Perillyl Alcohol Treatment Research Articles

Perillyl alcohol attenuates chronic restraint stress aggravated dextran sulfate sodium-induced ulcerative colitis by modulating TLR4/NF-κB and JAK2/STAT3 signaling pathways

BackgroundUlcerative colitis (UC) is the most prevalent chronic inflammatory immune bowel disease. The modernization of lifestyle accompanied by the stress to cope with the competition has resulted in a new range of complications where stress became a critical contributing factor for many diseases, including UC. Hence there is an urgent need to develop a dual role in curtailing both systemic and neuroinflammation. Perillyl alcohol (POH) is a natural essential oil found in lavender, peppermint, cherries etc and has been widely studied for its strong anti-inflammatory, antioxidant and anti-stress properties. Hypothesis/PurposePOH regulates the various inflammatory signaling cascades involved in chronic inflammation by inhibiting farnesyltransferase enzyme. Several studies reported that POH could inhibit the phosphorylation of NF-κB, STAT3 and promote the endogenous antioxidant enzymes like Nrf2 via farnesyltransferase enzyme inhibition. Also, the effects of POH against UC is not known yet. Thus, this study aims to explore the anti-ulcerative properties of POH on stress aggravated ulcerative colitis in C57BL/6 mice. MethodsUlcerative colitis was induced by duel exposure of chronic restraint stress (day 1 to day 28) and 2.5% dextran sulphate sodium (day8 to day14) in mice. POH treatment 100 and 200 mg/kg was administred from day14 ti day28 following oral route of administration. Disease activity index, colonoscopy, western blot analysis and histological analysis, neurotransmitter analysis and Gene expression studies were perofomerd to asses the anti-colitis effects of POH. ResultsThe treatment reversed the oxidative stress and inflammatory response by inhibiting TLR4/NF-kB pathway, and IL-6/JAK2/STAT3 pathway in both isolated mice colons and brains. The inhibition of these pathways resulted in a decrease in pro-inflammatory cytokines like IL-6, IL-1β and TNF-α. The treatment improved the physiological and histological changes with decreased ulcerations as examined by colonic endoscopy and Haematoxylin and Eosin staining. The treatment also improved the behavior response as it increased mobility time which was reduced by chronic restrained stress. This was due to increased satiety neurotransmitters like dopamine and serotonin and decreased cortisol in mice brains. ConclusionThese results infer that POH has significant anti-colitis activity on chronic restraint stress aggravated DSS-induced UC in mice.

Perillyl alcohol attenuates rheumatoid arthritis via regulating TLR4/NF-κB and Keap1/Nrf2 signaling pathways: A comprehensive study onin-vitro and in-vivo experimental models

BackgroundRheumatoid arthritis is a chronic and idiopathic autoimmune disorder. Perillyl alcohol (POH) is a monoterpene which can be extracted from widely available essential oils and is known for its strong anti-inflammatory and anti-oxidant properties. Hypothesis/PurposeRecent studies have been proven that inhibitors of farnesyltransferase enzyme showed significant anti-arthritic activity. POH is one such natural molecule having anti-inflammatory and anti-oxidant properties by inhibiting farnesyltransferase enzyme which further down regulates NF-κB and Nrf2 via Ras/Raf/MAPK pathway. Also, the effect of POH against rheumatoid arthritis is not known yet. Hence, the present research was intended to assess the anti-arthritic potential of POH in-vitro and in-vivo. MethodsThe in-vitro effects of POH on RAW 264.7 cells stimulated with LPS 1 µg/ml were investigated to its potential therapeutic effects. CFA 100 µl was intradermally administered to rats for the induction of arthritis. POH 100 mg/kg and 200 mg/kg administered topically from day 1 to day 28. Paw volumes measured, radiography analysis, anti-oxidant status, Gene expression studies, western blot analysis and histological analysis were performed to check the effects of POH. ResultsOur in-vitro findings suggested that POH inhibits inflammation by suppressing reactive oxygen species (ROS), NF-кB and Nrf2 signaling axis. Besides this, POH also rescinded the nitrate levels, pro-inflammatory cytokine levels like IL-1β, IL-6 and TNF-α also PGE2 and COX-2 levels induced by LPS in murine macrophages. Additionally, our in-vivo results revealed that POH conscientiously alleviated CFA induced inflammation by restoring arthritis index, body weight, nitrosative, lipid peroxidation assays. Macroscopically through measuring paw volumes and X-ray, it was evidenced that POH has decreased inflammation and bone erosion. Not only in-vitro but also in-vivo, POH has abridged cytokine levels IL-1β, IL-6, and TNF-α. Histopathological evaluation presented POH treatment alleviated joint inflammation, pannus formation, and bone erosion significantly. Moreover, POH suppressed the protein expression of NF-кB, COX-2, iNOS and improved Nrf2, and SOD2 levels in paw tissues estimated by western blotting. ConclusionPOH was effective in ameliorating LPS stimulation mediated oxidative stress and pro-inflammatory cytokines in RAW 264.7 cells in-vitro and FCA induced arthritis in rats in-vivo through its anti-inflammatory effects via regulating TLR4/NF-κB and Keap1/Nrf2 signaling pathways..

