1. Alterazioni dei livelli intracellulari di PCho e tCho durante la progressione tumorale o in risposta a trattamenti antitumorali mirati riflettono modifiche a livello genico, trascrizionale e/o post-trascrizionale, correlate alla deregolazione e riorganizzazione dei meccanismi molecolari responsabili della proliferazione cellulare, sopravvivenza e capacità invasiva della cellula (12).
2. Analisi MRS di cellule tumorali in coltura forniscono nuove informazioni sui meccanismi molecolari coinvolti in alterazioni del metabolismo e del signaling cellulare responsabili del fenotipo tumorale, e possono consentire l’identificazione e il successivo utilizzo clinico di nuovi biomarcatori di risposta a terapie mirate.
3. Lo studio del metabolismo della colina, in combinazione con analisi di espressione genica e proteica può aprire nuove aree di ricerca in oncologia molecolare, come illustrato dalle evidenze ottenute per la prima volta nel nostro laboratorio sull’esistenza di interazioni specifiche tra HER2 e PtdCho-PLC e sul ruolo dell’attivazione di questa fosfolipasi sulla overespressione del recettore e sullo stato proliferativo di cellule di carcinoma mammario. Risultati ottenuti nel nostro laboratorio mostrano analoghe evidenze per il carcinoma ovarico (L. Paris e collaboratori, manoscritto in preparazione e 21).
4. L’utilizzo di protocolli di quantificazione metabolica in opportuni modelli preclinici può rendere fattibile lo sviluppo e la valutazione di nuovi approcci di imaging multiparametrico e multimodale non invasivo in vivo (MRS, colina-PET), per il monitoraggio della risposta di pazienti oncologici a terapie mirate.
OVCA432 xenotrapianto sub cutaneo
OVCA432 xenotrapianto intraperitoneale
119
Ringraziamenti
Gli autori ringraziano la dott.ssa Silvana Canevari, la dott.ssa Delia Mezzanzanica e la dott.ssa Marina Bagnoli (Fondazione IRCCS Istituto Nazionale dei Tumori, Milano); dott. ZM Bhujwalla e dott. K Glunde (Johns Hopkins University, School of Medicine, Baltimore, MA, Stati Uniti); il dott. Pier Giorgio Natali (Istituto Tumori Regina Elena, Roma); il dott. Francesco Sardanelli (Policlinico San Donato e Università degli Studi di Milano); il dott. Filippo Belardelli e la dott.ssa Carmen Rozera (Dipartimento di Biologia Cellulare e Neuroscienze, ISS) per l’illuminante, esperta e amichevole collaborazione scientifica; il dott. Massimo Di Vito per la collaborazione nello studio del pathway di deacilazione in cellule di tumore ovarico; Massimo Giannini per l’assistenza tecnica di alto livello prestata negli esperimenti di risonanza magnetica in vitro e in vivo; Massimo Spada, Albino Cesolini, Massimo Venditti e Emiliano Surrentino per l’esperta assistenza nell’allestimento dei modelli preclinici e nei loro esami in vivo.
Gli studi del gruppo hanno ricevuto supporto finanziario da parte di: Associazione Italiana per la Ricerca sul Cancro (AIRC 2007-2009); Ministero della Salute (Programma Integrato Oncologia RO 06.5/N.ISS/Q; Programma Ordinario Oncologia OncOrd 37/07/N. ISS/70CF/4;
Programma Speciale Alleanza Contro il Cancro 2006, ACC3-AC5/D; Accordo di Collaborazione Italia-Stati Uniti ISS/530F/0F2).
Bibliografia
1. Pelech SL, Vance DE. Signal transduction via phosphatidylcholine cycle. Trends Biochem Sci 1989;14:28-30.
2. Negendank W. Studies of human tumors by MRS: a review. NMR Biomed 1992;5(5):303-24.
3. de Certaines JD, Larsen VA, Podo F, Carpinelli G, Briot O, Henriksen O. In vivo 31P MRS of experimental tumours. NMR Biomed 1993;6(6):345-65.
