Ayurveda Inspired Discovery

The word "Ayurveda" comprises of word "āyus" meaning life or "life principle", and the "veda", which refers to a system of "knowledge". Thus "Ayurveda" roughly translates as the "knowledge of life".

Ayurveda is a more than 5000 years-old traditional system of Indian medicine and continues to serve 70% of India's rural population. Recent research in Ayurveda has been dominated by studies on medicinal plants and the development of herbal drugs, which has a large market growing at 15% per year. However fundamental life science or biotechnology based studies to understand concepts, procedures, and products are still inadequate. RGCB has major research interests in elucidating the underlying biological and molecular mechanisms involved in ayurvedic therapeutics as well scientifically documenting the various plants used in composition of ayurvedic medicines through a Natural Products Research Platform that uses the tools of DNA fingerprinting, DNA barcoding, chemical fingerprinting, medicinal chemistry, cell biology, chemoinformatics and metabolomics.

Read the decadal vision document "Towards Ayurvedic Biology" by Professor M.S Valiathan and published by the Indian Academy of Sciences. http://www.ias.ac.in/academy/dvdocs/ayurvis.pdf

Select Examples of Ayurveda Research at RGCB

Efficacy of Varanadi Ghritha in preventing loco-regional recurrences and the occurrence of second malignancies in treated Head & Neck Cancers

[Clinical collaboration with Regional Cancer Centre, Thiruvananthapuram]

Cancer chemoprevention is defined as the use of natural, synthetic or biologic chemical agents to reverse, suppress, or prevent carcinogenic progression to invasive cancer. Tertiary prevention focuses on the prevention of second primary tumors (SPTs) in patients cured of their initial cancer or individuals definitively treated for their premalignant lesions. Chemoprevention trials are based on the hypothesis that interruption of the biological process involved in carcinogenesis will inhibit this process and in turn, reduce cancer incidence. Our pilot studies have shown immunomodulation and improvement in quality of life following the administration of Varanadi ghritha and no toxicity was observed. However its impact in preventing loco-regional recurrence and second primary tumor was not clear. Therefore this research investigation will study whether the administration of Varanadi ghritha, a poly-herbal formulation used in Ayurveda will be able to control loco-regional failure and development of second malignancy in treated Head and Neck cancer patients as well as to see whether this will improve the immunity and quality of life of treated patients. Uncontrolled loco-regional disease and development of second primary are the major causes of failure in Head and Neck cancer patients. Several ingredients in Varanadi ghritha have been found to have anti-tumor, apoptotic, immunomodulatory and antioxidant properties.

Divya Ravindran, Indhu Hariharan, Rejnish Kumar, RR Muwonge, M. Radhakrishna Pillai, Kunnambath Ramadas. Varanadi Ghritha as an Immunomodulator in Head & Neck Cancer patients - A Randomized trial.
In: Abstract book 3rd International Symposium on Translational Cancer Research 2009, organized by Institute of Life Sciences, Bhubaneswar and UT MD Anderson Cancer Centre, Houston. December 2009. Bhubaneswar, Orissa, India.

Divya Ravindran, Indhu Hariharan, Rejnish Kumar R, Richard Muwonge, M. Radhakrishna Pillai, Kunnambath Ramadas. Efficacy of Varanadi Ghritha as a Biological Response Modifier in Controlling the Recurrence of Treated Head and Neck Cancers- A Randomized Pilot Study.
In: Proceedings of 97th Indian Science Congress, January 2010, p 170.

Divya Ravindran, Indhu Hariharan, R. Rejnish Kumar, Richard Muwonge, M. Radhakrishna Pillai, K Ramadas. Immunomodulation by Varanadi Ghritha and its role in preventing loco-regional recurrence/ second malignancy in Head & Neck Cancer patients - a three-year follow up data.
In: Proceedings of International Cancer Research Symposium 2010: Defining & Translating Science behind the disease, December 2010,Thiruvananthapuram, India Page No 0:5.

