Patent EP-0418308-A4 (EP)
Title: Recombinant techniques for production of novel natriuretic and vasodilator peptides
IPC: C12N1/19
Filed: 1989-05-31
Abstract: The cDNA sequence encoding porcine brain natriuretic peptide and related genes encoding canine and human peptides with natriuretic activity are disclosed. The gene is shown to make accessible the DNAs encoding analogous natriuretic peptides in other vertebrate species. The genes encoding these NPs can be used to effect modifications of the sequence to produce alternate forms of the NPs and to provide practical amounts of these proteins. The NPs of the invention can also be synthesized chemically. The invention peptides have the formula: <tables id="TABLE-US-00001" num="00001"> <table frame="none" colsep="0" rowsep="0" tabstyle="monospace"> <tgroup align="left" colsep="0" rowsep="0" cols="2"> <colspec colname="1" colwidth="217pt" align="left"/> <colspec colname="2" colwidth="0pt" align="left"/> <TBODY VALIGN="TOP"> <ROW> <ENTRY>R<SUP>1</SUP>-Cys-Phe-Gly-Arg- Arg/ - Leu/ -Asp-Arg-</ENTRY> <ENTRY/> </ROW> <ROW> <ENTRY> Lys Met</ENTRY> </ROW> <ROW> <ENTRY> (1)</ENTRY> </ROW> <ROW> <ENTRY> Ile- Gly/ -Ser- Leu/ -Ser-Gly-Leu-Gly-Cys-R-<SUP>2</SUP></ENTRY> </ROW> <ROW> <ENTRY> Ser Ser</ENTRY> </ROW> </TBODY> </TGROUP> </TABLE> </TABLES> wherein R<SUP>1 </SUP>is selected from the group consisting of: <tables id="TABLE-US-00002" num="00002"> <table frame="none" colsep="0" rowsep="0" tabstyle="monospace"> <tgroup align="left" colsep="0" rowsep="0" cols="2"> <colspec colname="1" colwidth="217pt" align="left"/> <colspec colname="2" colwidth="0pt" align="left"/> <TBODY VALIGN="TOP"> <ROW> <ENTRY> (H);</ENTRY> <ENTRY/> </ROW> <ROW> <ENTRY> Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Asp/</ENTRY> </ROW> <ROW> <ENTRY> Lys/ -Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Gly</ENTRY> </ROW> <ROW> <ENTRY> Arg/ Asp/</ENTRY> </ROW> <ROW> <ENTRY> His/ - Lys/ -Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Gln Gly</ENTRY> </ROW> <ROW> <ENTRY> Arg/ Asp/</ENTRY> </ROW> <ROW> <ENTRY> Met/ - His/ - Lys/ -Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Val Gln Gly</ENTRY> </ROW> <ROW> <ENTRY> Arg/ Asp/</ENTRY> </ROW> <ROW> <ENTRY> Thr/ - Met/ - His/ - Lys/ -Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Met Val Gln Gly</ENTRY> </ROW> <ROW> <ENTRY> Arg/ Asp/</ENTRY> </ROW> <ROW> <ENTRY> Lys- Thr/ - Met/ - His/ - Lys/ -Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Met Val Gln Gly</ENTRY> </ROW> <ROW> <ENTRY> Arg/ Asp/</ENTRY> </ROW> <ROW> <ENTRY> Pro-Lys- Thr/ - Met/ - His/ - Lys/ -Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Met Val Gln Gly</ENTRY> </ROW> <ROW> <ENTRY> Arg/ Asp/</ENTRY> </ROW> <ROW> <ENTRY>Ser-Pro-Lys- Thr/ - Met/ - His/ - Lys/ -Ser-Gly-;</ENTRY> </ROW> <ROW> <ENTRY> Met Val Gln Gly</ENTRY> </ROW> </TBODY> </TGROUP> </TABLE> </TABLES> or a 10- to 109-amino acid sequence shown as the native upstream sequence for porcine, canine or human BNP in FIG. 8 , or a composite thereof; R<SUP>2 </SUP>is (OH), NH<SUB>2</SUB>, or NR'R'' wherein R' and R'' are independently lower alkyl (1-4C) or is <tables id="TABLE-US-00003" num="00003"> <table frame="none" colsep="0" rowsep="0" tabstyle="monospace"> <tgroup align="left" colsep="0" rowsep="0" cols="3"> <colspec colname="offset" colwidth="42pt" align="left"/> <colspec colname="1" colwidth="175pt" align="left"/> <colspec colname="2" colwidth="0pt" align="left"/> <TBODY VALIGN="TOP"> <ROW> <ENTRY/> <ENTRY>Asn/</ENTRY> <ENTRY/> </ROW> <ROW> <ENTRY/> <ENTRY>Lys</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Asn/ -Val</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Lys</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Asn/ -Val-Leu</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Lys</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Asn/ -Val-Leu-Arg</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Lys</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Asn/ -Val-Leu-Arg- Arg/</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Lys Lys</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Asn/ -Val-Leu-Arg- Arg/ - Tyr/</ENTRY> </ROW> <ROW> <ENTRY/> <ENTRY>Lys Lys His</ENTRY> </ROW> </TBODY> </TGROUP> </TABLE> </TABLES> or the amides (NH<SUB>2 </SUB>or NR'R'') thereof, with the proviso that if formula (1) is <tables id="TABLE-US-00004" num="00004"> <table frame="none" colsep="0" rowsep="0" tabstyle="monospace"> <tgroup align="left" colsep="0" rowsep="0" cols="3"> <colspec colname="offset" colwidth="21pt" align="left"/> <colspec colname="1" colwidth="196pt" align="left"/> <colspec colname="2" colwidth="0pt" align="left"/> <TBODY VALIGN="TOP"> <ROW> <ENTRY/> <ENTRY>R<SUP>1</SUP>-Cys-Phe-Gly-Arg-Arg-Leu-Asp-Arg-</ENTRY> <ENTRY/> </ROW> <ROW> <ENTRY/> <ENTRY> Ile-Gly-Ser-Leu-Ser-Gly-Leu-Gly-Cys-R<SUP>2</SUP></ENTRY> </ROW> </TBODY> </TGROUP> </TABLE> </TABLES> and R<SUP>1 </SUP>is Asp-Ser-Gly-, R<SUP>2 </SUP>cannot be Asn-Val-Leu-Arg-Arg-Tyr. The peptides of the invention can be formulated into pharmaceutical compositions and used to treat conditions associated with high extracellular fluid levels, especially congestive heart failure.