Biomedical subjects

Kevin R Kozak

Publications and source records attributed to Kevin R Kozak.

18 recordsLinked to original sources

PubMed2007

AZD2171, a pan-VEGF receptor tyrosine kinase inhibitor, normalizes tumor vasculature and alleviates edema in glioblastoma patients.

Using MRI techniques, we show here that normalization of tumor vessels in recurrent glioblastoma patients by daily administration of AZD2171-an oral tyrosine kinase inhibitor of VEGF receptors-has rapid onset, is prolonged but reversible, and has the significant clinical benefit of alleviating edema. Reversal of normalization began by 28 days, though some features persisted for as long as four months. Basic FGF, SDF1alpha, and viable circulating endothelial cells (CECs) increased when tumors escaped treatment, and circulating progenitor cells (CPCs) increased when tumors progressed after drug interruption. Our study provides insight into different mechanisms of action of this class of drugs in recurrent glioblastoma patients and suggests that the timing of combination therapy may be critical for optimizing activity against this tumor.

Brain Edema↗

PubMed2006

Accelerated partial-breast irradiation using proton beams: initial clinical experience.

PURPOSE: We present our initial clinical experience with proton, three-dimensional, conformal, external beam, partial-breast irradiation (3D-CPBI). METHODS AND MATERIALS: Twenty patients with Stage I breast cancer were treated with proton 3D-CPBI in a Phase I/II clinical trial. Patients were followed at 3 to 4 weeks, 6 to 8 weeks, 6 months, and every 6 months thereafter for recurrent disease, cosmetic outcome, toxicity, and patient satisfaction. RESULTS: With a median follow-up of 12 months (range, 8-22 months), no recurrent disease has been detected. Global breast cosmesis was judged by physicians to be good or excellent in 89% and 100% of cases at 6 months and 12 months, respectively. Patients rated global breast cosmesis as good or excellent in 100% of cases at both 6 and 12 months. Proton 3D-CPBI produced significant acute skin toxicity with moderate to severe skin color changes in 79% of patients at 3 to 4 weeks and moderate to severe moist desquamation in 22% of patients at 6 to 8 weeks. Telangiectasia was noted in 3 patients. Three patients reported rib tenderness in the treated area, and one rib fracture was documented. At last follow-up, 95% of patients reported total satisfaction with proton 3D-CPBI. CONCLUSIONS: Based on our study results, proton 3D-CPBI offers good-to-excellent cosmetic outcomes in 89% to 100% of patients at 6-month and 12-month follow-up and nearly universal patient satisfaction. However, proton 3D-CPBI, as used in this study, does result in significant acute skin toxicity and may potentially be associated with late skin (telangiectasia) and rib toxicity. Because of the dosimetric advantages of proton 3D-CPBI, technique modifications are being explored to improve acute skin tolerance.

Aged↗

PubMed2006

Dosimetric comparison of proton and photon three-dimensional, conformal, external beam accelerated partial breast irradiation techniques.

PURPOSE: To compare the dosimetry of proton and photon-electron three-dimensional, conformal, external beam accelerated partial breast irradiation (3D-CPBI). METHODS AND MATERIALS: Twenty-four patients with fully excised, Stage I breast cancer treated with adjuvant proton 3D-CPBI had treatment plans generated using the mixed-modality, photon-electron 3D-CPBI technique. To facilitate dosimetric comparisons, planning target volumes (PTVs; lumpectomy site plus 1.5-2.0 cm margin) and prescribed dose (32 Gy) were held constant. Plans were optimized for PTV coverage and normal tissue sparing. RESULTS: Proton and mixed-modality plans both provided acceptable PTV coverage with 95% of the PTV receiving 90% of the prescribed dose in all cases. Both techniques also provided excellent dose homogeneity with a dose maximum exceeding 110% of the prescribed dose in only one case. Proton 3D-CPBI reduced the volume of nontarget breast tissue receiving 50% of the prescribed dose by an average of 36%. Statistically significant reductions in the volume of total ipsilateral breast receiving 100%, 75%, 50%, and 25% of the prescribed dose were also observed. The use of protons resulted in small, but statistically significant, reductions in the radiation dose delivered to 5%, 10%, and 20% of ipsilateral and contralateral lung and heart. The nontarget breast tissue dosimetric advantages of proton 3D-CPBI were not dependent on tumor location, breast size, PTV size, or the ratio of PTV to breast volume. CONCLUSIONS: Compared to photon-electron 3D-CPBI, proton 3D-CPBI significantly reduces the volume of irradiated nontarget breast tissue. Both approaches to accelerated partial breast irradiation offer exceptional lung and heart sparing.