Perillyl alcohol inhibits keratinocyte proliferation and attenuates imiquimod-induced psoriasis like skin-inflammation by modulating NF-κB and STAT3 signaling pathways

Psoriasis is a chronic inflammatory and proliferative skin disease characterized by pathological skin lesions which significantly impact the quality of life. Recent studies have been proven that inhibitors of farnesyltransferase enzyme showed significant anti-psoriatic activity. Perillyl alcohol (POH) is one such natural molecule having anti proliferative, anti-inflammatory and anti-oxidant properties by inhibiting farnesyltransferase enzyme which further down regulates NF-κB and STAT3 via Ras/Raf/MAPK pathway. Hence, in the current study we aimed to find the effect of POH on human keratinocytes (HaCat) cells in in-vitro and IMQ induced psoriatic like skin inflammation model in mice. POH significantly decreased the intracellular ROS levels and inhibited the phosphorylation of NF-κB and STAT3 in in-vitro. It was found that POH (200 mg/kg, topical application) has reduced the epidermal hyperplasia, psoriasis area and severity index (PASI) scoring; splenomegaly in imiquimod (IMQ) induced psoriatic mice. Further, POH treatment has decreased the pro-inflammatory serum cytokine levels such as IL-6, IL-12/23, TNF-α and IL-1β and also reduced the expression levels of various inflammatory proteins, COX-2, iNOS, IL-17A, IL-22, NF-кB and STAT3 evidenced by Immunoblotting studies from skin samples. The levels of endogenous antioxidants like glutathione GSH, SOD, Nrf2 were restored to normal levels upon POH treatment. POH downregulated the proteins levels of TLR7, TLR8, CyclinD1 and mRNA expression of Bcl-2 in the skin samples when compared to the IMQ group. POH has ameliorated the hyper-keratosis and acanthosis which was evidenced by histopathology. Collectively, our results suggest that POH has a promising therapeutic application for ameliorating psoriasis-like skin inflammation.

Perillyl Alcohol Attenuates NLRP3 Inflammasome Activation and Rescues Dopaminergic Neurons in Experimental In Vitro and In Vivo Models of Parkinson's Disease.

NLRP3 activation plays a key role in the initiation and progression of a variety of neurodegenerative diseases. However, understanding the molecular mechanisms involved in the bidirectional signaling required to activate the NLRP3 inflammasomes is the key to treating several diseases. Hence, the present study aimed to investigate the role of lipopolysaccharide (LPS) and hydrogen peroxide (H2O2) in activating NLRP3 inflammasome-driven neurodegeneration and elucidated the neuroprotective role of perillyl alcohol (PA) in in vitro and in vivo models of Parkinson's disease (PD). Initial priming of microglial cells with LPS following treatment with H2O2 induced NF-κB translocation to the nucleus with a robust generation of free radicals that act as signal 2 in augmenting NLRP3 inflammasome assembly and its downstream targets. PA treatment suppresses the nuclear translocation of NF-κB, enhances PARKIN translocation into the mitochondria, and maintains cellular redox homeostasis in both mouse and human microglial cells that limit NLRP3 inflammasome activation along with processing of active caspase-1, IL-1β, and IL-18. To further correlate the in vitro study with the in vivo MPTP model, treatment with PA also inhibited the nuclear translocation of NF-κB and downregulated the NLRP3 inflammasome activation. PA administration upregulated various antioxidant enzymes' levels and restored the level of dopamine and other neurotransmitters in the striatum of the mouse brain, subsequently improving the behavioral activities. Therefore, we conclude that NLRP3 inflammasome activation required a signal from damaged mitochondria for its activation. Further pharmacological scavenging of free radicals restricts microglia activation and simultaneously supports neuronal survival via targeting the NLRP3 inflammasome pathway in PD.

