Stabilized transient gene expression

A method of enhancing the transient expression of a foreign gene in a eukaryotic cell in vitro comprising:

• introducing into the cell a molecule of foreign DNA that encodes a protein in a form capable of being expressed in the cell;
• contacting the cell with a first agent during the introduction into the cell of the foreign DNA, wherein the agent comprises at least one compound having the formula:
• ##STR4## wherein R.sub.1 is:
• CHNH.sub.2 R.sub.3, wherein R.sub.3 is the side chain of a naturally occurring amino acid;
• C.sub.6 H.sub.4 R.sub.4, wherein R.sub.4 is H, CH.sub.3, (CH.sub.2).sub.n CH.sub.3, NH.sub.2, COCH.sub.3, CO(CH.sub.2).sub.n CH.sub.3, C(CH.sub.3).sub.3, CH(CH.sub.3).sub.2, (CH.sub.2).sub.n CH(CH.sub.3).sub.2, (CH.sub.2).sub.n COCH.sub.3, OCH.sub.3, or O(CH.sub.2).sub.n CH.sub.3, wherein n=1-3;
• CHNH.sub.2 (CH.sub.2).sub.n R.sub.5, wherein n=1-7 and R.sub.5 is CH.sub.3, OH, CONH.sub.2, C.sub.6 H.sub.4 OH, or CONHNH.sub.2 ;
• (CH.sub.2).sub.n R.sub.6, wherein n=1-9 and R.sub.6 is an indole group, NCH.sub.3 C(.dbd.NH)NH.sub.2, SCH.sub.3, NH.sub.2, CH.sub.3, CO.sub.2 H, CONH.sub.2, or NHC(.dbd.NH)NH.sub.2, provided that when n=2 and R.sub.2 is H or M, R.sub.6 is not CH.sub.3 ;
• (CH.sub.2).sub.n CHNH.sub.2 CO.sub.2 H, wherein n=1-8;
• CH(CO.sub.2 H)NHCONH.sub.2 ;
• or,
• C.sub.5 H.sub.4 N;
• and
• wherein R.sub.2 is H, CH.sub.3, (CH.sub.2).sub.n CH.sub.3 wherein n=1-8, (CH.sub.2).sub.x O(CH.sub.2).sub.y CH.sub.3 or (CH.sub.2).sub.x CO(CH.sub.2).sub.y CH.sub.3 wherein x+y=2-7, or M, wherein M is a metal counterion or a low molecular weight organic cation;
• or the first agent comprises at least one chemical compound having the formula:
• R.sub.7 --SO.sub.2 --OR.sub.8,
• wherein R.sub.7 is a lower alkyl, aryl, substituted lower alkyl, or substituted aryl;
• and
• R.sub.8 is a hydrogen atom, a metal counterion, or a low molecular weight organic cation;
• and during and/or following the introduction of the foreign DNA, the cell is contacted with a second agent, wherein the second agent comprises at least one sulfonated amino polysaccharide or wherein the second agent comprises at least one chemical compound having the formula:
• ##STR5## wherein R.sub.1 is:
• CHNH.sub.2 R.sub.3, wherein R.sub.3 is the side chain of a naturally occurring amino acid;
• C.sub.6 H.sub.4 R.sub.4, wherein R.sub.4 is H, CH.sub.3, (CH.sub.2).sub.n CH.sub.3, NH.sub.2, COCH.sub.3, CO(CH.sub.2).sub.n CH.sub.3, C(CH.sub.3).sub.3, CH(CH.sub.3).sub.2, (CH.sub.2).sub.n CH(CH.sub.3).sub.2, (CH.sub.2).sub.n COCH.sub.3, OCH.sub.3, or O(CH.sub.2).sub.n CH.sub.3, wherein n=1-3;
• CHNH.sub.2 (CH.sub.2).sub.n R.sub.5, wherein n=1-7 and R.sub.5 is CH.sub.3, OH, CONH.sub.2, C.sub.6 H.sub.4 OH, or CONHNH.sub.2 ;
• (CH.sub.2).sub.n R.sub.6, wherein n=1-9 and R.sub.6 is an indole group, NCH.sub.3 C(.dbd.NH)NH.sub.2, SCH.sub.3, NH.sub.2, CH.sub.3, CO.sub.2 H, CONH.sub.2, or NHC(.dbd.NH)NH.sub.2, provided that when n=2 and R.sub.2 is H or M, R.sub.6 is not CH.sub.3 ;
• (CH.sub.2).sub.n CHNH.sub.2 CO.sub.2 H, wherein n=1-8;
• CH(CO.sub.2 H)NHCONH.sub.2 ;
• or,
• C.sub.5 H.sub.4 N;
• and
• wherein R.sub.2 is H, CH.sub.3, (CH.sub.2).sub.n CH.sub.3 wherein n=1-8, (CH.sub.2).sub.x O(CH.sub.2).sub.y CH.sub.3 or (CH.sub.2).sub.x CO(CH.sub.2).sub.y CH.sub.3 wherein x+y=2-7, or M, wherein M is a metal counterion or a low molecular weight organic cation;
• or the second agent comprises at least one chemical compound having the formula:
• R.sub.7 --SO.sub.2 --OR.sub.8,
• wherein R.sub.7 is a lower alkyl, aryl, substituted lower alkyl, or substituted aryl;
• and
• R.sub.8 is a hydrogen, a metal counterion, or a low molecular weight organic cation;
• and
• wherein the first agent is one that has a value for X that is greater than 25 when X is calculated according to the formula:
• ##EQU7## and wherein the second agent is one that has a value of G.sub.7 or G.sub.14 that is greater than 25, wherein G.sub.7 or G.sub.14 is calculated according to the formula:
• ##EQU8## wherein for both X or G.sub.7 or G.sub.14, "A" is the amount of the protein encoded by the foreign gene expressed in a first cell that is contacted with the first and the second agent, and "C" is the amount of protein expressed in a second cell that is not contacted with the first or the second agent.