Aged↗

PubMed2006

Dosimetric comparison of two different three-dimensional conformal external beam accelerated partial breast irradiation techniques.

PURPOSE: We compare the dosimetry of two techniques for three-dimensional, conformal, external beam, accelerated partial breast irradiation (3D-CPBI) in the supine position. METHODS AND MATERIALS: Sixteen patients with Stage I breast cancer had PBI treatment plans generated using the multiple, noncoplanar photon field technique and the three-field, mixed-modality technique. Planning target volumes (PTVs; lumpectomy site plus 1.5-2.0 cm margin) and total dose (32 Gy) were held constant to facilitate dosimetric comparisons. Plans were optimized for conformality and PTV coverage. RESULTS: Mixed-modality plans employed fewer fields than multiple, noncoplanar photon field plans (mean 3.2 vs. 4.1). Both techniques provided comparable PTV coverage and in all cases, 95% of the PTV received 90% of the prescribed dose. Volumes of ipsilateral breast receiving greater than 16 Gy were similar; however, the mean volume of ipsilateral breast receiving 8 Gy was significantly lower for mixed-modality plans (58% vs. 66%). No differences in the volumes of ipsilateral lung or heart receiving greater than 5 Gy were observed, however, the mixed-modality technique delivered 2.5 Gy to larger volumes of these organs. CONCLUSIONS: Both techniques for supine position, 3D-CPBI provides excellent normal tissue sparing with adequate PTV coverage. The multiple, noncoplanar photon field technique exposes smaller volumes of ipsilateral lung and heart to low dose radiation at the expense of increased plan complexity and larger irradiated breast volumes.

Adult↗

PubMed2006

Accelerated partial breast irradiation using proton beams: Initial dosimetric experience.

PURPOSE: The unique dosimetric features of proton radiotherapy make it an attractive modality for normal tissue sparing. We present our initial experience with protons for three-dimensional, conformal, external-beam accelerated partial breast irradiation (3D-CPBI). METHODS AND MATERIALS: From March 2004 to June 2005, 25 patients with tumors < or =2 cm and negative axillary nodes were treated with proton 3D-CPBI. The prescribed dose was 32 Cobalt Gray Equivalents (CGE) in 4 CGE fractions given twice daily. One to three fields were used to provide adequate planning target volume (PTV) coverage and dose homogeneity. RESULTS: Excellent PTV coverage and dose homogeneity were obtained in all patients with one to three proton beams. The median PTV receiving 95% of the prescribed dose was 100%. Dose inhomogeneity exceeded 10% in only 1 patient (4%). The median volume of nontarget breast tissue receiving 50% of the prescribed dose was 23%. Median volumes of ipsilateral lung receiving 20 CGE, 10 CGE, and 5 CGE were 0%, 1%, and 2%, respectively. The contralateral lung and heart received essentially no radiation dose. Cost analysis suggests that proton 3D-CPBI is only modestly more expensive (25%) than traditional whole-breast irradiation (WBI). CONCLUSION: Proton 3D-CPBI is technically feasible, providing both excellent PTV coverage and normal tissue sparing. It markedly reduces the volume of nontarget breast tissue irradiated compared with photon-based 3D-CPBI, addressing a principle disadvantage of external-beam approaches to PBI. As proton therapy becomes more widely available, it may prove an attractive tool for 3D-CPBI.