Perillyl alcohol as a treatment for cancer: A systematic review

BackgroundCancer is the second leading cause of death in the world. Cancer treatment has many adverse effects, making the search for new therapeutic agents necessary. Objectivethis systematic review aimed to assess the efficacy of perillyl alcohol (POH) in the treatment of cancer based on the objective response (OR). MethodsThis review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) Statement. The search was conducted in the PubMmed, Scopus, Web of Science, and Cochrane Library databases, and included clinical trials which reported in their results the anti-tumor activity of POH as an OR, and studies with isolated POH as treatment. The review excluded preclinical studies, in vitro trials, review articles, meta-analyses, abstracts, conference proceedings, editorials/letters, case reports, and studies assessing POH in other ways, not investigating it as an anticancer treatment. Data were extracted by two independent reviewers, using a table with pre-specified items. The risk of bias (RoB) in the included studies was assessed using the Cochrane risk of bias assessment tool. ResultsThirteen studies were identified, 7 phase I and 6 phase II trials. Eleven studies administrated POH by oral route and 2 by inhalation. Ten studies reported adverse effects. Among the selected studies, the cancer types most studied were colorectal, ovarian, breast, and prostate cancers. None of the selected studies achieved clinical complete response (CR) after POH treatment, three achieved partial response (PR) and several reported stable disease (SD). Of the three which achieved PR, two used the inhalation route for POH. ConclusionThe best evidence available shows that POH has little or no significant effect on objective response in different cancer types. However, the administration of POH by inhalation showed promising objective responses and outcomes. Taken together, these findings suggest a promising potential cytostatic effect of POH.

Intranasal administration of perillyl alcohol-loaded nanoemulsion and pharmacokinetic study of its metabolite perillic acid in plasma and brain of rats using ultra-performance liquid chromatography/tandem mass spectrometry.

Perillyl alcohol (POH) is a monocyclic terpene that has strong antitumor activity. Brain tumors are particularly difficult to treat with therapeutic agents, and clinical trials have shown their low tolerance through oral administration. We proposed the entrapment of POH into an oil-in-water chitosan nanoemulsion aiming its intranasal administration for brain targeting. An ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the quantitation of total metabolite perillic acid (PA) in plasma and brain of rats. The rat samples containing the metabolite were treated by liquid-liquid extraction with acetonitrile. The mobile phase was 0.1% formic acid in water (solvent A) and 0.1% formic acid in methanol (solvent B), at a flow rate of 0.3 mL min-1 in gradient elution. The chromatography was run for 10 min, and analytical curves were built in acetonitrile, plasma, and brain. The PA was detected in positive ion mode with multiple reaction monitoring. The method has shown high selectivity, sensitivity, and throughput. The low quantification limits of 162, 178, and 121 ng mL-1 for acetonitrile, brain, and plasma, respectively, indicate a good detectability of the method. The repeatability and precision observed were within the limits recommended in the literature. The accuracy of the method was verified through high recovery rates (106-118%). The validated method was successfully applied to the pharmacokinetic study of the metabolite PA after the intranasal administration of free or POH-loaded nanoemulsion in rats. The results showed that chitosan nanoemulsion improved the plasma and brain bioavailability of POH, representing a promising alternative to free POH treatment.

Adjuvant effect of low-carbohydrate diet on outcomes of patients with recurrent glioblastoma under intranasal perillyl alcohol therapy.

Background: Standard of care for glioblastoma (GB), consisting of cytotoxic chemotherapy, steroids, and high-dose radiation, induces changes in the tumor microenvironment through its effects on glucose availability, which is a determinant for tumor progression (TP). Low-carbohydrate diet (LCD) reduces the glucose levels needed to drive the Warburg effect.Methods: To investigate LCD’s effect on GB therapy, we have begun a clinical trial using LCD as an addition to intranasal perillyl alcohol (POH) for recurrent GB (rGB) patients. This study involved 29 individuals and evaluated, over a period of 1 year, the adjuvant effect of LCD associated with POH therapy in terms of toxicity, extent of peritumoral edema, reduced corticosteroid use, seizure frequency, and overall survival. POH group (n = 14) received solely intranasal POH without specific diet regimen, whereas POH/LCD group (n = 15) received intranasal POH in combination with nutritional intervention. Patients’ assessment was based on clinical reviews and magnetic resonance data.Results: In the 1-year follow-up, the POH/LCD group showed a 4.4-fold decrease in the proportion of patients who needed treatment with corticosteroids, as well as a reduction in tumor size and peritumoral edema, as compared to the POH group. While 75% of patients undergoing POH treatment experienced seizures, this fraction was reduced to 56% in the POH/LCD group. A 2.07-fold increase in the proportion of patients with stable disease, along with a 2.8-fold decrease in the proportion of patients with TP, was seen in the POH/LCD group.Conclusion: The results presented in this study show that the LCD associated with intranasal POH therapy may represent a viable option as adjunctive therapy for rGB to improve survival without compromising patients’ quality of life. Prospective cohort studies are needed to confirm these findings and validate the efficacy of this novel therapeutic strategy.

