The code above generates results you ask for. If that example data does not represent real data then you need to provide an appropriate example.

if the columns are not at the beginning of the table, you can extract the columns using cut:

cut -f2,4 *myfile* | sort | uniq > output

In that case, use the following modification:

$ awk -v FS=',' '!a[$2$4]++' input.txt > output.txt

This will report the first line seen for the combination of the 2nd and 4th columns. Second and subsequent "hits" are not reported.

If you want to instead use sort, you will need to use some additional options:

$ sort -u -k2,2 -k4,4 -t, input.txt > output.txt

Without reading the man pages, I'm unsure if sort is stable, so you might get a different answer on repeated runs.

In addition to flexibility on the keys used for filtering, the awk approach runs much faster on very large input (at the expense of memory), so if you're working with whole-genome scale input, then you may want to use awk, instead of sort | uniq or sort -u -based approaches.