Clinical value of a mouthwash composed of essential ingredients in Ayurveda for management of radiation-Induced oral mucositis

[Clinical collaboration with Regional Cancer Centre, Thiruvananthapuram]

Oral mucositis is thought to be a complex biological process, involving direct damage to the dividing cell of the oral epithelium with depletion of the basal epithelium modulated by the immune system, the inflammatory process and by super added infection by the oral bacterial flora, especially the aerobic gram negative bacteria. Oral mucositis besides being extremely painful prevents adequate food and water intake and also is the major dose limiting toxicity in patients undergoing radical radiation therapy for oral cancers. The purpose and objectives of the study was to evaluate the effectiveness of an ayurvedic herbal mouthwash containing five herbal ingredients in reducing severity of oral mucositis. We carried out a clinical trial on 148 patients with biopsy proven squamous cell carcinoma of oral cavity and scheduled to be treated with radiation. Patients were randomized using a computer generated randomization chart, 75 to the intervention arm and 73 to the control arm. Patients in intervention arm were advised to rinse the mouth with herbal mouthwash 4 times daily and those in control arm were advised soda saline mouthwash during the entire period of radiation therapy. All patients received external beam radiation therapy 5 days a week up to a total dose of 5250 c Gy in 15 fractions over 3 weeks period. All patients were evaluated weekly by a single physician, who was blinded to randomization procedure (blinded assessor). RTOG grading of mucositis was used to assess the grade of mucositis. Pain intensity was scored on a 10-point scale using a Visual Analogous Scale (VAS). Analgesics use among these patients was assessed on the basis of WHO step ladder analgesic requirement. The response to treatment was assessed at 5-6 months after completion of radiation therapy. Base line characteristics of patients in both arms were comparable. The intensity of pain at the end of radiation therapy was less in intervention arm. Severe pain (7-10 score on VAS) was experienced by 5.4% patients in intervention arm compared to 45.7% in control arm (p < 0.001). Onset of mucositis was significantly delayed in patients in intervention arm. The analgesic requirement was low in intervention arm. Step 2 and Step 3 analgesic usage was 8.1% and 1.4% respectively in intervention arm compared to 21.4 % 2.9% in control arm (p < 0.001). Treatment interruptions and hospitalizations were also significantly low in intervention arm. Good oral hygiene was observed in patients in intervention arm at the end of radiation therapy; 37.8% vs 10.0% in control arm (p < 0.001). Side effects of radiation therapy such as intolerance to spicy food and dryness of mouth were significantly less in patients in intervention arm. Complete response to radiation therapy was 92.6% in intervention arm compared to 79% in control arm. The compliance to treatment was better in intervention arm and no toxicity was observed. The ayurvedic herbal mouthwash found to be very effective in minimizing radiation induced oral mucositis and improving treatment outcome. Although not tested this formulation may be tried in patients undergoing radiotherapy for uterine cervical cancer as retention enema or as a gel in preventing proctitis.

Ramadas K, Ravindran D, Kumar RR, Muwonge R, Pillai MR. Management of radiation-induced oral mucositis with an herbal mouthwash. International Journal of Radiation Oncology 87 (25) Supplement: S143 - S144, 2013.

Amalaki rasayana in cardiac failure

Heart failure is a leading cause of hospitalization and mortality worldwide. Despite considerable therapeutic advances, the morbidity and mortality associated with heart failure continues to be unacceptably high. There has been considerable progress in the treatment of chronic heart failure with angiotensin converting-enzyme inhibitors, aldosterone antagonists, beta-receptor blockers, and resynchronization therapy. Even with the very best of these modern therapies, heart failure is still associated with an annual mortality rate of 10%. Developing new therapies with improved efficacy and minimum side effects for prevention and management of heart failure remains a major challenge in cardiology. Our study aims to evaluate Amalaki Rasayana in the prevention and reversal of remodeling changes in cardiac failure in a pressure overload model of left ventricular cardiac hypertrophy and failure in rats and age associated cardiac dysfunction.

Antiviral molecules from natural products and Ayurvedic leads

RGCB has a strong interest in identifying antiviral leads against chikungunya virus and dengue virus from natural resources. Both whole-virus based phenotypic assays as well as targeted assays focusing on specific viral protein-inhibition are being developed for this purpose. We have developed a prototype NS2B protease assay for evaluating potent anti-dengue molecules, which is being refined for increased sensitivity. Three natural compounds, with a predicted strong capacity to bind to E1 envelope protein of Chikungunya virus, were identified earlier in our laboratory. In vitro evaluation of these compounds shows encouraging results, with one of the molecules exhibiting complete viral inhibition in very low doses. Cytopathogenicity inhibition, plaque reduction assays and viral binding assays using various susceptible cell lines are currently being employed for identification of anti-chikungunya virus molecules in natural products and plant extracts. We initially short-listed 22 plants based on ethno-botanical information on medicinal plants, and from this four plants were screened for anti-CHIKV activity. Interestingly, a non-polar solvent extracted fraction from one of these plants showed promising activity. All these promising candidates are being evaluated further. Initiatives are also being taken to carry out in vivo analysis of these molecules in mice models.