Perillyl Alcohol Mitigates Behavioural Changes and Limits Cell Death and Mitochondrial Changes in Unilateral 6-OHDA Lesion Model of Parkinson’s Disease Through Alleviation of Oxidative Stress

In this study, we aim to assess the phytomedicinal potential of perillyl alcohol (PA), a dietary monoterpenoid, in a unilateral 6-hydroxydopamine (6-OHDA) lesion rat model of Parkinson's disease (PD). We observed that PA supplementation alleviated behavioural abnormalities such as loss of coordination, reduced rearing and motor asymmetry in lesioned animals. We also observed that PA-treated animals exhibited reduced oxidative stress, DNA fragmentation and caspase 3 activity indicating alleviation of apoptotic cell death. We found reduced mRNA levels of pro-apoptotic regulator BAX and pro-inflammatory mediators IL18 and TNFα in PA-treated animals. Further, PA treatment successfully increased mRNA and protein levels of Bcl2, mitochondrial biogenesis regulator PGC1α and tyrosine hydroxylase (TH) in lesioned animals. We observed that PA treatment blocked BAX and Drp1 translocation to mitochondria, an event often associated with the inception of apoptosis. Further, 6-OHDA exposure reduced expression of electron transport chain complexes I and IV, thereby disturbing energy metabolism. Conversely, expression levels of both complexes were upregulated with PA treatment in lesioned rats. Finally, we found that protein levels of Nrf2, the transcription factor responsible for antioxidant gene expression, were markedly reduced in cytosolic and nuclear fraction on 6-OHDA exposure, and PA increased expression of Nrf2 in both fractions. We believe that our data hints towards PA having the ability to provide cytoprotection in a hemiparkinsonian rat model through alleviation of motor deficits, oxidative stress, mitochondrial dysfunction and apoptosis.

Intranasal perillyl alcohol therapy improves survival of patients with recurrent glioblastoma harboring mutant variant for MTHFR rs1801133 polymorphism

BackgroundPolymorphisms in MTHFR gene influence risk and overall survival of patients with brain tumor. Global genomic DNA (gDNA) methylation profile from tumor tissues is replicated in peripheral leukocytes. This study aimed to draw a correlation between rs1801133 MTHFR variants, gDNA methylation and overall survival of patients with recurrent glioblastoma (rGBM) under perillyl alcohol (POH) treatment.MethodsgDNA from whole blood was extracted using a commercially available kit (Axygen) and quantified by spectrophotometry. Global gDNA methylation was determined by ELISA and rs1801133 polymorphism by PCR-RFLP. Statistical analysis of gDNA methylation profile and rs1801133 variants included Mann-Whitney, Kruskal-Wallis, Spearman point-biserial correlation tests (SPSS and Graphpad Prism packages; significant results for effect size higher than 0.4). Prognostic value of gDNA methylation and rs1801133 variants considered survival profiles at 25 weeks of POH treatment, having the date of protocol adhesion as starting count and death as the final event.ResultsMost rGBM patients showed global gDNA hypomethylation (median = 31.7%) and a significant, moderate and negative correlation between TT genotype and gDNA hypomethylation (median = 13.35%; rho = − 0.520; p = 0.003) compared to CC variant (median = 32.10%), which was not observed for CT variant (median = 33.34%; rho = − 0.289; p = 0.06). gDNA hypermethylated phenotype (median = 131.90%) exhibited significant, moderate and negative correlations between TT genotype (median = 112.02%) and gDNA hypermethylation levels when compared to CC (median = 132.45%; rho = − 0,450; p = 0.04) or CT (median = 137.80%; rho = − 0.518; p = 0.023) variants. TT variant of rs1801133 significantly decreased gDNA methylation levels for both patient groups, when compared to CC (d values: hypomethylated = 1.189; hypermethylated = 0.979) or CT (d values: hypomethylated = 0.597; hypermethylated = 1.167) variants. Positive prognostic for rGBM patients may be assigned to gDNA hypermethylation for survivors above 25 weeks of treatment (median = 88 weeks); and TT variant of rs1801133 regardless POH treatment length.ConclusionrGBM patients under POH-based therapy harboring hypermethylated phenotype and TT variant for rs1801133 had longer survival. Intranasal POH therapy mitigates detrimental effects of gDNA hypomethylation and improved survival of patients with rGBM harboring TT mutant variant for MTHFR rs1801133 polymorphism.Trial registrationCONEP -9681- 25,000.009267 / 2004. Registered 12th July, 2004.

