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 method for treating cancer, comprising administering an effective amount of a composition comprising a fusion protein to a patient in need thereof, 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, and wherein said modified human TNFs have reduced affinity towards its receptor as compared to wild type human TNF;
(ii) a linker sequence; and
(iii) a targeting moiety directed to a cellular target, wherein the linker sequence links the modified human TNFs to targeting moiety, 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 shows significant biological activity towards cells that are targeted by the targeting moiety.
2. The method of claim 1 , wherein the Y87 mutation is selected from Y87Q, Y87L, Y87A, or Y87F.
3. The method of claim 1 , wherein the Y115 mutation is selected from Y115A or Y115G.
4. The method of claim 1 , wherein the I97 mutation is selected from I97A, I97Q or I97S.
5. The method of claim 1 , wherein the linker sequence comprises GGS.
6. The method of claim 5 , wherein the linker sequence comprises GGS repeats.
7. The method of claim 1 , wherein the targeting moiety is a variable domain of a camelid heavy chain antibody (VHH).
8. The method of claim 1 , wherein the targeting moiety is directed towards CD20.
9. The method of claim 1 , wherein the targeting moiety is directed towards Her2.
10. The method of claim 8 , wherein the targeting moiety is a VHH.
11. The method of claim 9 , wherein the targeting moiety is a VHH.
12. The method of claim 1 , wherein Y87 and the targeting moiety is a VHH directed towards CD20.
13. The method of claim 12 , wherein the mutation is selected from Y87Q, Y87L, Y87A, or Y87F.