Cytotoxic Effect of the Pentacyclic Oxindole Alkaloid Mitraphylline Isolated from Uncaria tomentosa Bark on Human Ewing's Sarcoma and Breast Cancer Cell Lines

Abstract

Preparations from Uncaria tomentosa, a South American Rubiaceae, have been used in the Peruvian traditional medicine for the treatment of infective, inflammatory and tumoral processes. In this study, the pentacyclic oxindole alkaloid mitraphylline was isolated from the dried inner bark of this plant species, and its structure elucidated by analysis of NMR spectroscopic data. Mitraphylline was differentially identified from its stereoisomeric pair isomitraphylline by ¹⁵N‐NMR. Its antiproliferative and cytotoxic effects have been tested on human Ewing's sarcoma MHH‐ES‐1 and breast cancer MT-3 cell lines, using cyclophosphamide and vincristine as reference controls. A Coulter counter was used to determine viable cell numbers, followed by the application of the tetrazolium compound MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxy phenyl)-2-(4-sulfophenyl)-2H-tetrazolium] an inner salt. A colorimetric method was employed to evaluate cell viability in this cytotoxic assay. Micromolar concentrations of mitraphylline (5 µM to 40 µM) inhibited the growth of both cell lines in a dose-dependent manner. The IC₅₀ ± SE values were 17.15 ± 0.82 µM for MHH‐ES‐1 and 11.80 ± 1.03 µM for MT-3 for 30 hours, smaller than those obtained for the reference compounds. This action suggests that the pentacyclic oxindole alkaloid mitraphylline might be a new promising agent in the treatment of both human sarcoma and breast cancer.

Key words

Uncaria tomentosa - Rubiaceae - cat's claw - oxindole alkaloid - mitraphylline - cytotoxic activity