Efficacy of a ketogenic diet with concomitant intranasal perillyl alcohol as a novel strategy for the therapy of recurrent glioblastoma.

It has been hypothesized that persistent ketotic hypoglycemia represents a potential therapeutic strategy against high-grade gliomas. Perillyl alcohol (POH) is a non-toxic, naturally-occurring, hydroxylated monoterpene that exhibits cytotoxicity against temozolomide-resistant glioma cells, regardless of O6-methylguanine-methyltransferase promoter methylation status. The present study aimed to evaluate the toxicity and therapeutic efficacy of intranasal POH when administered in combination with a ketogenic diet (KD) program for the treatment of patients with recurrent glioblastoma. The 32 enrolled patients were divided into two groups, KD or standard diet, with intranasal POH treatment (n=17 and n=15, respectively). The nutritional status and anthropometric parameters of the patients were measured. Patients that adhered to the KD maintained a strict dietary regimen, in addition to receiving 55 mg POH four times daily, in an uninterrupted administration schedule for three months. Neurological examination and magnetic resonance imaging analysis were used to monitor disease progression. A total of 9/17 patients in the KD group survived and maintained compliance with the KD. After three months of well-tolerated treatment, a partial response (PR) was observed for 77.8% (7/9) of the patients, stable disease (SD) in 11.1% (1/9) and 11.1% (1/9) presented with progressive disease (PD). Among the patients assigned to the standard diet group, the PR rate was 25% (2/8 patients), SD 25% (2/8) and PD 50% (4/8 patients). The patients assigned to the KD group presented with reduced serum lipid levels and decreased low-density lipoprotein cholesterol levels. These results are encouraging and suggest that KD associated with intranasal POH may represent a viable option as an adjunct therapy for recurrent GBM.

Inhibition of perillyl alcohol on cell invasion and migration depends on the Notch signaling pathway in hepatoma cells.

Cell metastasis, especially the process of invasion and migration, is considered as the main cause for the high mortality rate of hepatocellular carcinoma (HCC), which has become the sixth most common cancer worldwide and the third leading cause of cancer death. In this present study, we aimed to exploit the effects of perillyl alcohol on cell invasion and migration and the underlying molecular mechanisms in HCC. According to the transwell assays, cell invasiveness and migratory capacity were markedly higher in hepatoma cells (HepG2, SMMC-7721 and MHCC97H) than those in normal liver cells (HL-7702), and then significantly suppressed by perillyl alcohol treatment (P<0.05). Meanwhile, the mRNA levels of Notch signaling pathway downstream target genes, HES1, HES5, and HEY1, were notably higher in hepatoma cells detected with real-time reverse transcription polymerase chain reaction (RT-PCR) (P<0.05). After treated with perillyl alcohol, these mRNA levels were significantly decreased in hepatoma cells (P<0.05). In addition, compared with the normal liver cells, the protein expression levels of Notch1 intracellular domain (N1ICD) and Snail were significantly increased, while E-cadherin protein expression was significantly decreased in hepatoma cells (P<0.05). However, perillyl alcohol treatment significantly decreased N1ICD and Snail protein expressions and increased E-cadherin protein expression in hepatoma cells (P<0.05). In conclusion, perillyl alcohol might play an important role in the process of hepatoma cell invasion and migration via decreasing the activity of Notch signaling pathway and increasing E-cadherin expression regulated by Snail.