4. Podo F. Tumour phospholipid metabolism. NMR Biomed 1999;12(7):413-39.
5. Gillies RJ, Morse DL. In vivo magnetic resonance spectroscopy in cancer. Annu Rev Biomed Eng 2005;7:287-326.
6. Podo F, Sardanelli F. Iorio E, Canese R, Carpinelli, G, Fausto F. and Canevari S. Abnormal choline phospholipid metabolism in breast and ovary cancer: molecular bases for non invasive imaging approaches. Current Med Imaging Rev 2007;3:123-37.
7. Glunde K, Artemov D, Penet MF, Jacobs MA, Bhujwalla ZM. Magnetic resonance spectroscopy in metabolic and molecular imaging and diagnosis of cancer. Chem Rev 2010;110(5):3043-59.
8. Ackerstaff E, Glunde K, Bhujwalla ZM. Choline phospholipid metabolism: a target in cancer cells? J Cell Biochem 2003;90(3):525-33.
9. Glunde K, Ackerstaff E, Mori N, Jacobs MA, Bhujwalla ZM. Choline phospholipid metabolism in cancer: consequences for molecular pharmaceutical interventions. Mol Pharm 2006;3(5):496-506.
10. Glunde K, Serkova NJ. Therapeutic targets and biomarkers identified in cancer choline phospholipid metabolism. Pharmacogenomics 2006;7(7):1109-23.
11. Beloueche-Babari M, Chung YL, Al-Saffar NM, Falck-Miniotis M, Leach MO. Metabolic assessment of the action of targeted cancer therapeutics using magnetic resonance spectroscopy.
Br J Cancer 2010;102(1):1-7.
12. Podo F. Canevari S, Canese R, Pisanu ME, Ricci A, Iorio E. Magnetic resonance evaluation of response to targeted treatment in cancer cells. NMR Biomed 2011, Mar 8 (online prima della pubblicazione).
120
13. Ramoni C, Spadaro F, Menegon M, Podo F. Cellular localization and functional role of phosphatidylcholine-specific phospholipase C in NK cells. J Immunol 2001;167(5):2642-50.
14. Ferretti A, D’Ascenzo A, Knijn A, Iorio E, Dolo V, Pavan A, Podo F. Detection of polyol accumulation in a new ovarian carcinoma cell line, CABA I: a 1H NMR study. Br J Cancer 2002;86(7):1180-87.
15. Ramoni C, Spadaro F, Barletta B, Dupuis ML, Podo F. Phosphatidylcholine-specific phospholipase C in mitogen-stimulated fibroblasts. Exp Cell Res 2004;299(2):370-82.
16. Iorio E, Mezzanzanica D, Alberti P, Spadaro F, Ramoni C, D’Ascenzo S, Millimaggi D, Pavan A, Dolo V, Canevari S, Podo F. Alterations of choline phospholipid metabolism in ovarian tumor progression. Cancer Res 2005;65(20):9369-76.
17. Spadaro F, Cecchetti S, Sanchez M, Podo F, Ramoni C. Expression and role of phosphatidyl-choline-specific phospholipase C in human NK and T lymphocyte subsets. Eur J Immunol 2006;36(12):3277-87.
18. Cecchetti S, Spadaro F, Lugini L., Podo F, Ramoni C. Functional role of phosphatidylcholine-specific phospholipase C in regulating CD16 membrane expression in natural killer cells. Eur J Immunol 2007;37(10):2912-22.
19. Sardanelli F, Fausto A, Podo F. MR spectroscopy of the breast. Radiol Med 2008;113(1):56-64.
20. Fantuzzi L, Spadaro F, Purificato C, Cecchetti S, Podo F, Belardelli F, Gessani S, Ramoni C.
Phosphatidylcholine-specific phospholipase C activation is required for CCR5-dependent, NF-kB-driven CCL2 secretion elicited in response to HIV-1 gp120 in human primary macrophages. Blood 2008;111(7):3355-63.
21. Spadaro F, Ramoni C, Mezzanzanica D, Miotti S, Alberti P, Cecchetti S, Iorio E, Dolo V, Canevari S, Podo F. Phosphatidylcholine-specific phospholipase C activation in epithelial ovarian cancer cells. Cancer Res 2008;68(16):6541-49.