References

• 1 Keplinger K, Laus G, Wurm M, Dierich M P, Teppner H. Uncaria tomentosa (Willd.) DC. ethnomedicinal use and new pharmacological, toxicological and botanical results.  J Ethnopharmacol. 1999;  64 23-34
• 2 Heitzman M E, Neto C C, Winiarz E, Vaisberg A J, Hammond G B. Ethnobotany, phytochemistry and pharmacology of Uncaria (Rubiaceae).  Phytochemistry. 2005;  66 5-29
• 3 Montoro P, Carbone V, Quiroz De Dioz J, De Simone F, Pizza C. Identification and quantification of components in extracts of Uncaria tomentosa by HPLC-ES/MS.  Phytochem Anal. 2004;  15 55-64
• 4 Kitajima M, Hashimoto K, Yokoya M, Takayama H, Sandoval M, Aimi N. Two new nor-triterpene glycosides from peruvian “Uña de Gato” (Uncaria tomentosa).  J Nat Prod. 2003;  66 320-323
• 5 Kitajima M, Yokoya M, Takayama H, Aimi N. Synthesis and absolute configuration of a new 3, 4-dihydro-beta-carboline-type alkaloid, 3, 4-dehydro-5(S)-5-carboxy-strictosidine, isolated from Peruvian Uña de Gato (Uncaria tomentosa).  Chem Pharm Bull. 2002;  50 1376-1378
• 6 Muhammad I, Dunbar D C, Khan R A, Ganzera M, Khan I A. Investigation of Uña De Gato I. 7-Deoxyloganic acid and ¹⁵N NMR spectroscopic studies on pentacyclic oxindole alkaloids from Uncaria tomentosa.  Phytochemistry. 2001;  57 781-785
• 7 Muhammad I, Khan I A, Fischer N H, Fronczek F R. Two stereoisomeric pentacyclic oxindole alkaloids from Uncaria tomentosa: uncarine C and uncarine E.  Acta Crystallogr C. 2002;  57 480-482
• 8 Aquino R, De Simone F, Vincieri F F, Pizza C. New polyhydroxylated triterpenes from Uncaria tomentosa.  J Nat Prod. 1990;  53 559-564
• 9 De Matta S M, Monache F D, Ferrari F, Marini-Bettolo G B. Alkaloids and procianidins of Uncaria sp. from Peru.  Pharmacology. 1976;  31 527-535
• 10 Rizzi R, Re F, Bianchi A, De Feo V, De Simone F, Bianchi L, Stivala L A. Mutagenic and antimutagenic activities of Uncaria tomentosa and its extracts.  J Ethnopharmacol. 1993;  38 63-77
• 11 Phillipson J D, Hemingway S R. Chromatographic and spectroscopic methods for the identification of alkaloids from herbarium samples of the genus Uncaria.  J Chromatogr. 1975;  105 163-178
• 12 Laus G, Keplinger D. Separation of stereoisomeric oxindole alkaloids from Uncaria tomentosa by high performance liquid chromatography.  J Chromatogr A. 1994;  662 243-249
• 13 Ganzera M, Muhammad I, Khan R A, Khan I A. . Improved method for the determination of oxindole alkaloids in Uncaria tomentosa by high performance liquid chromatography.  Planta Med. 2001;  67 447-450
• 14 Philp R B. Herbal-drug interaction and adverse effects. An evidence-based quick reference guide. New York; McGraw-Hill Editions 2004: 63-65
• 15 Reinhard K H. Uncaria tomentosa (Will.) DC: cat's claw, uña de gato, or saventaro.  J Altern Compl Med. 1999;  5 143-151
• 16 Sandoval M, Charbonnet R M, Okuhama N N, Roberts J, Krenova Z, Trentacosti A M, Miller M J S. Cat's claw inhibits TNF [alpha] production and scavenges free radicals: role in cytoprotection.  Free Radic Biol Med. 2000;  29 71-78
• 17 Sandoval-Chacon M, Thompson J H, Zhang X J, Liu X, Mannick E E, Sadowska-Krowicka H, Charbonnet R M, Clark D A, Miller M. Antiinflammatory actions of cat's claw: the role of NF-kB.  Aliment Pharmacol Ther. 1998;  12 1279-1289
• 18 Aguilar J L, Rojas P, Marcelo A, Plaza A, Bauer R, Reininger E, Klaas C A, Merfort I. Antiinflammatory activity of two different extracts of Uncaria tomentosa (Rubiaceae).  J Ethnopharmacol. 2002;  81 271-276
• 19 Wurm M, Kacani L, Laus G, Keplinger K, Dierich M P. Pentacyclic oxindole alkaloids from Uncaria tomentosa induce human endothelial cells to release a lymphocyte-proliferation regulating factor.  Planta Med. 1998;  64 701-704
• 20 Akesson C, Pero R W, Ivars F. C-Med 100, a hot water extract of Uncaria tomentosa, prolongs lymphocyte survival in vivo.  Phytomedicine. 2003;  10 23-33
• 21 Sheng Y, Pero R W, Amiri A, Bryngelsson C. Induction of apoptosis and inhibition of proliferation in human tumor cells treated with extracts of Uncaria tomentosa.  Anticancer Res. 1998;  18 3363-3368
• 22 Riva L, Coradini D, Di Fronzo G. The antiproliferative effects of Uncaria tomentosa extracts and fractions on the growth of breast cancer cell line.  Anticancer Res. 2001;  21 2457-2461
• 23 Pilarski R, Poczekaj-Kostrzewska M, Ciesiolka D, Szyfter K, Gulewicz K. Antiproliferative activity of various Uncaria tomentosa preparations on HL-60 promyelocytic leukemia cells.  Pharmacol Rep. 2007;  59 565-572
• 24 García Prado E, García Gimenez M D, De la Puerta Vazquez R, Espartero Sanchez J L, Sáenz Rodríguez M T. Antiproliferative effects of mitraphylline, a pentacyclic oxindole alkaloid of Uncaria tomentosa on human glioma and neuroblastoma cell lines.  Phytomedicine. 2007;  14 280-284
• 25 Muñoz M, Rosso M, Pérez A, Coveñas R, Rosso R, Zamarriego C, Piruat J I. The NK1 receptor is involved in the antitumoral action of L-733, 060 and in the mitogenic action of substance P on neuroblastoma and glioma cell lines.  Neuropeptides. 2005;  39 427-432
• 26 Seki A, Takayama H, Aimi N, Salai S L, Ponglux D. A nuclear magnetic resonance study on the eleven stereoisomers of heteroyohimbine-type oxindole alkaloids.  Chem Pharm Bull. 1993;  41 2077-2086
• 27 Paradowska K, Wolniak M, Pisklak M, Gliński J A, Davey M H, Wawer I. ¹³C, ¹⁵N CPMAS NMR and GIAO DFT calculations of stereoisomeric oxindole alkaloids from Cat's Claw (Uncaria tomentosa).  Solid State Nucl Magn Reson. 2008;  34 202-209
• 28 Stuppner H, Sturm S, Geisen G, Zillian U, Konwalinka G. A differential sensitivity of oxindole alkaloids to normal and leukemic cell lines.  Planta Med. 1993;  59 A583
• 29 Bacher N, Tiefenthaler M, Sturm S, Stuppner H, Ausserlechner M J, Kofler R, Konwalinka G. Oxindole alkaloids from Uncaria tomentosa induce apoptosis in proliferating, G0/G1-arrested and bcl-2-expressing acute lymphoblastic leukaemia cells.  Br J Haematol. 2006;  132 615-622