Pomegranate extract demonstrate a selective estrogen receptor modulator profile in human tumor cell lines and in vivo models of estrogen deprivation

Pomegranate (Punica granatum, Punicaceae) is a native to Mediterranean region and has been used extensively in the folk medicine of many cultures Pomegranate fruit is a rich source of polyphenolic compounds like anthocyanidins (delphinidin, cyanidine and pelargonidin) and hydrolysable tannins (such as punicalgin, pedunculgin, punicalin, gallagic, ellagic acid esters of glucose), which account for 92% of antioxidant activity of whole fruit Various parts of the pomegranate fruit have been shown to exert anti-proliferative, anti-angiogenic, anti-aromatase and pro-apoptotic effects on human breast cancer cell lines and chemopreventive properties in mouse mammary organ culture. Previous studies have proved the high antioxidant activities of the methanol extract of pomegranate peel in various in vitro and in vivo models. Estrogen is implicated in the development of breast cancer, based on the data from both clinical and animal studies and an ideal strategy for prevention and treatment of estrogen-dependent breast cancer is to selectively block estrogen activity in the affected tissues without compromising its beneficial effects. Such selective estrogen receptor modulators (SERMs) bind ER and exert estrogen agonist action in some target tissues while acting as estrogen antagonists in others. SERMs may be possibly considered therapeutically for the inhibition of proliferation of breast ductal epithelium with maintenance of bone density and reduction in cholesterol levels without uterine endometrial proliferation. We analyzed the methanol extract of pomegranate pericarp (PME) for its possible SERM like property using human breast, endometrial, cervical and ovarian cancer cell lines as well as in vivo models (ovariectomized Swiss albino mice) using biochemical markers of SERM activity. Our findings demonstrate that PME binds to ER and down-regulates the ERE-mediated transcription in breast cancer cells without being agonistic in the uterine endometrium and has cardio-protective effects comparable to that of 17β-estradiol.

Sreeja S, Hima Sithul, Parvathy Muraleedharan, Juberiya Mohammed Azeez, and Sreeja S, "Pomegranate Fruit as a Rich Source of Biologically Active Compounds", BioMed Research International Volume (2014), http://dx.doi.org/10.1155/2014/686921

S Sreeja, T R Santhosh Kumar, S Lakshmi, S Sreeja. Pomegranate extract demonstrate a selective estrogen receptor modulator profile in human tumor cell lines and in vivo models of estrogen deprivation. Journal of Nutritional Biochemistry, DOI :10.1016/j.nutbio.2011.03.015.

Plumbagin as a possible therapeutic agent for hereditary breast cancers

The hereditary breast and ovarian cancers harbor germline mutations in BRCA1 while in sporadic breast cancers, the BRCA1-defect could arise from a functional deficiency of BRCA1 due to hyper-methylation of its promoter. Standard chemotherapeutic regimens that were initially found to be effective in BRCA1-defective breast cancers have succumbed to the problem of insensitivity and relapse. We have demonstrated for the first time, that a naphthaquinone derivative, Plumbagin (5-hydroxy-2-methyl-1,4-naphthoquinone) isolated from Plumbago zeylanica has shown promising specific anticancer activity to target BRCA1-defective ovarian cancers and BRCA1-defective breast cancers in vitro and in vivo. Plumbagin may interfere with ER-a signaling pathway and induce apoptosis in a death receptor-mediated pathway in ER-a overexpressing BRCA1 defective ovarian cancer cells. Plumbagin can also generate Reactive Oxygen Species (ROS) that induces damage to the nucleic acids that will not be repaired by BRCA1 defective cells. Using genetically modified mouse models we have proved that plumbagin has potential to be a more effective anti-cancer agent than carboplatin against such cancers.

