ABSTRACT
Codomax has developed the Epi-MAX platform, an integrated mass spectrometry omics and computational
technology for enhanced protein synthesis. Current protein manufacturing processes typically employ
expression vectors inserted into heterologous host cells, where gene, regulatory and cell features limit protein
production; some proteins are difficult to express in high yields due to inefficiently translated or sub-optimally
transcribed mRNA. Codomax's innovation hinges on the discovery that cells regulate translation in response to
stress by altering the tRNA pools to enhance translation of mRNAs with distinct codon usage patterns. The Epi-
MAX platform has leveraged this tRNA codon-based mechanism to increase recombinant protein expression in
cells by matching codon usage in the target gene to the stress-reprogrammed tRNAs. Previously, Codomax
demonstrated stress-induced tRNA reprogramming in mammalian systems and propose to extend the platform
to antibody production in Chinese hamster ovary (CHO) cells. Underlying antibody production in CHO cells is
the stress of high protein expression, evidenced by Codomax's observed changes in the levels of mRNAs and
proteins specific to stress response, protein folding and metabolic genes when comparing high to low expressing
clones. High antibody expression reprograms protein expression machinery with dramatic changes in ribosome
components and translation factors. Codomax has shown that available levels of the CREB1 transcription factor,
which drives target antibody expression, decrease in high expressing clones. Thus, the specific aims for
Codomax's project include application of the Epi-MAX platform to analyze high and low productivity CHO clones
for the model therapeutic antibody to identify tRNA systems that limit heavy chain production, and then identify
tRNAs that when over-expressed, boost antibody expression (Aim 1). Codomax will then increase target heavy
and light chain mRNA transcription 200%, by re-engineering the CMV-regulating CREB1 transcription factor to
have increased stress-induced translation (Aim 2). Codon optimized CREB1 will be used to enhance mRNA
translation during the stress of antibody expression, with maximized CREB1 transcription factor levels driving
increased mRNA transcription from the CMV promoter. The CREB1 gene will be re-engineered using Epi-MAX
data from Aim 1. Then CRISPR-Cas9 approaches will be used to replace the native CREB1 gene with the
CREB1-codon optimized sequence and mRNA expression will be evaluated for the heavy and light chains under
high expression conditions. Aim 2 will result in the generation of gene sequences, plasmids and CRISPR
reagents for CMV-based transcription in several CHO lines. The successful completion of Codomax's Phase I
project will demonstrate proof-of-concept that the Epi-MAX platform can be used to increase antibody yield from
CHO cells and generate designer reagents that promote boosted antibody production.