Aged↗

PubMed2006

Initial dosimetric experience using simple three-dimensional conformal external-beam accelerated partial-breast irradiation.

PURPOSE: Several accelerated partial-breast irradiation (APBI) techniques are being investigated in patients with early-stage breast cancer. We present our initial experience using three-dimensional conformal radiation therapy (3D-CRT). METHODS AND MATERIALS: Sixty-one patients with tumors of 2 cm or less and negative axillary nodes were treated with 3D-CRT accelerated partial-breast irradiation (APBI) between August 2003 and March 2005. The prescribed radiation dose was 32 Gy in 4-Gy fractions given twice daily. Efforts were made to minimize the number of beams required to achieve adequate planning target volume (PTV) coverage. RESULTS: A combination of photons and electrons was used in 85% of patients. A three-field technique that consisted of opposed, conformal tangential photons and enface electrons was employed in 43 patients (70%). Nine patients (15%) were treated with a four-field arrangement, which consisted of three photon fields and enface electrons. Mean PTV volumes that received 100%, 95%, and 90% of the prescribed dose were 93% +/- 7%, 97% +/- 4%, and 98% +/- 2%, respectively. Dose inhomogeneity exceeded 10% in only 7 patients (11%). Mean doses to the ipsilateral lung and heart were 1.8 Gy and 0.8 Gy, respectively. CONCLUSIONS: Simple 3D-CRT techniques of APBI can achieve appropriate PTV coverage while offering significant normal-tissue sparing. Therefore, this noninvasive approach may increase the availability of APBI to patients with early-stage breast cancer.

Adult↗

PubMed2006

Hypertrophic osteoarthropathy pathogenesis: a case highlighting the potential role for cyclo-oxygenase-2-derived prostaglandin E2.

BACKGROUND: A 65-year-old woman presented with weakness, 9 kg weight loss, dysphagia, facial and bilateral upper-extremity swelling, and debilitating, bilateral lower-extremity pain. The patient had undergone a right upper lobectomy for a 5 mm, poorly differentiated adenocarcinoma of the lung 4 years previously. Medical history included chronic obstructive pulmonary disease (emphysema), hypertension, cerebrovascular disease and multinodular goiter. Surgical history included a right carotid endarterectomy. The patient's history was remarkable for 50+ pack-years of smoking. INVESTIGATIONS: Physical examination, comprehensive metabolic panel and complete blood counts, CT, bone scintigraphy, quantification of urinary 11a-hydroxy-9,15-dioxo-2,3,4,5-tetranor-prostane-1,20-dioic acid (also known as PGE-M). DIAGNOSIS: Recurrent non-small-cell lung cancer with adrenal metastasis, hypertrophic osteoarthropathy associated with non-small-cell lung cancer, and hyperprostaglandinuria. MANAGEMENT: Rofecoxib 25 mg daily for hypertrophic osteoarthropathy, palliative external-beam radiation (44 Gy in 22 fractions) for mediastinal mass, palliative external-beam radiation (30 Gy in 12 fractions), followed 2 years later with radiofrequency ablation, for left adrenal metastasis.

Aged↗

PubMed2006

Combined vascular endothelial growth factor-targeted therapy and radiotherapy for rectal cancer: theory and clinical practice.