Sinha, S., Pal, K., Elkhanany, A., Dutta, S., Cao, Y., Mondal, G., Iyer, S., Somasundaram, V., Couch, F. J., Shridhar, V., Bhattacharya, R., Mukhopadhyay, D. and Srinivas, P. Plumbagin inhibits tumorigenesis and angiogenesis of ovarian cancer cells in vivo. Int. J. Cancer: 132, 1201-1212 (2013).

Thasni KA, Ratheeshkumar T, Rojini G, Sivakumar KC, Rakesh Sathish Nair, Srinivas G, Asoke Banerji, Veena Somasundaram, Priya Srinivas. Structure activity relationship of Plumbagin in BRCA1 related Cancer Cells. Mol Carcinog. 52, 392-403 (2013).

Priya Srinivas, Chitta Ranjan Patra, Santanu Bhattacharya and Debabrata Mukhopadhyay. Cytotoxicity and ROS generating capacity of naphthaquinones is quenched when conjugated with gold nanoparticles. Int J Nanomed: 6 2113-2122, 2011.

Thasni KA, Rakesh S, Rojini G, Ratheeshkumar T, Srinivas G, Priya Srinivas. Estrogen-dependent cell signaling and apoptosis in BRCA1-blocked BG1 ovarian cancer cells in response to plumbagin and other chemotherapeutic agents. Ann Oncol 19: 4; 696-705, 2008.

Molecular biology and biological standardization interphase in ayurveda based pharmaceutical and herbal product industry

The quality of primary raw materials, either whole or part of medicinal plants, constitutes the backbone of ayurvedic and herbal medicine industry. Three factors negatively affect the quality of herbal materials used in the pharmaceutical industry: scanty morphological descriptors to identify authentic materials; use of the same common name for different species; and intentional mixing with inexpensive herbs. It has become imperative to implement a code of practices for biological standardization of raw materials used in each Ayurveda medicine in order to ensure batch-to-batch consistency in the efficacy of herbal drugs. The nucleotide polymorphisms, which sort each biological material at the genome level, can be used for the accurate circumscription of different varieties of medicinal species and for the precise delimitation of authentic medicinal species by employing DNA fingerprinting technology for the former and DNA barcoding for the latter. Utilizing DNA markers, we successfully circumscribed the medicinal rice Njavara, the single largest used rice in Ayurveda and developed a DNA barcode repository for the accurate identification of Zingiberaceae species, whose rhizomes are indispensable in the preparation of several Ayurveda drugs. Development of a DNA marker/barcode repository for all commonly used herbal species supported with herbaria would validate Ayurveda traditions and expand its credibility.

Sreejayan, Suresh Kumar, U., Varghese, G., Jacob, T. M. and Thomas, G. 2011. Stratification and population structure of the genetic resources of ancient medicinal rice (Oryza sativa L.) landrace Njavara. Genetic Resources and Crop Evolution 58:697-711

Varghese, G., Jose, M., Dinesh Raj, R., Bocianowski, J., Thomas, G. and Omanakumar, N. 2014. Quantitative and molecular analyses reveal a deep genetic divergence between the ancient medicinal rice (Oryza sativa) Njavara and syntopic traditional cultivars. Annals of Applied Biology 164: 95-106.

Vinitha, M. R., Suresh Kumar, U., Aishwarya, K., Sabu, M. and Thomas, G. 2014. Prospects for discriminating Zingiberaceae species in India using DNA barcodes. Journal of Integrative Plant Biology 56: 760-773.

Structural and Biological Characterization of anti-hepatocellular carcinoma compound from Glycosmis pentaphylla

