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EPIGENETIC IMMUNOTHERAPY OF HUMAN MELANOMA: MOLECULAR BASES

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Presentation on theme: "EPIGENETIC IMMUNOTHERAPY OF HUMAN MELANOMA: MOLECULAR BASES"— Presentation transcript:

1 EPIGENETIC IMMUNOTHERAPY OF HUMAN MELANOMA: MOLECULAR BASES
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 Heritable changes in the expression
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 Morgan et al., Hum Mol Genet (2005)
Hypomethylation (gene expression) Hypermethylation (gene silencing)

5 EPIGENETIC ALTERATIONS
DYNAMIC IRREVERSIBLE

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

9 Epigenetically-regulated
immune molecules in cutaneous melanoma

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

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

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

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

22 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 7 5 MAGE-3 mol/b-actin mol (x10-3) 3 1 Mel 313 cell line 1 2 3 4 5 6 7 8 9 10 11 12 13 14 Clone

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

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

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
10 20 30 25:1 12:1 6:1 3:1 1.5:1 Ctrl 5-AZA-CdR % Lysis E:T ratio

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

32 CTA-positive/ total samples T0 2/33 T15 32/33 T30 10/15 T40 3/3
DE NOVO EXPRESSION OF CTA IN AML AND MDS PATIENTS TREATED WITH A SINGLE COURSE OF 5-AZA-CdR CTA-positive/ total samples T0 2/33 T15 32/33 T30 10/15 T40 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 cell cycle apoptosis angiogenesis invasion & metastasis Epigenetic drugs immune recognition

35 “Epigenetic” chemoimmunotherapy
Systemic Administration of 5-AZA-CdR Methylation “Epigenetic” chemoimmunotherapy So (therefore), the final design of our approach will combine these two therapeutic modalities. Further support to the most effective design of this clinical approach will also be provided clinical and laboratory data that are being generated …… decitabine treatment will hamper the emergence of tumor immune escape, boosting the clinical effects of CTA-based vaccines ……. CTA-based Vaccine(s)

36 MEDICAL ONCOLOGY AND IMMUNOTHERAPY DEPT
MEDICAL ONCOLOGY AND IMMUNOTHERAPY DEPT. OF MEDICAL ONCOLOGY UNIVERSITY HOSPITAL OF SIENA CANCER BIOIMMUNOTHERAPY UNIT DEPT. OF MEDICAL ONCOLOGY, CRO AVIANO 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 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


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