Abstract
In Drosophila, the Slit gene product, a secreted glycoprotein, acts as a midline repellent to guide axonal development during embryogenesis. Three human Slit gene orthologues have been characterised and recently we reported frequent promoter region hypermethylation and transcriptional silencing of
SLIT2
in lung, breast, colorectal and glioma cell lines and primary tumours. Furthermore, re-expression of
SLIT2
inhibited the growth of cancer cell lines so that
SLIT2
appears to function as a novel tumour suppressor gene (TSG). We analysed the expression of
SLIT3
(5q35–34) and
SLIT1
(1q23.3–q24) genes in 20 normal human tissues. Similar to
SLIT2
expression profile,
SLIT3
is expressed strongly in many tissues, while
SLIT1
expression is neuronal specific. We analysed the 5′ CpG island of
SLIT3
and
SLIT1
genes in tumour cell lines and primary tumours for hypermethylation.
SLIT3
was found to be methylated in 12 out of 29 (41%) of breast, one out of 15 (6.7%) lung, two out of six (33%) colorectal and in two out of (29%) glioma tumour cell lines. In tumour cell lines, silenced
SLIT3
associated with hypermethylation and was re-expressed after treatment with 5-aza-2′-deoxycytidine. In primary tumours,
SLIT3
was methylated in 16% of primary breast tumours, 35% of gliomas and 38% of colorectal tumours. Direct sequencing of bisulphite-modified DNA from methylated tumour cell lines and primary tumours demonstrated that majority of the CpG sites analysed were heavily methylated. Thus, both
SLIT2
and
SLIT3
are frequently methylated in gliomas and colorectal cancers, but the frequency of
SLIT3
methylation in lung and breast cancer is significantly less than that for
SLIT2
. We also demonstrated
SLIT1
promoter region hypermethylation in glioma tumour lines (five out of six; 83%), the methylation frequency in glioma tumours was much lower (two out of 20; 10%). Hence, evidence is accumulating for the involvement of members of the guidance cues molecules and their receptors in tumour development.