Hepatocellular carcinoma (HCC) or Liver cancer has a poor survival rate and is the fifth most common cancer in the world and the third leading cause of cancer related death. A complete cure for this disease is not available today. Unlike synthetic drugs, herbal medicines are mixture of structurally diverse compounds, which may or may not provide therapeutic activity. A complete characterization of all the chemical constituents from a natural product and its mechanism of biological activity are essential for its standardization. The search for new lead compounds from plant sources is a crucial element of modern pharmaceutical research. Glycosmis pentaphylla (Retz) correa is commonly known as ashvashakota, vananimbuka (Sanskrit) and paanal (Malayalam). It belongs to Rutaceae family. In Ayurveda G. pentaphyllais considered as one an important option for curing fever, cough, rheumatism, anaemia, cancer and liver disorders. However a scientific validation regarding the active compounds, mechanism of action, safety and efficacy is still not yet completely elucidated. The present study aimed for the isolation and characterization of anti-HCC compounds from G. pentaphylla and molecular mechanism behind its activity using HCC cell lines (Hep3 B).The cytotoxic and apoptosis inducing effect of the active extract and its fractions were valued on Hep3 B, HepG2, HEK293, LX2 and RAW264.7 cell lines by MTT assay. Molecular mechanism behind the activity of extract was proved by using morphological studies, Hoechst staining, DNA fragmentation, reverse transcription polymerase chain reaction and western blotting. The phytochemical profiling of the active extract was accomplished by different biochemical assays. Compound purification and identification were done by using chromatography and different spectroscopic methods (NMR, IR, DTA, UV/VIS, CHN element analysis and MS). The preliminary investigation showed a selective anti-HCC activity of G. pentaphylla alcohol extract on HCC cell lines (Hep3B and HepG2) without significant cytotoxicity on non-hepatocyte cell lines, HEK 293 and RAW 264.7. Morphological observation, Hoechst staining, DNA fragmentation, transcript and translational studies showed the cytotoxicity of active extract is by regulating the apoptosis cascade. The mechanism of action of active alcohol extract showed that it induces apoptosis by regulating expression of Bax/Bcl2 gene expression time and dose dependently. Phytochemical screening of the active alcohol extract confirmed the presence of alkaloids, flavonoid and sterol. Chromatography techniques were used to isolate and purify the most active compound from the active alcohol extract. Compound characterization did by chromatography, phytochemical tests and spectroscopic techniques. The results showed that the alcohol extract of G. pentaphylla contain a modified precursor of flavonoid which is responsible for its selective anti HCC activity.

PS Sreejith, RR Mascarenhas, RJ Praseeja, VV Asha. The apoptosis inducing effect of Glycosmis pentaphylla (Retz.) Correa and its influence on gene expression in hepatocellular carcinoma cell line, Hep3 B. Journal of Ethnopharmacology. 2012, 139: 359-365.

K. A. Krishnakumar, K.H. Muneeb Hamza and V.V. Asha.. Antifibrotic activity of Phyllanthus maderaspatensis L. in Wistar rats. Annals of Phytomedicine 2014, 3: 68-79.

Suresh V. Asha V.V. Anti-HBV activity of the different extracts from Phyllanthus rheedi Wight in cell culture assay based system.Journal of Ethnopharmacology 2014 (in press)

Exploring the metabolite diversity in pharmacologically significant plants leading to natural product synthesis

Plants are rich natural source of medicinally important metabolic compounds and are vital to Ayurvedic healthcare. Type III Polyketide synthases (PKSs) represent the largest group of enzymes, which have potential role in the biosynthesis of biologically active secondary metabolites with important roles in both plant and human health. Our research aims for the identification and characterization of Type III PKSs that are involved in the biosynthesis of alkaloids, flavonoids and various other pharmacologically active compounds by molecular approaches from various medicinal plants including Aegle marmelos, (Bael), Emblica officinalis (Indian Gooseberry), Curcuma longa (Turmeric) and Zingiber officinale (Ginger). We also focus on chemical, structural and biochemical analyses, which will provide an atomic resolution of plant PKSs and will serve as a catalyst for bioengineering and scalable production of various nutraceuticals. In addition to that, the elucidation of molecular networks and metabolic pathways through transcriptome approach will help to understand the biosynthesis of secondary metabolites especially piperine alkaloids from black pepper. Further our research sheds light into the metabolic redesign of lead molecules for the development of novel plant natural product.

Resmi MS, Verma P, Gokhale RS, Soniya EV (2013). Identification and characterization of a type III polyketide synthase involved in quinolone alkaloid biosynthesis from Aegle marmelos Correa. J Biol Chem. 288(10):7271-81.

Resmi, M.S., Soniya, E.V (2011). Molecular cloning and differential expressions of two cDNA encoding Type III polyketide synthase in different tissues of Curcuma longa L, Gene: 491(2012)278-283.

Radhakrishnan, E.K. and Soniya, E.V (2009). Molecular Characterization of Novel form of Type III Polyketide Synthase from Zingiber Officinale Rosc. and its Analysis using Bioinformatics Method. J Proteomics Bioinform,2(7): 310-315 - 310.