Perillyl alcohol improves functional and histological outcomes against ischemia–reperfusion injury by attenuation of oxidative stress and repression of COX-2, NOS-2 and NF-κB in middle cerebral artery occlusion rats

Perillyl alcohol (PA) is a monoterpene found in essential oils of mints, cherries, citreous fruits and lemon grass, reported to have antioxidant and anti-inflammatory properties. However, the role of PA in stroke is still illusive. Since oxidative stress and inflammation play a pivotal role in ischemia–reperfusion (I–R) injury, this study was designed to elucidate the potential effects of PA against I–R induced pathology in rat׳s brain. Middle cerebral artery occlusion (MCAO) for 2h followed by 22h reperfusion in Wistar male rats (250–280g, 14–16 weeks old) induced the behavioral and histological alterations along with exhausted antioxidant status and enhanced inflammatory mediators. However, PA administration (25, 50 and 100mg/kg b.wt orally once daily for 7 days) prior to MCAO significantly attenuated neurological deficits related to flexion test and spontaneous motor activity, improved grip strength and motor coordination in a dose dependent manner. PA treatment also inhibited oxidative stress in MCAO rats as evident from decreased lipid peroxidation and augmented level of reduced glutathione and restored activities of catalase, glutathione peroxidase, and glutathione reductase and thus, reduced infarct volume and protected the brain histology after I–R injury. Furthermore, PA markedly suppressed the level of proinflammatory cytokines (IL-1β, TNF α and IL-6) and down regulated expressions of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (NOS-2) and nuclear factor κB (NF-κB) in MCAO group. In conclusion, PA mediates neuroprotection against I–R injury via mitigation of oxidative stress and inflammation and thus, may be a good therapeutic approach in stroke prone patient.

Característica clínica e topográfica do glioma maligno em adolescente – Influência do tratamento intranasal com álcool perílico

Resumo Objetivo: Analisar a influência da topografia da lesão tumoral na resposta ao tratamento intranasal com álcool perílico (POH) em jovens com glioma maligno recidivo. Método: Tendo como padrão a faixa etária de 0 a 19 anos, foram incluídos pacientes do sexo masculino (#153; #31) e feminino (#178) com glioma maligno em estágio terminal, recebendo terapia de suporte paliativa e administração intranasal diária de 440 mg de POH. Resultados: Cefaleia intensa, tontura, vômito, crise convulsiva, alteração de comportamento, fraqueza muscular, alteração visual e hemiplegia à direita foram os sintomas prevalentes antes da confirmação diagnóstica de glioma. Análise de imagem mostrou lesão tumoral nas regiões troncocerebral (#153), talamomesencefálica esquerda (#178) e frontotemporal e insular direita (#31). Paciente #178 não respondeu ao tratamento, evoluindo a óbito em três semanas, e paciente #31 permaneceu em tratamento com POH por aproximadamente 54 semanas. Apesar de nova recidiva, paciente #153 apresenta doença estável, sem qualquer evidência clínica de recorrência para mais de 200 semanas em tratamento exclusivo com álcool perílico por via intranasal. Conclusão: Pacientes adolescentes com glioma maligno recidivo apresentaram heterogeneidade de sintomas compatível com a região anatômica comprometida, indicando que a topografia da lesão tumoral foi um fator prognóstico de sobrevida, influenciando inclusive na resposta ao tratamento intranasal com o álcool perílico.

Perillyl alcohol suppresses antigen-induced immune responses in the lung

Perillyl alcohol (POH) is an isoprenoid which inhibits farnesyl transferase and geranylgeranyl transferase, key enzymes that induce conformational and functional changes in small G proteins to conduct signal production for cell proliferation. Thus, it has been tried for the treatment of cancers. However, although it affects the proliferation of immunocytes, its influence on immune responses has been examined in only a few studies. Notably, its effect on antigen-induced immune responses has not been studied. In this study, we examined whether POH suppresses Ag-induced immune responses with a mouse model of allergic airway inflammation. POH treatment of sensitized mice suppressed proliferation and cytokine production in Ag-stimulated spleen cells or CD4+ T cells. Further, sensitized mice received aerosolized OVA to induce allergic airway inflammation, and some mice received POH treatment. POH significantly suppressed indicators of allergic airway inflammation such as airway eosinophilia. Cytokine production in thoracic lymph nodes was also significantly suppressed. These results demonstrate that POH suppresses antigen-induced immune responses in the lung. Considering that it exists naturally, POH could be a novel preventive or therapeutic option for immunologic lung disorders such as asthma with minimal side effects.

