PTO Agrees to Reexamine Broad Codon-Optimization Patents

Many important applications of biotechnology involve the expression of recombinant genes in heterologous systems. Since the genetic code is degenerate (ie each amino acid can be coded by on average three different codons), the DNA sequence can be modified by synonymous nucleotide substitutions without altering the amino acid sequence of the encoded protein. A biotechnologist might find it useful to make a synonymous substitution for any of a variety of reasons, e.g., to introduce or remove restriction sites, or to remove repeats.

Today genes are also often altered for the specific purpose of optimizing expression, i.e., the entire coding region of the gene is recoded to fit one of a variety of codon optimization schemes. Examples of popular codon optimization schemes used in designing a gene for recombinant expression include:

A) Replace a degenerate codon with the most common codon present in the host chromosome; the rationale being that the most common codon correspond to the most common tRNA results in most protein yield.

B) Reengineer the gene so that it uses the same frequency of codons that are present in the host chromosome; the rationale being that the recombinant gene should have the same codon bias as the host chromosome to fit the translational machinery.

C) Re-code the pattern of rare and common codons found in the wild type by replacing it with the corresponding rare/common codons in the heterologous host; the rationale being that protein folding and expression requires rare (slow) and common (fast) codons distributed according to the folding domains in the protein.

D) Empirically identify codons and other variables that correlate with high protein expression in a systematically varied recombinant gene.

Codon optimization has become standard practice in the recombinant expression of heterologous proteins such as biologic drugs. Gene synthesis companies such as DNA2.0 and Blue Heron assist their customers by providing codon optimization as a part of their services.

Two U.S. patents (both assigned to the Massachusetts General Hospital) include claims directed towards codon optimization, both the method and the resulting codon-optimized gene (US Patent Numbers 5,786,464 and 6,114,148). These patents are very broad, purporting to cover any "synthetic gene encoding a protein normally expressed in an eukaryotic cell wherein at least one non-preferred or less preferred codon . . . has been replaced by a preferred codon encoding the same amino acid, [and wherein the replacement results in at least 10% increase in the level of protein expressed] in an in vitro mammalian cell culture system under identical conditions”

These patents appear to not only cover any successfully codon-optimized recombinant gene, but also many genes that have been reengineered for purposes other than codon optimization. For example, a silent mutation introduced for the purpose of removing a restriction site could easily constitute infringement if it results in the introduction of any of the 17 codons identified in the patent as “preferred,” if it turns out that the substitution results in a modest 10% increase in expression in any in vitro mammalian cell culture system. Rarely would an experimenter test for such an increase in expression at that level of accuracy, and a practical matter it would be virtually impossible to rule out the possibility of a 10% increase in expression in some mammalian cell culture system.

In view of their scope, and the pervasive use of codon replacement in the expression of high-value heterologous proteins (such as biologic drugs), these patents could potentially have sweeping implications for biotechnology. However, although the patents issued in 1998 and 2000, they have never been asserted in court and until recently appear to have received little attention.

However, the patents have recently become a thorny issue for those involved in the business of expressing heterologous proteins, particularly gene synthesis companies providing codon optimized genes for their customers. Last year Geneart (a German gene synthesis company owned by Life Technologies) began sending letters to companies engaged in the expression of heterologous proteins announcing that "we are very pleased to inform you that Geneart has acquired a license under [the Massachusetts General Hospital (MGH) Patents].” The letter goes on to state that under the license “Geneart is in the excellent position to offer a broad range of sublicensing opportunities." The licensing “opportunities” include a royalty-free sublicense to use synthetic genes purchased from Geneart in internal R&D, and a royalty-bearing commercial sublicense for synthetic genes used for purposes other than internal research and development. According to the letter, Geneart has executed a non-prosecution agreement with respect to synthetic genes delivered by Geneart to customers prior to May 30, 2010, so these customers are apparently free to use the genes in any manner without liability for infringement.

The letter goes on to imply that customers who choose to purchase synthetic genes from competing gene synthesis companies could be sued for infringing the MGH Patents, stating that "to Geneart’s knowledge, no other Gene Synthesis Service Provider has obtained a respective license under the MGH Patents."

This poses a problem for any biotechnologist wishing to use a synthetic or partially modified gene. Even slight variations to the wild-type sequence, such as restriction site removal or addition, could infringe.
It certainly poses a problem to other synthetic gene companies, whose customers might switch to Geneart in order to avoid a perceived threat of patent infringement liability for using synthetic genes not purchased from Geneart.


DNA2.0, a leading provider of synthetic genes headquartered in Menlo Park California, has responded to this perceived threat by successfully petitioning for reexamination of both patents, arguing that the broad claims are anticipated and/or rendered obvious by prior art not previously considered by the PTO. The orders granting the ex parte reexamination of the ‘148 and ‘464 patents were issued on December 14, 2010 and January 26, 2011, respectively.

The orders granting reexamination state that a number of references cited by DNA2.0 in the request for reexamination raise substantial new questions of patentability with respect to most of the claims in the patents, particularly the broadest claims. For example, an article published in the Journal of Virology in 1992 by Schwartz et al. describes the characterization of inhibitory RNA elements in the gag region of human immunodeficiency virus type I (HIV-1). This involved replacing many of the codons in the wild type gene with synonymous codons defined as "preferred" in the MGH Patents, resulting in at least a five-fold increase in expression. As another example, the PTO found an article disclosing a computer program for optimizing DNA sequences for protein expression, in combination with another article disclosing the most frequent human codon usage together with preferred codon choice patterns, also raise substantial new questions of patentability for many of the claims.

It is interesting to note that the corresponding codon optimization patent issued to MGH in Europe (EP 0 781 329 B2) is much narrower in scope, claiming a method for preparing a synthetic gene wherein at least 50% of the non-preferred codons and less preferred codons are replaced by preferred codons and resulting in a 10% increase in expression. In contrast, the US patents purport to claim any method that involves replacing even one codon with a preferred codon, and any gene made by such a process.

It is my understanding that, as a practical matter, the European patent only poses a substantial impediment to the “A” form of codon optimization described above, i.e., replacement of all degenerate codons with the most common codon present in the host chromosome. Most other codon optimizations do not require the substitution of 50% of non- or less preferred codons. In contrast, the U.S. patents would appear to cover the majority of recombinant protein expression that has been done in recent years. The patents should expire 2015, but the pending reexamination could be important if it results in cancellation or narrowing of the claims in the US patents, thereby providing assurance that companies will not be sued for expressing recombinant proteins prior to patent expiration.