Despite the routine use of adjuvant and neoadjuvant chemoradiotherapy, patients with advanced rectal tumors experience significant rates of treatment failure and disease recurrence. Resistance to radiation is a particular problem. Adding a vascular endothelial growth factor (VEGF)-targeted therapy may improve outcomes in these patients. Epidemiologic studies have shown that tumor expression of VEGF predicts disease recurrence and lower overall survival in patients treated with radiation. In tumor xenograft models in mice, VEGF-targeted agents increase the response to radiation, with a greater probability of tumor control and a greater delay in tumor growth. In addition to killing cancer cells indirectly by damaging tumor blood vessels (antivascular effect), VEGF-targeted therapy may sensitize tumors to radiation through two mechanisms: by normalizing the tumor vasculature, leading to greater tumor oxygenation, and thereby increasing the cytotoxicity of radiation to cancer cells, and by increasing the radiosensitivity of tumor-associated endothelial cells. In addition, anti-VEGF agents may inhibit the regrowth of tumors after radiation by decreasing the number of circulating endothelial cells and endothelial progenitor cells. A phase I dose-escalation study has shown the safety of bevacizumab at a dose of 5 mg/kg in combination with 5-fluorouracil and radiation in patients with rectal carcinoma, and has provided evidence of both vascular normalization and antivascular mechanisms. Phase II evaluation of bevacizumab in this setting is under way.

Animals↗

PubMed2004

The glyceryl ester of prostaglandin E2 mobilizes calcium and activates signal transduction in RAW264.7 cells.

Glyceryl prostaglandins (PG-Gs) are generated by the oxygenation of the endocannabinoid, 2-arachidonylglycerol, by cyclooxygenase 2. The biological consequences of this selective oxygenation are uncertain because the cellular activities of PG-Gs have yet to be defined. We report that the glyceryl ester of PGE(2), PGE(2)-G, triggers rapid, concentration-dependent Ca(2+) accumulation in a murine macrophage-like cell line, RAW264.7. Ca(2+) mobilization is not observed after addition of PGE(2), PGD(2)-G, or PGF(2alpha)-G but is observed after addition of PGF(2alpha). Moreover, PGE(2)-G, but not PGE(2), stimulates a rapid but transient increase in the levels of inositol 1,4,5-trisphosphate (IP(3)) as well as the membrane association and activation of PKC. PGE(2)-G induces a concentration-dependent increase in the levels of phosphorylated extracellular signal regulated kinases 1 and 2 through a pathway that requires the activities of PKC, IP(3) receptor, and phospholipase C beta. The results indicate that PGE(2)-G triggers Ca(2+) mobilization, IP(3) synthesis, and activation of PKC in RAW264.7 macrophage cells at low concentrations. These responses are independent of the hydrolysis of PGE(2)-G to PGE(2).

Animals↗

PubMed2004

Studies on the metabolism of the novel, selective cyclooxygenase-2 inhibitor indomethacin phenethylamide in rat, mouse, and human liver microsomes: identification of active metabolites.

The metabolism of 2-[1-(4-chlorobenzoyl)-5-methoxy-2-methyl-1H-indol-3-yl]-N-phenethyl-acetamide (indomethacin phenethylamide, LM-4108), a highly selective cyclooxygenase-2 inhibitor, was studied in rat, mouse, and human liver microsomes. The primary site of oxidation in all species examined was on the methylene carbons of the phenethyl side chain to form the 1'- and 2'-hydroxy and 2'-oxo metabolites as determined by electrospray ionization liquid chromatography-tandem mass spectrometry. Half-lives for the disappearance of 10 microM LM-4108 in rat, human, and mouse liver microsomes (0.15 pmol P450/ml) were 11 min, 21 min, and 51 min, respectively. Indomethacin formation was not observed in incubations with rat, mouse, or human liver microsomes. Both the 2'-hydroxy-LM-4108 and 2'-oxo-LM-4108 metabolites were synthesized and found to be equipotent to the parent compound with regard to COX-2 inhibitory potency and selectivity [2'-hydroxy-LM-4108: IC(50)(COX-2) = 0.06 microM, IC(50)(COX-1) >66 microM; 2'-oxo-LM-4108: IC(50)(COX-2) = 0.05 microM, IC(50)(COX-1) >66 microM]. The formation of the metabolites was strongly inhibited by specific CYP3A4 inhibitors ketoconazole and troleandomycin but not by other isoform-selective inhibitors. These findings were confirmed by demonstrating that cloned, expressed CYP3A4 catalyzed side chain oxidation. O-Demethylation was a minor oxidative pathway in contrast to the metabolism of indomethacin and was catalyzed by CYP2D6. Upon intravenous administration of LM-4108 to Sprague-Dawley rats, oxidative metabolism on the phenethyl side chain constituted the rate-limiting steps in its clearance. The active metabolites, 2'-oxo- and 2'-hydroxy-LM-4108, as well as 1'-hydroxy-LM-4108, were all observed in rat plasma and thus may contribute to COX-2 inhibition in vivo. The glucuronides of 2'hydroxy-LM-4108 and O-desmethyl-2'-hydroxy-LM-4108 were also identified in rat bile.