Influence of GSTM1 and GSTT1 polymorphisms on the survival rate of patients with malignant glioma under perillyl alcohol-based therapy

GSTM1 (glutathione S-transferase mu 1) and GSTT1 (glutathione S-transferase theta 1) are critical enzymes for detoxification of endogenous and environmental carcinogens. Constitutive GST gene polymorphisms may be associated with increased risk for cancer development. We made an explorative study of a Brazilian population with malignant glioma to determine whether GSTM1 and GSTT1 genetic polymorphisms influence the response to intranasal administration of perillyl alcohol and the survival rate. Patients were stratified into groups according to clinical presentation, tumor classification, and tumor location. Circulating DNA was extracted from blood plasma or serum, and genotypes were detected by multiplex PCR. The cohort included 95 patients with recurrent malignant glioma included in a Phase I/II clinical trial with perillyl alcohol and 100 matched healthy control subjects. GSTM1 frequency was similar in patients with glioma (44%) and healthy controls (54%), but GSTT1 deletion was found in 11.5% patients, contrasting with 36% in controls. A longer survival rate was associated with a lack of GSTM1 deletion (31 weeks) and a deletion for GSTT1 (28 weeks). A poor survival rate was associated with GSTM1 deletion (23 weeks) and with a lack of a GSTT1 deletion (19 weeks). A significantly lower frequency of GSTT1 deletion in glioma patients compared to healthy controls indicates that GSTT1 deletion may exert a protective role against gliomagenesis, influence therapeutic response to intranasal perillyl alcohol treatment, and increase overall survival, especially considering tumor topography.

Prevention of transplant coronary artery disease by prenylation inhibitors

In this study we systematically dissect the prenylation pathway to better define the mechanism behind statin inhibition in chronic allograft rejection in heart transplants, or transplant coronary artery disease (TCAD). Utilizing a murine heterotopic heart transplant model, animals received daily treatments of either statin or selective isoprenoid blockade inhibitors to block the four major downstream branches of the mevalonate pathway. TCAD was assessed by morphometric analysis at Day 52. Graft-infiltrating cells, cytokine production, smooth muscle cell proliferation and migration and endothelial cell MHC II expression were detected on Day 7. Atorvastatin and two prenylation inhibitors, NE-10790 and manumycin A, significantly reduced TCAD lesions compared with untreated animals. Perillyl alcohol treatment resulted in a trend toward decreased luminal narrowing. Finally, zaragozic acid (cholesterol blockade only) did not alter TCAD severity. Statins and prenylation inhibitors reduced inflammatory cell allograft recruitment, but did not always correlate with TCAD reduction. Cytokine production was decreased in recipient spleens in all treatment groups. Both in vitro and in vivo IFN-γ-stimulated MHC II expression was decreased in a dose-dependent manner in the atorvastatin, perillyl alcohol and NE-10790 groups. In vitro smooth muscle cell proliferation was decreased in all treatment groups. Finally, in vitro smooth muscle cell migration was decreased in the atorvastatin, NE-10790 and manumycin A groups only. FPT and GGPT-2 (inhibition) are the key enzymes in the HGM-CoA reductase pathway and most influential in TCAD prevention. TCAD reduction is most closely related to smooth muscle cell migration, but not its anti-inflammatory properties.

Efficacy of monoterpene perillyl alcohol upon survival rate of patients with recurrent glioblastoma

The monoterpene perillyl alcohol (POH) a Ras inhibitor with potential capacity to arrest gliomagenesis is being used in a phase I/II clinical trial in adults with recurrent malignant glioma. The present study aimed to investigate the efficacy of intranasal administration of monoterpene POH upon survival rate of patients with recurrent glioblastoma (GBM) in comparison with historical control group of GBM patients. It was included 89 adults with recurrent GBM receiving daily intranasal administration of 440mg POH and 52 matched GBM patients as historical control untreated group only with supportive treatment. Patients with recurrent primary GBM treated with POH survived significantly longer (log rank test, P<0.0001) than untreated group. Patients with recurrent primary GBM in deep location survived significantly longer than with lobar location (log rank test, P<0.0001). Median survival rate of secondary GBM was 11.2months, longer (log rank test, P=0.0366) than primary GBM (5.9months). Radiographic improvement and reduction of corticosteroid dosage (36%) further associated with a delay towards progression. Intranasal administration of POH increased the overall survival of patients with recurrent GBM in comparison with historical untreated controls, but especially patients with secondary GBM and primary GBM with tumor localized in deep regions of the brain. The side effects of POH treatment were almost nonexistent, even in patients treated for over 4years.

