Targeted modified TNF family members
The present invention relates to a modified cytokine of the TNF superfamily, with reduced activity to its receptor, wherein said modified cytokine is specifically delivered to target cells. Preferably, said modified cytokine is a single chain variant of the TNF superfamily, even more preferably, one or more of the chains carry one or more mutations, resulting in a low affinity to the receptor, wherein said mutant cytokine is specifically delivered to target cells. The targeting is realized by fusion of the modified cytokine of the TNF superfamily to a targeting moiety, preferably an antibody or antibody-like molecule. The invention relates further to the use of such targeted modified cytokine of the TNF superfamily to treat diseases.
1. A composition comprising a fusion protein, wherein the fusion protein comprises:
(i) a single chain polypeptide comprising three modified human TNFs, wherein each modified human TNF comprises a modified amino acid residue by substitution at the same position selected from Y87, I97, or Y115 relative to wild type human TNF (SEQ ID NO: 14), and wherein the modified human TNFs have reduced affinity towards its receptor as compared to wild type human TNF;
(ii) a targeting moiety directed to a cellular target, wherein the targeting moiety is directed towards a target selected from CD20, Her2, c-Met, EGFR, tenascin C, αvβ3 integrin, CD13, CD33, CD47, CD70, Axl, PSCA, and PSMA, and
wherein the fusion protein has significant biological activity towards cells that are targeted by the targeting moiety.
2. The composition of claim 1 , wherein the Y87 substitution is selected from Y87Q, Y87L, Y87A, and Y87F.
3. The composition of claim 1 , wherein the I97 substitution is selected from I97A, I197Q and I97S.
4. The composition of claim 1 , wherein the Y115 substitution is selected from Y115A and Y115G.
5. The composition of claim 1 , wherein the targeting moiety comprises an antibody or a variable domain of a camelid heavy chain antibody (VHH).
6. The composition of claim 5 , wherein the targeting moiety is a VHH.
7. The composition of claim 1 , wherein the targeting moiety is directed towards CD20.
8. The composition of claim 1 , wherein the targeting moiety is directed towards Her2.
9. The composition of claim 7 , wherein the targeting moiety is a VHH.
10. The composition of claim 8 , wherein the targeting moiety is a VHH.
11. The composition of claim 1 , wherein the substitution position is Y87 and the targeting moiety is a VHH directed towards CD20.
12. The composition of claim 11 , wherein the mutation is selected from Y87Q, Y87L, Y87A, or Y87F.