Animals↗

PubMed2004

Oxidative metabolism of endocannabinoids by COX-2.

The last decade has witnessed a rapid expansion in our understanding of the mammalian endogenous cannabinoid system. In just a few short years since the discovery of endogenous lipids that serve as cannabinoids in vivo, these molecules have been shown to participate in a broad array of physiological and pathological processes. Consequently, attention has been directed at defining the proteins responsible for endocannabinoid synthesis, transport, and metabolism. Recently, multiple fatty acid oxygenases including, most notably, cyclooxygenase-2 (COX-2), have been implicated in endocannabinoid metabolism. This review will highlight connections between COX-2 and the endogenous cannabinoid system. The available biochemical evidence supporting a role for COX-2 in endocannabinoid metabolism will be presented. Finally, the potential biological consequences of COX-2-mediated endocannabinoid oxygenation will be discussed.

Binding Sites↗

PubMed2003

A novel mechanism of cyclooxygenase-2 inhibition involving interactions with Ser-530 and Tyr-385.

A variety of drugs inhibit the conversion of arachidonic acid to prostaglandin G2 by the cyclooxygenase (COX) activity of prostaglandin endoperoxide synthases. Several modes of inhibitor binding in the COX active site have been described including ion pairing of carboxylic acid containing inhibitors with Arg-120 of COX-1 and COX-2 and insertion of arylsulfonamides and sulfones into the COX-2 side pocket. Recent crystallographic evidence suggests that Tyr-385 and Ser-530 chelate polar or negatively charged groups in arachidonic acid and aspirin. We tested the generality of this binding mode by analyzing the action of a series of COX inhibitors against site-directed mutants of COX-2 bearing changes in Arg-120, Tyr-355, Tyr-348, and Ser-530. Interestingly, diclofenac inhibition was unaffected by the mutation of Arg-120 to alanine but was dramatically attenuated by the S530A mutation. Determination of the crystal structure of a complex of diclofenac with murine COX-2 demonstrates that diclofenac binds to COX-2 in an inverted conformation with its carboxylate group hydrogen-bonded to Tyr-385 and Ser-530. This finding represents the first experimental demonstration that the carboxylate group of an acidic non-steroidal anti-inflammatory drug can bind to a COX enzyme in an orientation that precludes the formation of a salt bridge with Arg-120. Mutagenesis experiments suggest Ser-530 is also important in time-dependent inhibition by nimesulide and piroxicam.

Animals↗

PubMed2003

Amino acid determinants in cyclooxygenase-2 oxygenation of the endocannabinoid anandamide.

