Method and system for optimizing minigenes and peptides encoded thereby

A computerized method for designing a multi-epitope construct having multiple epitopes, the method comprising the steps of:

• storing a plurality of input parameters in a memory of a computer system, said input parameters comprising a plurality of epitopes, at least one motif for identifying junctional epitopes, a plurality of amino acid insertions, a C+1 enhancement weight value for each insertion, a N−1 enhancement weight value for each insertion, and a maximum number of insertions (MaxInsertions);
• generating a list of epitope pairs from said plurality of epitopes;
• for each combination of insertions for each epitope pair, wherein the number of insertions is in the range of 0 to MaxInsertions, calculating a function value (F) using the equation F=(C+N)/J, when J>0, and F=2(C+N), when J=0, wherein C equals a C+1 enhancement weight value of a respective flanking amino acid insertion, N equals a N−1 enhancement weight value of a respective N−1 flanking amino acid insertion, and J equals the number of junctional epitopes detected for each respective combination of insertions in an epitope pair based on said at least one motif;
• determining for each epitope pair at least one optimal combination of insertions yielding a maximum function value F;
• generating a list of optimal combinations of insertions;
• and
• based on said list of optimal combinations of insertions, identifying at least one optimum permutation of said multi-epitope construct comprising said plurality of epitopes arranged in an order that yields a maximum sum of function values, wherein a respective one of said optimal combinations of insertions are inserted at a respective junction of two epitopes of said multi-epitope construct.

A computer system for designing a multi-epitope construct having multiple epitopes, the system comprising:

• a memory for storing a plurality of input parameters, said input parameters comprising a plurality of epitopes, at least one motif for identifying junctional epitopes, a plurality of amino acid insertions and at least one enhancement weight value for each insertion;
• a processor for retrieving said input parameters from said memory and generating a list of epitope pairs from said plurality of epitopes;
• said processor further determining for each of said epitope pairs at least one optimum combination of amino acid insertions, based on said at least one motif, said plurality of insertions and said at least one enhancement weight value for each insertion;
• said processor further identifying at least one optimum arrangement of said plurality of epitopes, wherein a respective one of said optimum combinations of amino acid insertions are inserted at a respective junction of two epitopes, so as to provide an optimized multi-epitope construct;
• and
• a display monitor, coupled to said processor, for displaying said at least one optimum arrangement of said plurality of epitopes to a user.

A computer system for designing an optimized multi-epitope construct having multiple epitopes, the system comprising:

• an input device for inputting a plurality of input parameters specified by a user;
• a memory, coupled to the input device, for storing said plurality of input parameters, said input parameters comprising a plurality of epitopes, at least one motif for identifying junctional epitopes, a plurality of amino acid insertions, a C+1 enhancement weight value for each insertion, a N−1 enhancement weight value for each insertion, and a maximum number of insertions (Maxinsertions);
• a processor for retrieving said input parameters from said memory and generating a list of epitope pairs from said plurality of epitopes;
• wherein said processor, for each combination of insertions for each epitope pair, wherein the number of insertions is in the range of 0 to Maxinsertions, calculates a function value (F) using the equation F=(C+N)/J, when J>0, and F=2(C+N), when J=0, wherein C equals a C+1 enhancement weight value of a respective flanking amino acid insertion, N equals a N−1 enhancement weight value of a respective N−1 flanking amino acid insertion, and J equals the number of junctional epitopes detected for each respective combination of insertions in an epitope pair based on said at least one motif;
• and
• wherein said processor further determines for each epitope pair at least one optimal combination of insertions yielding a maximum function value F, generates a list of optimal combinations of insertions, and, based on said list of optimal combinations of insertions, identifies at least one optimum permutation of said multi-epitope construct comprising said plurality of epitopes arranged in an order that yields a maximum sum of function values, wherein a respective one of said optimal combinations of insertions are inserted at a respective junction of two epitopes of said optimized multi-epitope construct.

