvement of CYP3A4 and CYP2D6 in the Metabolism of Haloperidol

1. Human cytochrome P450 (CYP) isoenzymes expressed in a human cell line were used to elucidate their involvement in the metabolism of haloperidol (HAL).2. It was found that CYP3A4 catalyzes the metabolism of HAL to HAL 1,2,3,6-tetrahydropyridine (HTP). HTP is further metabolized to HAL pyridinium (HP⁺) by both CYP3A4 and CYP2D6.3. CYP3A4 and CYP2D6 are also responsible for the N-dealkylation of HAL. The N-dealkylation of reduced HAL (RH) was observed, which is catalyzed by CYP3A4. In addition, CYP3A4 also catalyzes the oxidation of RH back to HAL.4. These results are discussed in terms of the metabolic interactions of HAL with other drugs and how this knowledge may be used to reduce the movement disorders induced by HAL.     

Allele and genotype frequencies of the polymorphic cytochrome P450 genes (CYP1A1, CYP3A4, CYP3A5, CYP2C9 and CYP2C19) in the Jordanian population

Drug metabolizing enzymes participate in the neutralizing of xenobiotics and biotransformation of drugs. Human cytochrome P450, particularly CYP1A1, CYP2C9, CYP2C19, CYP3A4 and CYP3A5, play an important role in drug metabolism. The genes encoding the CYP enzymes are polymorphic, and extensive data have shown that certain alleles confer reduced enzymatic function. The goal of this study was to determine the frequencies of important allelic variants of CYP1A1, CYP2C9, CYP2C19, CYP3A4 and CYP3A5 in the Jordanian population and compare them with the frequency in other ethnic groups. Genotyping of CYP1A1(m1 and m2), CYP2C9 (*2 and *3), CYP2C19 (*2 and *3), CYP3A4*5, CYP3A5 (*3 and *6), was carried out on Jordanian subjects. Different variants allele were determined using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). CYP1A1 allele frequencies in 290 subjects were 0.764 for CYP1A1*1, 0.165 for CYP1A1*2A and 0.071 for CYP1A1*2C. CYP2C9 allele frequencies in 263 subjects were 0.797 for CYP2C9*1, 0.135 for CYP2C9*2 and 0.068 for CYP2C9*3. For CYP2C19, the frequencies of the wild type (CYP2C19*1) and the nonfunctional (*2 and *3) alleles were 0.877, 0.123 and 0, respectively. Five subjects (3.16?%) were homozygous for *2/*2. Regarding CYP3A4*1B, only 12 subjects out of 173 subjects (6.9?%) were heterozygote with none were mutant for this polymorphism. With respect to CYP3A5, 229 were analyzed, frequencies of CYP3A5*1,*3 and *6 were 0.071, 0.925 and 0.0022, respectively. Comparing our data with that obtained in several Caucasian, African-American and Asian populations, Jordanians are most similar to Caucasians with regard to allelic frequencies of the tested variants of CYP1A1, CYP2C9, CYP2C19, CYP3A4 and CYP3A5.     

Genetic evidence that the two isozymes of sucrose synthase present in developing maize endosperm are critical, one for cell wall integrity and the other for starch biosynthesis

In maize, two paralogous genes, Sh1 and Sus1, encode two biochemically similar isozymes of sucrose synthase, SS1 and SS2, respectively. Previous studies have attributed the mild starch deficiency of the shrunken1 (sh1) endosperm to the loss of the SS1 isozyme in the mutant. Here we describe the first mutation in the sucrose synthase1 (Sus1) gene, sus1-1, and the isolation of a double recessive genotype, sh1 sus1-1. Combined data from diverse studies, including Northern and Western analyses, RT-PCR and genomic PCR, cloning and sequencing data for the 3′ region, show that the mutant sus1-1 gene has a complex pattern of expression, albeit at much reduced levels as compared to the Sus1 gene. Endosperm sucrose synthase activity in sh1 sus1-1 was barely 0.5% of the total activity in the Sh1 Sus1 genotype. Significantly, comparative analyses of Sh1 Sus1, sh1 Sus1 and sh1 sus1-1 genotypes have, for the first time, allowed us to dissect the relative contributions of each isozyme to endosperm development. Starch contents in endosperm of the three related genotypes were 100, 78 and 53%, respectively. Anatomical analyses, which confirmed the previously described early cell degeneration phenotype unique to the sh1 Sus1 endosperm, revealed no detectable difference between the two sh1 genotypes. We conclude that the SS1 isozyme plays the dominant role in providing the substrate for cellulose biosynthesis, whereas the SS2 protein is needed mainly for generating precursors for starch biosynthesis.     

Genetic evidence that the two isozymes of sucrose synthase present in developing maize endosperm are critical, one for cell wall integrity and the other for starch biosynthesis

In maize, two paralogous genes, Sh1 and Sus1, encode two biochemically similar isozymes of sucrose synthase, SS1 and SS2, respectively. Previous studies have attributed the mild starch deficiency of the shrunken1 (sh1) endosperm to the loss of the SS1 isozyme in the mutant. Here we describe the first mutation in the sucrose synthase1 (Sus1) gene, sus1-1, and the isolation of a double recessive genotype, sh1 sus1-1. Combined data from diverse studies, including Northern and Western analyses, RT-PCR and genomic PCR, cloning and sequencing data for the 3′ region, show that the mutant sus1-1 gene has a complex pattern of expression, albeit at much reduced levels as compared to the Sus1 gene. Endosperm sucrose synthase activity in sh1 sus1-1 was barely 0.5% of the total activity in the Sh1 Sus1 genotype. Significantly, comparative analyses of Sh1 Sus1, sh1 Sus1 and sh1 sus1-1 genotypes have, for the first time, allowed us to dissect the relative contributions of each isozyme to endosperm development. Starch contents in endosperm of the three related genotypes were 100, 78 and 53%, respectively. Anatomical analyses, which confirmed the previously described early cell degeneration phenotype unique to the sh1 Sus1 endosperm, revealed no detectable difference between the two sh1 genotypes. We conclude that the SS1 isozyme plays the dominant role in providing the substrate for cellulose biosynthesis, whereas the SS2 protein is needed mainly for generating precursors for starch biosynthesis. Received: 22 January 1998 / Accepted: 30 March 1998     

Study the polymorphism of CYP3A5 and CYP3A4 loci in Iranian population with laryngeal squamous cell carcinoma

Cancer reflects a complicated network of interactions between genes and environmental factors. Cytochrome P450 (CYP) is a multi-gene superfamily participating in the metabolism of xenobiotics. The aim of our study was to examine whether polymorphisms in the CYP enzyme genes affect the risk of developing larynx squamous cell carcinoma (SCC). Polymorphism of CYP3A5 and CYP3A4 genes were investigated in 50 patients with laryngeal SCC and 100 control subjects by polymerase chain reaction- restriction fragment length polymorphism (PCR-RFLP). In patients the CYP3A5 3*/3* and 1*/3*genotypes were detected in 92% and 8% respectively. There was no relation between genotype, allele frequency and grade/stage of tumor. In control group, the frequency of CYP3A5 3*/3* and CYP3A5 1*/3* genotype were 98% and 2% respectively. There was no significant difference in genotype and allele frequency of this gene between patient and control group. In respect of CYP3A41A*/B*, people in both patient and control groups had the same genotype of CYP3A41A*/1A*. In this study, the CYP gene variants were not associated with increased risk of laryngeal SCC. Study on the other genetic factors which are involved in activation/detoxication of procarcinogenes, such as CYP1A1, CYP1B1, CYP2E1 and gluthation S transferase is recommended.