The endocannabinoid arachidonylethanolamide (AEA, anandamide) is an endogenous ligand for the cannabinoid receptors and has been shown to be oxygenated by cyclooxygenase-2 (COX-2). We examined the structural requirements for COX-mediated, AEA oxygenation using a number of substrate analogues and site-directed mutants of COX-2. Fourteen AEA analogues were synthesized and tested as COX substrates. These studies identified the hydroxyl moiety of AEA as a critical determinant in the ability of COX enzymes to effect robust endocannabinoid oxygenation. In addition, these studies suggest that subtle structural modifications of AEA analogues near the ethanolamide moiety can result in pronounced changes in their ability to serve as COX-2 substrates. Site-directed mutagenesis studies have permitted the development of a model of AEA binding within the COX-2 active site. As with arachidonic acid, the omega-terminus of AEA binds in a hydrophobic alcove near the top of the COX-2 active site. The polar ethanolamide moiety of AEA, like the carboxylate of arachidonate, interacts with Arg-120 at the bottom of the COX-2 active site. Mutation of Tyr-385 prevents AEA oxygenation, suggesting that, as in the case of other COX substrates, AEA metabolism is initiated by Tyr-385-mediated hydrogen abstraction. Thus, AEA binds within the COX-2 active site in a conformation roughly similar to that of arachidonic acid. However, important differences have been identified that account for the isoform selectivity of AEA oxygenation. Importantly, the COX-2 side pocket and Arg-513 in particular are critical determinants of the ability of COX-2 to efficiently generate prostaglandin H(2) ethanolamide. The reduced efficiency of COX-1-mediated, AEA oxygenation can thus be explained by the absence of an arginine residue at position 513 in this isoform. Mutational analysis of Leu-531, an amino acid located directly across from the COX-2 side pocket, suggests that AEA is shifted away from this hydrophobic residue and toward Arg-513 relative to arachidonic acid. Coupled with earlier observations with the endocannabinoid 2-arachidonylglycerol, these results indicate that one possible function of the highly conserved COX-2 active site side pocket is to promote endocannabinoid oxygenation.

Acetylation↗

PubMed2003

Hand-assisted laparoscopic radical nephrectomy-associated rhabdomyolysis with ARF.

Intraoperative rhabdomyolysis with resultant acute renal failure is a rare complication seen, most commonly, with urologic surgical procedures. Since the early 1990s, the refinement of laparoscopic techniques has permitted their application more broadly. Among the procedures to benefit from these less invasive surgical methods has been radical nephrectomy. In general, this has resulted in less postoperative pain and shorter convalescence. Nonetheless, laparoscopic radical nephrectomy still represents major surgery and is not free from operative risks. To highlight one of these risks, we present a case of a young, obese man with renal cell carcinoma who underwent a hand-assisted laparoscopic radical nephrectomy that was complicated by rhabdomyolsis resulting in acute renal failure (ARF). We discuss the clinical insults that contributed to the development of azotemia with particular attention paid to our current understanding of the pathogenesis of myoglobinuric ARF. In addition, we review the literature concerning surgery-associated, rhabdomyolytic ARF with the aim of providing clinicians guidance for the avoidance and early recognition of this rare, but very serious, surgical complication.

Acute Kidney Injury↗

PubMed2002

Metabolism of the endocannabinoids, 2-arachidonylglycerol and anandamide, into prostaglandin, thromboxane, and prostacyclin glycerol esters and ethanolamides.

Cyclooxygenase-2 (COX-2) action on the endocannabinoids, 2-arachidonylglycerol (2-AG) and anandamide (AEA), generates prostaglandin glycerol esters (PG-G) and ethanolamides (PG-EA), respectively. The diversity of PG-Gs and PG-EAs that can be formed enzymatically following COX-2 oxygenation of endocannabinoids was examined in cellular and subcellular systems. In cellular systems, glycerol esters and ethanolamides of PGE(2), PGD(2), and PGF(2alpha) were major products of the endocannabinoid-derived COX-2 products, PGH(2)-G and PGH(2)-EA. The sequential action of purified COX-2 and thromboxane synthase on AEA and 2-AG provided thromboxane A(2) ethanolamide and glycerol ester, respectively. Similarly, bovine prostacyclin synthase catalyzed the isomerization of the intermediate endoperoxides, PGH(2)-G and PGH(2)-EA, to the corresponding prostacyclin derivatives. Quantification of the efficiency of prostaglandin and thromboxane synthase-directed endoperoxide isomerization demonstrated that PGE, PGD, and PGI synthases catalyze the isomerization of PGH(2)-G at rates approaching those observed with PGH(2). In contrast, thromboxane synthase was far more efficient at catalyzing PGH(2) isomerization than at catalyzing the isomerization of PGH(2)-G. These results define the in vitro diversity of endocannabinoid-derived prostanoids and will permit focused investigations into their production and potential biological actions in vivo.