A data storage device storing a computer program for designing a multi-epitope construct having multiple epitopes, the computer program, when executed by a computer system, performing a process comprising the steps of:

• retrieving a plurality of input parameters from a memory of a computer system, said input parameters comprising a plurality of epitopes, at least one motif for identifying junctional epitopes, a plurality of amino acid insertions and at least one enhancement weight value for each insertion;
• generating a list of epitope pairs from said plurality of epitopes;
• determining for each of said epitope pairs at least one optimum combination of amino acid insertions based on said at least one motif, said plurality of insertions and said at least one enhancement weight value for each insertion;
• and
• identifying at least one optimum arrangement of said plurality of epitopes, wherein a respective one of said at least one optimum combination of amino acid insertions is inserted at a respective junction of two epitopes, so as to provide an optimized multi-epitope construct.

An apparatus for designing a multi-epitope construct having multiple epitopes, comprising:

• means for storing a plurality of input parameters in a memory of a computer system, said input parameters comprising a plurality of epitopes, at least one motif for identifying junctional epitopes, a plurality of amino acid insertions and at least one enhancement weight value for each insertion;
• means for generating a list of epitope pairs from said plurality of epitopes;
• means for determining for each of said epitope pairs at least one optimum combination of amino acid insertions based on said at least one motif, said plurality of insertions and said at least one enhancement weight value for each insertion;
• and
• means for identifying at least one optimum arrangement of said plurality of epitopes, wherein a respective one of said at least one optimum combination of amino acid insertions is inserted at a respective junction of two epitopes, so as to provide an optimized multi-epitope construct.

A method for designing a multi-epitope construct that comprises two or more CTL epitope nucleic acids wherein the construct is presented to an HLA Class I processing pathway, the method comprising steps of:

• (i) sorting the CTL epitope nucleic acids to minimize the number of junctional epitopes;
• (ii) introducing a flanking amino acid residue selected from the group consisting of K, R, N, Q, G, A, S, C, and T at a C+1 position of a CTL epitope nucleic acids;
• (iii) introducing one or more amino acid spacer residues between two epitope nucleic acids, wherein the spacer prevents the occurrence of a CTL or HTL junctional epitope;
• and,
• (iv) selecting one or more multi-epitope constructs that have a minimal number of junctional epitopes, a minimal number of amino acid spacer residues, and a maximum number of K, R, N, G,A, S., C, or T at a C+1 position relative to each CTL epitope nucleic acids.

A method for designing a multi-epitope construct that comprises two or more HTL epitope nucleic acids wherein the construct is presented to an HLA Class II processing pathway, the method comprising steps of:

• (i) sorting said epitope nucleic acids to minimize the number of junctional epitopes;
• (ii) introducing a flanking amino acid residue selected from the group consisting of G, P, N or A positioned between said nucleic acid epitopes;
• and
• (iii) introducing one or more amino acid spacer residues between two epitope nucleic acids, wherein the spacer prevents the occurrence of a HTL junctional epitope.

A multi-epitope construct comprising a plurality of CTL epitope nucleic acids and a plurality of spacer nucleic acids, wherein:

• the CTL epitope nucleic acids encode class I HLA epitopes;
• the CTL epitope nucleic acids encode epitope peptides of about eight to about thirteen amino acids in length;
• the spacer nucleic acids are positioned between the CTL epitope nucleic acids;
• the spacer nucleic acids encode between one and eight amino acids;
• one or more of the spacer nucleic acids encodes an amino acid sequence that is different than the amino acid sequence encoded by other spacer nucleic acids;
• and
• each of the spacer nucleic acids optimizes epitope processing and minimizes junctional epitopes.

A multi-epitope construct comprising a plurality of HTL epitope nucleic acids and a plurality of spacer nucleic acids, wherein:

• the HTL epitope nucleic acids encode class II HLA epitopes of about seven to about seventeen amino acids in length;
• the spacer nucleic acids are positioned between the HTL epitope nucleic acids;
• and
• the spacer nucleic acids encode five or more amino acids, wherein each of the spacer nucleic acids optimizes epitope processing and minimizes junctional epitopes.

A multi-epitope construct comprising fifteen or more epitope nucleic acids and ten or more spacer nucleic acids, wherein:

• the epitope nucleic acids encode class I HLA epitopes or class II HLA epitopes;
• the epitope nucleic acids encode epitope peptides of about seven to about seventeen amino acids in length;
• the spacer nucleic acids are positioned between the epitope nucleic acids;
• and
• the spacer nucleic acids encode between one and eight amino acids when inserted between the class I HLA epitope nucleic acids and five or more amino acids when inserted between the class II HLA epitope nucleic acids.