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Kidney cancer and melanoma: progresses in clinical and translational research ROME, November 23-24, 2007 EPIGENETIC IMMUNOTHERAPY OF HUMAN MELANOMA: MOLECULAR.

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Presentation on theme: "Kidney cancer and melanoma: progresses in clinical and translational research ROME, November 23-24, 2007 EPIGENETIC IMMUNOTHERAPY OF HUMAN MELANOMA: MOLECULAR."— Presentation transcript:

1 Kidney cancer and melanoma: progresses in clinical and translational research ROME, November 23-24, 2007 EPIGENETIC IMMUNOTHERAPY OF HUMAN MELANOMA: MOLECULAR BASES AND PERSPECTIVE CLINICAL APPLICATIONS Luca Sigalotti Cancer Bioimmunotherapy Unit, Centro di Riferimento Oncologico, I.R.C.C.S, Aviano

2 EPIGENETIC ALTERATIONS

3 EPIGENETICS Heritable changes in the expression of single genes or patterns of genes not based on modifications of the DNA sequence Methylation in C5 of cytosine within CpG dinucleotides Histone modifications Changes in chromatin structure TO TRANSCRIBE OR NOT TO TRANSCRIBE ?

4 Hypomethylation (gene expression) Hypermethylation (gene silencing) Morgan et al., Hum Mol Genet (2005)

5 GENETIC ALTERATIONS IRREVERSIBLE EPIGENETIC ALTERATIONS DYNAMIC

6 EPIGENETIC MODIFICATIONS ARE REVERSIBLE PHARMACOLOGICALLY Inhibitors of DNMT (e.g., 5-AZA-cytidine, 5-AZA-2- deoxycytidine, Zebularine) Inhibitors of HDAC (e.g., TSA, depsipeptide, SAHA,….)

7 5-AZA-2'-DEOXYCYTIDINE (5-AZA-CdR)

8 CH3 z 5-AZA-CdR DNA replication DNA methylation X CH3 z DNMT X CH3 z z z DNMT CH3 z z Hypomethylated DNA DNMT

9 Epigenetically-regulated immune molecules in cutaneous melanoma

10 CYTOTOXIC T CELLS AND/OR ANTIBODY-DEFINED TUMOR-ASSOCIATED ANTIGENS

11 Different families of related antigens: MAGE, NY-ESO and SSX gene families and GAGE/PAGE/XAGE super- families ……….. CANCER TESTIS ANTIGENS (CTA)

12 ADVANTAGES OF CTA AS THERAPEUTIC TARGETS

13 EXPRESSION OF MAA IN SEQUENTIAL AND CONCOMITANT MELANOMA METASTASES

14 CTA EXPRESSION IN MELANOMA STEM CELLS

15 However…….

16 CO-EXPRESSION OF 7 CTA IN 53 METASTATIC MELANOMAS 0 CTA (11 %) 1 CTA (24 %) 2 CTA (21 %) 3 CTA (13 %) 4 CTA (13 %) 5 CTA (9 %) 6CTA (9 %) 7CTA (0 %)

17 Roeder et al., Arch Dermatol Res (2005)

18 Can epigenetic drugs help?

19

20 REGULATION OF CTA EXPRESSION IN HUMAN MELANOMA XENOGRAFTS BY 5-AZA-CdR

21 PERSISTENCY OF 5-AZA-CdR-INDUCED CTA EXPRESSION IN HUMAN MELANOMA XENOGRAFTS NY-ESO-1 mol/ -actin mol

22 OD 405 IN VIVO GENERATION OF ANTI-NY-ESO-1 ANTIBODIES BY 5-AZA-CdR-TREATED MELANOMA CELLS

23 INTRATUMOR HETEROGENEITY

24 LEVELS OF MAGE-A3 mRNA EXPRESSED BY DIFFERENT SINGLE CELL CLONES FROM MEL 313 MELANOMA CELL LINE Clone 1 3 5 7 1234567891011121314Mel 313 cell line MAGE-3 mol/ -actin mol (x10 -3 )

25 CLONE 5 CLONE 14 ANALYSIS OF MAGE-A3 PROMOTER METHYLATION IN MEL 313 MELANOMA CLONES

26 0 4x10 -3 8x10 -3 1,2x10 -2 1,6x10 -2 0 1x10 -3 2x10 -3 3x10 -3 UP-REGULATION OF MAGE-3 EXPRESSION IN SINGLE CELL CLONES FROM MEL 313 MELANOMA CELL LINE BY 5-AZA-CdR Clone 5Clone 14 MAGE-3 mol/ -actin mol

27 RECOGNITION OF MEL313 MELANOMA CLONES BY A MAGE-3-SPECIFIC HLA-B37-RESTRICTED CTL CLONE

28

29 EFFECT OF 5-AZA-CdR ON LEVELS OF HLA CLASS I AND CO-STIMULATORY MOLECULES IN MEL 275 MELANOMA CELLS

30 RECOGNITION OF HLA-A2-POSITIVE MEL 275 MELANOMA CELLS BY AN ANTI-GP100 CTL CLONE 0 10 20 30 25:112:16:13:11.5:1 Ctrl 5-AZA-CdR % Lysis E:T ratio

31 5-AZA-CdR -HLA Class I -ICAM-1 ENHANCED AMOUNT OF IFN- RELEASING GP100-SPECIFIC HLA-A2- RESTRICTED CTL IN RESPONSE TO 5-AZA-CDR-TREATED MEL 275 MELANOMA CELLS

32 DE NOVO EXPRESSION OF CTA IN AML AND MDS PATIENTS TREATED WITH A SINGLE COURSE OF 5-AZA-CdR CTA-positive/ total samples T0T0 2/33 T 15 32/33 T 30 10/15 T 40 3/3 T: indicates time (days) from the beginning of Decitabine treatment

33

34 PHARMACOLOGIC DNA HYPOMETHYLATION AS A POSITIVE REGULATOR OF THE BIOLOGY OF CANCER CELLS Epigenetic drugs immune recognition cell cycle apoptosis angiogenesis invasion & metastasis

35 CTA-based Vaccine(s) Systemic Administration of 5-AZA-CdR Epigenetic chemoimmunotherapy Methylation

36 MEDICAL ONCOLOGY AND IMMUNOTHERAPY DEPT. OF MEDICAL ONCOLOGY UNIVERSITY HOSPITAL OF SIENA CANCER BIOIMMUNOTHERAPY UNIT DEPT. OF MEDICAL ONCOLOGY, CRO AVIANO Maresa Altomonte Lucia Anzalone Edi Bolzanaro Lorelai Brasoveanu Luana Calabrò Ilaria Cattarossi Francesca Colizzi Enzo Cortini Sandra Coral Alessia Covre Riccardo Danielli Chiara De Nardo Anna Maria Di Giacomo Elisabetta Fratta Ester Fonsatti Massimo Guidoboni Annunziata Gloghini Elda Lamaj Antonia Anna Lettini Samuele Massarut Giampaolo Nardi Hugues Nicolay Laura Pezzani Cristina Santantonio Luca Sigalotti Daniela Marconi


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