A method of enhancing the transient expression of a foreign gene in a eukaryotic cell in vitro comprising:

• introducing into the cell a molecule of foreign DNA that encodes a protein in a form capable of being expressed in the cell;
• and,
• contacting the cell before, during, or after introducing the DNA with a transient expression enhancing agent;
• wherein the transient expression enhancing agent comprises:
• a compound selected from the group consisting of 3-methyl-L-histidine, .alpha.-ketoglutaric acid, 9-alanine, carnosine, citrulline, creatine, glutathione, hippuric acid, homoserine, N-carbamyl aspartic acid, N-formyl-L-methionine, ornithine, N-(4-aminobenzyl)-L-glutamic diethylester, ethyl-4-acetylbutyrate, adrenaline, methylcobalamin, benzoic acid, benzoate buffer, 4-ethylbenzoic acid and a sulfonated N-acetylated amino polysaccharide, disaccharide monomeric units derived from Type C chondroitin sulfate, polymannose, mannose;
• or
• a sulfonic acid derivative having the formula:
• R.sub.7 --SO.sub.2 --OR.sub.8
• wherein R.sub.7 is a lower alkyl, aryl, substituted lower alkyl, aryl, substituted lower alkyl, or substituted aryl, and R.sub.8 is a hydrogen, a metal counterion, or a low molecular weight organic cation;
• or
• benzoic acid and 4-ethylbenzoic acid;
• or
• benzoic acid and 4-ethylbenzoic acid and chondroitin sulfate;
• or
• benzoic acid and L-glutamine;
• or
• benzoate buffer and chondroitin sulfate;
• or
• benzoate buffer and glutamic acid;
• or
• lipoic acid and chondroitin sulfate;
• or
• benzoate buffer, chondroitin sulfate, and L-glutamine;
• or
• chondroitin sulfate and L-glutamine;
• or
• butyrate buffer and L-glutamine;
• or
• a mixture comprising glutathione, methionine, glycine, .alpha.-amino-n-butyric acid, taurine, phenylalanine, benzoate buffer, and alanine.