22. Sardanelli F, Fausto A, Di Leo G, de Nijs R, Verbuchner M, Podo F In vivo proton MR spectroscopy of the breast using the total choline peak integrals as a marker of malignancy. Am J Roentgen 2009;192(6):1608-17.
23. Canese R, Iorio E, Ricci A, Pisanu ME, Giannini M, Podo F. Metabolite quantification in tumours by magnetic resonance spectroscopy: objectives, results and perspectives. Current Med Imaging Rev 2009;5:110-27.
24. Iorio E, Ricci A, Bagnoli M, Pisanu ME, Castellano G, Di Vito M, Venturini E, Glunde K, Bhujwalla ZM, Mezzanzanica D, Canevari S, Podo F. Activation of phosphatidylcholine cycle enzymes in human epithelial ovarian cancer cells. Cancer Res 2010;70(5):2126-35.
25. Paris L, Cecchetti S, Spadaro F, Abalsamo L, Lugini L, Pisanu ME, Iorio E, Natali PG, Ramoni C, Podo F. Inhibition of phosphatidylcholine-specific phospholipase C downregulates HER2 overexpression on plasma membrane of breast cancer cells. Breast Cancer Res 2010;12(3)R27 (16 pp).
26. Esseridou A, Di Leo G, Sconfienza LM, Caldiera V, Raspagliesi F, Grijela B, Hanozet F, Podo F, Sardanelli F. In vivo detection of choline in ovarian tumors using 3D magnetic resonance spectroscopy. Invest Radiol 2011;46(6):377-82.
27. Abalsamo L, Spadaro F, Bozzuto G, Paris L, Cecchetti S, Lugini L, Iorio E, Molinari A, Ramoni C, Podo F. Inhibition of phosphatidylcholine-specific phospholipase C induces loss of mesenchymal traits in metastatic. Breast cancer cells 2011 (inviato per la pubblicazione).
28. Canese R, Pisanu ME, Mezzanzanica D, Ricci A, Paris L, Bagnoli M, Valeri B, Spada M, Venditti M, Cesolini A, Rodomonte A, Giannini M, Canevari S, Podo F, Iorio E. Characterization of the in vivo ovarian modles by quantitative 1H magnetic resonance spectroscopy and diffusion-weighted imaging. NMR Biomed 2011; Oct 24 (online prima della stampa).
121
29. Vance JE, Vance DE. Phospholipid biosynthesis in mammalian cells. Biochem Cell Biol 2004;82(1):113-28.
30. Cuadrado A, Carnero A, Dolfi F, Jimenez B, Lacal JC. Phosphorylcholine: a novel second messenger essential for mitogenic activity of growth factors. Oncogene 1993;8(11):2959-68.
31. Aoyama C, Liao H, Ishidate K. Structure and function of choline kinase isoforms in mammalian cells. Prog Lipid Res 2004;43(3):266-81.
32. Janardhan S, Srivani P, Sastry GN. Choline kinase: an important target for cancer. Curr Med Chem 2006;13(10):1169-86.
33. Aboagye EO, Bhujwalla ZM. Malignant transformation alters membrane choline phospholipid metabolism of human mammary epithelial cells. Cancer Res 1999;59(1):80-4.
34. Ackerstaff E, Pflug BR, Nelson JB, Bhujwalla ZM. Detection of increased choline compounds with proton nuclear magnetic resonance spectroscopy subsequent to malignant transformation of human prostatic epithelial cells.Cancer Res 2001;61(9):3599-603.
35. Eliyahu G, Kreizman T, Degani H. Phosphocholine as a biomarker of breast cancer: molecular and biochemical studies. Int J Cancer 2007;120(8):1721-30.
36. Ramirez de Molina A, Bañez-Coronel M, Gutierrez R, Rodriguez-Gonzalez A, Olmeda D, Megias D, Lacal JC. Choline kinase activation is a critical requirement for the proliferation of primary human mammary epithelial cells and breast tumor progression. Cancer Res 2004;64(18):6732-39.
37. Glunde K, Shah T, Winnard PT, Jr, Raman V, Takagi T, Vesuna F, Artemov D, Bhujwalla ZM.
Hypoxia regulates choline kinase expression through hypoxia-inducible factor-1 alpha signaling in a human prostate cancer model. Cancer Res 2008;68(1):172-80.
