METHODS AND IMMUNOGENIC COMPOSITIONS RELATING TO HER2 WITH SELECTIVE SEQUENCE MODIFICATIONS
Only limited success has been previously achieved from cancer vaccines targeting unmodified tumor-associated self-antigens and new compositions and methods are needed. Immunogenic compositions and methods of use thereof are provided according to the present disclosure which include a protein effective to stimulate immune activity against a tumor-associated self-antigen, or a variant thereof which is a tumor-associated self-antigen.
1 . An immunogenic composition, comprising:
the amino acid sequence of SEQ ID NO:2 or a variant thereof, wherein the variant is not SEQ ID NO:1.
2 . The immunogenic composition of claim 1 , further comprising a pharmaceutically acceptable carrier.
3 . The immunogenic composition of claim 1 , further comprising an adjuvant.
4 . A recombinant expression construct, comprising:
a nucleic acid encoding the immunogenic composition of claim 1 , operably linked to a heterologous regulatory nucleic acid sequence.
5 . A host cell comprising the recombinant expression construct of claim 4 .
6 . A method of treatment or prevention of a HER2+ cancer in a subject, comprising:
administering a therapeutically effective amount of the immunogenic composition of claim 1 , to the subject.
7 . The method of claim 6 , wherein the immunogenic composition further comprises a pharmaceutically acceptable carrier.
8 . The method of claim 6 , wherein the immunogenic composition further comprises an immunostimulating adjuvant.
9 . The method of claim 6 , wherein the immunogenic composition is administered as a nucleic acid encoding the amino acid sequence of SEQ ID NO:2 operably linked to a heterologous regulatory nucleic acid sequence.
10 . The method of claim 9 , wherein the nucleic acid is present in an expression vector.
11 . The method of claim 10 , wherein the expression vector is a virus.
12 . The method of claim 1 , wherein the immunogenic composition is characterized by one or more of: 1) effectiveness to stimulate immune activity against a specified tumor-associated self-antigen in a subject, 2) effectiveness to overcome self-tolerance of the specified tumor-associated self-antigen, and 3) substantial similarity to the native three dimensional structure of the specified tumor-associated self-antigen.
13 . A method of generating an immunogenic composition, comprising:
identifying a reference sequence;
identifying at least a first amino acid sequence homologous to the reference sequence, wherein the reference sequence and the first amino acid sequence homologous to the reference sequence are not identical;
comparing the homologous amino acid sequence and the reference sequence to identify at least a first difference between the homologous amino acid sequence and the reference sequence at a first position in the reference sequence and a corresponding first position in the homologous amino acid sequence;
assigning a BLOSUM62 score to the first difference between the homologous amino acid sequence and the reference sequence, wherein the score represents a probability of substitution of an amino acid at the first position of the reference sequence with the amino acid at the corresponding first position in the homologous sequence, wherein the score falls within a numerical probability range of −4 to +3, where −4 is a number which indicates an extremely non-conservative substitution of the amino acid at the first position of the reference sequence with the amino acid at the corresponding first position of the homologous sequence such that the occurrence of substitution of the extremely non-conservative substitution is relatively rare or unlikely, where +3 is a number which indicates an extremely conservative substitution of the amino acid at the first position of the reference sequence with the amino acid at the corresponding first position of the homologous sequence such that the occurrence of substitution of the extremely non-conservative substitution is relatively frequent or likely, where a BLOSUM score of 0 indicates neutrality such that the occurrence of substitution has an equal probability;
synthesizing a new amino acid sequence identical to the reference amino acid sequence with the proviso that that the new amino acid sequence has at least one substitution at the first position with an amino acid present at the corresponding first position in the homologous amino acid sequence where the score assigned to the first difference is in the range of 0 to 1 and indicates neutrality such that the occurrence of substitution has an equal probability, thereby generating an immunogenic composition.
14 . The method of claim 13 , wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, or at least 15, amino acid sequences homologous to the reference sequence are identified and compared with the reference amino acid sequence.
15 . The method of claim 13 , wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, or more, differences between the homologous amino acid sequence and the reference sequence at corresponding 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions in the reference sequence and a corresponding position in the homologous amino acid sequence are identified and assigned a BLOSUM score.
16 . The method of claim 15 , wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, or at least 15, substitutions are made in the reference amino acid sequence at 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions in the reference sequence with an amino acid present at the corresponding 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions in the homologous amino acid sequence where the BLOSUM score assigned to the differences identified at the 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions is in the range of 0 to 1.
17 . A method of generating an immunogenic composition, comprising:
identifying a reference sequence;
identifying at least a first amino acid sequence homologous to the reference sequence, wherein the reference sequence and the first amino acid sequence homologous to the reference sequence are not identical;
comparing the homologous amino acid sequence and the reference sequence to identify at least a first difference between the homologous amino acid sequence and the reference sequence at a first position in the reference sequence and a corresponding first position in the homologous amino acid sequence;
assigning a score to the first difference between the homologous amino acid sequence and the reference sequence, wherein the score represents a probability of substitution of an amino acid at the first position of the reference sequence with the amino acid at the corresponding first position in the homologous sequence, wherein the score falls within a numerical probability range of x to y, where x is a number which indicates an extremely non-conservative substitution of the amino acid at the first position of the reference sequence with the amino acid at the corresponding first position of the homologous sequence such that the occurrence of substitution of the extremely non-conservative substitution is relatively rare or unlikely, where y is a number which indicates an extremely conservative substitution of the amino acid at the first position of the reference sequence with the amino acid at the corresponding first position of the homologous sequence such that the occurrence of substitution of the extremely non-conservative substitution is relatively frequent or likely, where a number intermediate between x and y indicates neutrality such that the occurrence of substitution has an equal probability;
synthesizing a new amino acid sequence identical to the reference amino acid sequence with the proviso that that the new amino acid sequence has at least one substitution at the first position with an amino acid present at the corresponding first position in the homologous amino acid sequence where the score assigned to the first difference is intermediate between x and y indicates neutrality such that the occurrence of substitution has an equal probability, thereby generating an immunogenic composition.
18 . The method of claim 17 , wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, or at least 15, amino acid sequences homologous to the reference sequence are identified and compared with the reference amino acid sequence.
19 . The method of claim 17 , wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, or more, differences between the homologous amino acid sequence and the reference sequence at corresponding 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions in the reference sequence and a corresponding position in the homologous amino acid sequence are identified and assigned a score.
20 . The method of claim 19 , wherein at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, or at least 15, substitutions are made in the reference amino acid sequence at 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions in the reference sequence with an amino acid present at the corresponding 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions in the homologous amino acid sequence where the score assigned to the differences identified at the 2 nd , 3 rd , 4 th , 5 th , 6 th , 7 th , 8 th , 9 th , 10 th , 11 th , 12 th , 13 th , 14 th , 15th or more, positions is intermediate between x and y indicating that the occurrence of substitution has an equal probability.