Evaluation of Curcumin as a chemosensitizer and chemopreventive

Several studies have shown that curcumin exhibits remarkable anticancer activity in vitro. However its efficacy as an anticancer agent is yet to be proved in clinics, due to its poor bioavailability and retention time within the body. The maximum available concentration of curcumin in the physiological condition is between 2-5 µM, at which curcumin does not produce any anticancer effect. Our studies illustrate that though curcumin cannot act as an anticancer drug at this concentration, it can act as a potent chemosensitizer with paclitaxel against cervical cancer. We also observed that curcumin can inhibit all survival signals induced by nicotine in lung cancer cells and can inhibit Benzo[a]pyrene-induced lung carcinogenesis, illustrating its role as an effective chemopreventive against environmental carcinogenesis. We have also established in vitro that nano-encapsulation can make curcumin aqueous soluble and cell permeable

Therapeutic potential of tryptanthrin, isolated from Wrightia tinctoria agianst malignant melanoma

We isolated tryptanthrin from the indigenous medicinal plant, Wrightia tinctoria, belonging to Apocyanaceae. It was found to be most cytotoxic to melanoma cells, while being non-toxic to normal skin fibroblasts and melanocytes. It induced apoptosis in melanoma cells and down-regulated various survival signals that are over-expressed in melanoma. It also down-regulate the key molecules involved in angiogenesis and metastasis. The in vivo experiments also confirm the pharmacological safety of the compound and provide an idea about the selection of starting doses for phase 1 human studies. In vivo xenograft studies using melanoma cells confirmed the therapeutic and antiangiogenic potential of the compound. Tryptanthrin also inhibited metastasis of melanoma cells as assessed by different metastasis models. Taken together, this is the first study reporting the anticancer property of Wrightia tinctoria and is the first detailed report of extraction and purification of tryptanthrin from this plant. It is also the first study reporting the efficacy of tryptanthrin towards malignant melanoma and its antimetastatic potential. This is also the first study conducted for the toxicological evaluation of the compound.

BS Vinod, J Antony, HH Nair, VT Puliyappadamba, M Saikia, S Shyam Narayanan, A Bevin and Ruby John Anto. Mechanistic evaluation of the signaling events regulating curcumin-mediated chemosensitization of breast cancer cells to 5-fluorouracil. Cell Death and Disease 4, e 505; doi:10.1038/cddis.2013.26, 2013.

Lekha Nair K, Arun Kumar T Thulasidasan, Deepa G, Ruby John Anto, GS Vinod Kumar. Purely aqueous PLGA nanoparticulate formulations of curcumin exhibit enhanced anticancer activity with dependence on the combination of the carrier. Int J Pharm.4:425 (1-2), 44-52, 2012.

Chanickal N Sreekanth, Smitha V Bava, E Sreekumar and Ruby John Anto. Molecular evidences for the chemosensitizing efficacy of liposomal curcumin in paclitaxel chemotherapy in mouse models of cervical cancer. Oncogene. 30, 3139-3152, 2011.

Smitha V Bava, Chanickal N Sreekanth, Arun Kumar T Thulasidasan, Nikhil P Anto, Vino T Cheriyan, Vineshkumar T Puliyappadamba, Sajna G Menon, Santhosh D Ravichandran, and Ruby John Anto. Akt is upstream and MAPKs are downstream of NF-?B in paclitaxel-induced survival signaling events, which are down-regulated by curcumin contributing to their synergism. Int J Biochem Cell Biol. 43, 331-341, 2011.

Vineshkumar T Puliyappadamba, Vino T Cheriyan, Arun Kumar T Thulasidasan, Smitha V Bava, Balachandran S Vinod, Priya R Prabhu, Ranji Varghese, Arathy Bevin, Shalini Venugopal and Ruby John Anto. Nicotine-induced survival signaling in lung cancer cells is dependent on their p53 status while its down-regulation by curcumin is independent. Mol Cancer, 9, 220, 2010. (Highly Accessed).

Smitha V. Bava, Vineshkumar T. Puliappadamba, Ayswaria Deepti, Asha Nair, Devarajan Karunagaran, and Ruby John Anto (2005) Sensitization of taxol-induced apoptosis by curcumin involves down regulation of NF-kB and Akt and is independent of tubulin polymerization: J. Biol. Chem., 280: 6301 - 6308.