Contributions
Abstract: PB1851
Type: Publication Only
Background
PRAME is the most frequently expressed non-X-chromosomal cancer-testis gene in solid and hematological cancer. It is important, because PRAME often has a bad prognostic significance. In early studies was found that PRAME frequently coexpressed in translocation-harboring (like t(8;21), t(15;17) and t(9;22)) haematological diseases. Authors supposed that chimeric genes are activators of PRAME expression. But in large cases with normal karyotype PRAME is also expressed. Another reason for PRAME expression is promoter demethylation. But demethylating agents cannot activate PRAME expression in hematological cells taken from healthy donor. So presence of chimeric genes and methylation status only are not enough to explain why PRAME can be expressed in high level. Wadelin et al. found that PRAME expression level was increased in cell during lipopolysaccharide-treatment conditions. Role of MYD88 in this process still be unknown.
Aims
To check if MYD88 participates in activating PRAME expression in leukemia cell lines.
Methods
Three cell lines were used for incubation with anti-PRAME antibody: chronic myeloid leukemia cell line K562 with high PRAME expression level (645%), acute monocytic leukemia cell line THP-1 with intermediate PRAME expression level (2,92% relative to ABL) and acute myeloid leukemia cell line NOMO-1 with low PRAME expression level (0,46%). All cell lines were incubated in RPMI 1640 with addition of LPS in final concentration 10 ng/ml. After 1 and 4 hour of incubation total RNA was extracted and PRAME and MYD88 expression levels were measured.
Results
After 1 and 4 hours of experiment in K562 cell line PRAME expression level was increased in 2,7 and 7 fold under control, respectively, and MYD88 expression level increased in 1,1 and 2,5 fold under control. In THP-1 line PRAME expression level was increased in 20 and 25 fold, respectively, and MYD88 expression level was increased in 5,5 and 6,5 fold. In cell line NOMO-1 PRAME expression level was increased in 10 fold after 1 hour and in 14 fold after 4 hours, and MYD88 expression level was increased in 2,4 and 3,2 fold after 1 and 4 hours of experiment, respectively. Strong correlation between MYD88 and PRAME expression levels was observed (Pearson correlation coefficient 0,98).
Conclusion
We conclude that LPS after binding with TLRs initiates activating signal to PRAME gene via MYD88.
Session topic: 24. Gene therapy, cellular immunotherapy and vaccination
Keyword(s): Signaling, Innate Immunity
Abstract: PB1851
Type: Publication Only
Background
PRAME is the most frequently expressed non-X-chromosomal cancer-testis gene in solid and hematological cancer. It is important, because PRAME often has a bad prognostic significance. In early studies was found that PRAME frequently coexpressed in translocation-harboring (like t(8;21), t(15;17) and t(9;22)) haematological diseases. Authors supposed that chimeric genes are activators of PRAME expression. But in large cases with normal karyotype PRAME is also expressed. Another reason for PRAME expression is promoter demethylation. But demethylating agents cannot activate PRAME expression in hematological cells taken from healthy donor. So presence of chimeric genes and methylation status only are not enough to explain why PRAME can be expressed in high level. Wadelin et al. found that PRAME expression level was increased in cell during lipopolysaccharide-treatment conditions. Role of MYD88 in this process still be unknown.
Aims
To check if MYD88 participates in activating PRAME expression in leukemia cell lines.
Methods
Three cell lines were used for incubation with anti-PRAME antibody: chronic myeloid leukemia cell line K562 with high PRAME expression level (645%), acute monocytic leukemia cell line THP-1 with intermediate PRAME expression level (2,92% relative to ABL) and acute myeloid leukemia cell line NOMO-1 with low PRAME expression level (0,46%). All cell lines were incubated in RPMI 1640 with addition of LPS in final concentration 10 ng/ml. After 1 and 4 hour of incubation total RNA was extracted and PRAME and MYD88 expression levels were measured.
Results
After 1 and 4 hours of experiment in K562 cell line PRAME expression level was increased in 2,7 and 7 fold under control, respectively, and MYD88 expression level increased in 1,1 and 2,5 fold under control. In THP-1 line PRAME expression level was increased in 20 and 25 fold, respectively, and MYD88 expression level was increased in 5,5 and 6,5 fold. In cell line NOMO-1 PRAME expression level was increased in 10 fold after 1 hour and in 14 fold after 4 hours, and MYD88 expression level was increased in 2,4 and 3,2 fold after 1 and 4 hours of experiment, respectively. Strong correlation between MYD88 and PRAME expression levels was observed (Pearson correlation coefficient 0,98).
Conclusion
We conclude that LPS after binding with TLRs initiates activating signal to PRAME gene via MYD88.
Session topic: 24. Gene therapy, cellular immunotherapy and vaccination
Keyword(s): Signaling, Innate Immunity