Perillyl alcohol and methyl jasmonate sensitize cancer cells to cisplatin

Breast cancer is the second leading cause of cancer deaths among women in the United States. Several treatment options exist, with different side effects. To alleviate the side effects, several research groups have studied chemotherapeutic effects of plant compounds on cancer cells. These could be used as an alternative treatment option either alone or in combination with other chemotherapeutic drugs. The aim of this study was to evaluate the activity of a combination of perillyl alcohol (POH), methyl jasmonate (MJ) with cisplatin to define the most effective schedule and to investigate the mechanism of action in breast cancer cells. POH and MJ treatment (20% decrease in cell viability concentration) enhanced the cytotoxicity for subsequent exposure to cisplatin in MDA-MB-435 and MDA-MB-231 cells. Combination treatment of POH and MJ blocked cells at the G0/G1 phase of the cell cycle and the addition of cisplatin forced the cells to progress through the cell cycle and induced apoptosis. Apoptotic mechanistic studies indicated that POH and MJ treatment activated tumor necrosis factor receptor 1 and this was further increased by the addition of cisplatin. It was also found that mitochondrial membrane potential decreased with POH and MJ treatment; this effect was further enhanced by cisplatin treatment. These findings contributed to a better understanding of molecular mechanism of apoptosis in combination treatment of POH, MJ, and cisplatin. Results also showed that the combination treatment of three drugs is more effective than single drug alone or two drugs together.

A Pilot Study of Perillyl Alcohol in Pancreatic Cancer

Chemotherapy has been largely unsuccessful in pancreatic cancer. Measurement of cell-specific biological endpoints may clarify the evaluation of a newer generation of compounds. Perillyl alcohol has shown chemotherapeutic activity in preclinical systems through enhancing apoptosis. To pilot a new trial template for testing novel agents in pancreatic cancer and to assess the biological activity of perillyl alcohol in patients with resectable pancreatic cancer. Apoptosis was quantified with ApopTag in situ, Bak staining, and light microscopy. Tumor size, serum CA 19-9 level, and survival were also measured. Eight patients enrolled. Toxicity was mild and perillyl alcohol was generally well tolerated. Tumor size and CA 19-9 level were unchanged with perillyl alcohol treatment. Survival time was longer in patients who received full perillyl alcohol treatment (288 +/- 32 days) compared to those who did not (204 +/- 96 days), but this result did not achieve statistical significance (P = 0.2). There was a trend toward greater apoptosis in patients receiving perillyl alcohol compared to fresh operative controls; there was also a suggestion of greater apoptosis in tumor compared to normal pancreatic tissue in the same patient. Incorporation of cell-specific biological endpoints is challenging but feasible and should be used in clinical studies of pancreatic cancer treatment. Our pilot study suggests that perillyl alcohol may indeed have effects on biological endpoints. This study will serve as a useful template for examining cell-specific biological endpoints in the testing of future agents that are thought to induce apoptosis in pancreatic cancer.

QS104. A Pilot Study of Perillyl Alcohol in Pancreatic Cancer

Chemotherapy has been unsuccessful in pancreatic cancer. Measurement of cell specific biological endpoints may clarify the evaluation of a newer generation of compounds. Perillyl alcohol (POH) has shown chemotherapeutic activity in pre-clinical systems through enhancing apoptosis. Objective: To assess the biological activity of POH in patients with resectable pancreatic cancer. Methods: Apoptosis was quantified with ApopTag in situ, Bak staining, and light microscopy. Tumor size, serum CA 19-9 level, and survival were also measured. Results: Eight patients enrolled. Toxicity was mild and POH was generally well tolerated. Tumor size and CA 19-9 level were unchanged with POH treatment. Survival time was longer in patients who received 100% of POH treatment (288 + 32 days) compared to those who did not (204 + 96 days), but this result did not achieve statistical significance (p= 0.2). There was a trend toward greater apoptosis in patients receiving POH compared to fresh operative controls; there was also a suggestion of greater apoptosis in tumor compared to normal pancreatic tissue in the same patient. Conclusions: Pancreatic cancer continues to be a deadly cancer. Incorporation of cell specific biological endpoints is challenging but feasible and should be employed in clinical studies of pancreatic cancer treatment. Our pilot study suggests that POH may indeed have effects on biologic endpoints. The study is underpowered, but will serve as a useful template for examining cell specific biologic endpoints in the testing of future agents that are thought to induce apoptosis in pancreatic cancer.