38. Chua BT, Gallego-Ortega D, Ramirez de Molina A, Ullrich A, Lacal JC, Downward J. Regulation of Akt(ser473) phosphorylation by choline kinase in breast carcinoma cells. Mol Cancer 2009;8:131.
39. Jimenez B, del Peso L, Montaner S, Esteve P, Lacal JC. Generation of phosphorylcholine as an essential event in the activation of Raf-1 and MAP-kinases in growth factors-induced mitogenic stimulation. J Cell Biochem 1995;57(1):141-49.
40. Ramirez de Molina A, Gutierrez R, Ramos MA, Silva JM, Silva J, Bonilla F, Sanchez JJ, Lacal JC.
Increased choline kinase activity in human breast carcinomas: clinical evidence for a potential novel antitumor strategy. Oncogene 2002;21(27):4317-22.
41. Ramirez de Molina A, Sarmentero-Estrada J, Belda-Iniesta C, Taron M, Ramirez de Molina V, Cejas P, Skrzypski M, Gallego-Ortega D, de Castro J, Casado E, Garcia-Cabezas MA, Sanchez JJ, Nistal M, Rosell R, Gonzalez-Baron M, Lacal JC. Expression of choline kinase alpha to predict outcome in patients with early-stage non-small-cell lung cancer: a retrospective study. Lancet Oncol 2007;8(10):889-97.
42. Hernando E, Sarmentero-Estrada J, Koppie T, Belda-Iniesta C, Ramirez de Molina V, Cejas P, Ozu C, Le C, Sanchez JJ, Gonzalez-Baron M, Koutcher J, Cordon-Cardo C, Bochner BH, Lacal JC, Ramirez de Molina A. A critical role for choline kinase-alpha in the aggressiveness of bladder carcinomas. Oncogene 2009;28(26):2425-35.
43. Lee Z, Swaby RF, Liang Y, Yu S, Liu S, Lu KH, Bast RC, Jr., Mills GB, Fang X.
Lysophosphatidic acid is a major regulator of growth-regulated oncogene alpha in ovarian cancer.
Cancer Res 2006;66(5):2740-48.
44. Foster DA, Xu L. Phospholipase D in cell proliferation and cancer. Mol Cancer Res 2003;1(11):789-800.
45. Buczynski MW, Dumlao DS, Dennis EA. Thematic Review Series: Proteomics. An integrated omics analysis of eicosanoid biology. J Lipid Res 2009; 50:1015-38.
46. Glunde K, Jie C, Bhujwalla ZM. Molecular causes of the aberrant choline phospholipid metabolism in breast cancer. Cancer Res 2004;64(12):4270-76.
122
47. Cifone MG, Rocaioli P, De Maria R, Camarda G, Santoni A, Ruberti G, Testi R. Multiple pathways origiated at the Fas/Apo-1 8CD95) receptor: sequential involvement of phopshatidylcholine-specific phospholipase c and acidic sphingomyelinase in the propagation of the apoptotic signal. EMBO J 1995;14:5859-68.
48. Wang N, Sun C, Huo S, Zhang Y, Zhao J, Zhang S, Miao J. Coopoeration of phosphatidylcholine-specific phospholipase C and basic fibroblast growth factor in the neural differentiation of mesenchymal stem cells in vitro. Int J Biochem Cell Biol 2008;40:294-306.
49. Al-Saffar NM, Troy H, Ramirez de Molina A, Jackson LE, Madhu B, Griffiths JR, Leach MO, Workman P, Lacal JC, Judson IR, Chung YL. Noninvasive magnetic resonance spectroscopic pharmacodynamic markers of the choline kinase inhibitor MN58b in human carcinoma models.
Cancer Res 2006;66(1):427-34.
50. Krishnamachary B, Glunde K, Wildes F, Mori N, Takagi T, Raman V, Bhujwalla ZM.
Noninvasive detection of lentiviral-mediated choline kinase targeting in a human breast cancer xenograft. Cancer Res 2009;69(8):3464-71.
51. Kyriazi S, Kaye SB, Desouza NM, Imaging ovarian cancer and peritoneal metastases - current and emerging techniques. Nat Rev Clin Oncol 2010;7(7):381-93.
123