Animals↗

PubMed2002

Selective oxygenation of N-arachidonylglycine by cyclooxygenase-2.

Nonsteroidal anti-inflammatory drugs prevent hyperalgesia and inflammation by inhibiting the cyclooxygenase-2 (COX-2) catalyzed oxygenation of arachidonic acid to prostaglandin (PG) H(2). The lipoamino acid N-arachidonylglycine (NAGly) has also been shown to suppress tonic inflammatory pain and is naturally present at significant levels in many of the same mammalian tissues that express COX-2. Here, we report that COX-2 selectively metabolizes NAGly to PGH(2) glycine (PGH(2)-Gly) and hydroxyeicosatetraenoic glycine (HETE-Gly). Site-directed mutagenesis experiments identify the side pocket residues of COX-2, especially Arg-513, as critical determinants of the COX-2 selectivity towards NAGly. This is the first report of a charged arachidonyl derivative that is a selective substrate for COX-2. These results suggest a possible role for COX-2 in the regulation of NAGly levels and the formation of a novel class of eicosanoids from NAGly metabolism.

Animals↗

PubMed2002

Enantiospecific, selective cyclooxygenase-2 inhibitors.

Cyclooxygenase inhibition studies with novel indomethacin alkanolamides demonstrate the potential for dramatic differences in inhibitor properties conferred by subtle structural modifications. The transformation of non-selective alpha-(S)-substituted indomethacin ethanolamides to potent, COX-2 selective inhibitors by simple stereocenter inversion highlights this property.

Cyclooxygenase 2↗

PubMed2002

15-Lipoxygenase metabolism of 2-arachidonylglycerol. Generation of a peroxisome proliferator-activated receptor alpha agonist.

The recent demonstrations that cyclooxygenase-2 and leukocyte-type 12-lipoxygenase (LOX) efficiently oxygenate 2-arachidonylglycerol (2-AG) prompted an investigation into related oxygenases capable of metabolizing this endogenous cannabinoid receptor ligand. We evaluated the ability of six LOXs to catalyze the hydroperoxidation of 2-AG. Soybean 15-LOX, rabbit reticulocyte 15-LOX, human 15-LOX-1, and human 15-LOX-2 oxygenate 2-AG, providing 15(S)-hydroperoxyeicosatetraenoic acid glyceryl ester. In contrast, potato and human 5-LOXs do not efficiently metabolize this endocannabinoid. Among a series of structurally related arachidonyl esters, arachidonylglycerols serve as the preferred substrates for 15-LOXs. Steady-state kinetic analysis demonstrates that both 15-LOX-1 and 15-LOX-2 oxygenate 2-AG comparably or preferably to arachidonic acid. Furthermore, 2-AG treatment of COS-7 cells transiently transfected with human 15-LOX expression vectors or normal human epidermal keratinocytes results in the production and extracellular release of 15-hydroxyeicosatetraenoic acid glyceryl ester (15-HETE-G), establishing that lipoxygenase metabolism of 2-AG occurs in an eukaryotic cellular environment. Investigations into the potential biological actions of 15-HETE-G indicate that this lipid, in contrast to its free-acid counterpart, acts as a peroxisome proliferator-activated receptor alpha agonist. The results demonstrate that 15-LOXs are capable of acting on 2-AG to provide 15-HETE-G and elucidate a potential role for endocannabinoid oxygenation in the generation of peroxisome proliferator-activated receptor alpha agonists.

Animals↗