PepTalk 2017
PepTalk 2017
2014 Archived Content

Cambridge Healthtech Institute’s Sixth Annual
Engineering Genes, Vectors, Constructs and Clones
Upstream Decisions Lead to Downstream Success
January 13-14, 2014

The demand for high-quality biotherapeutic proteins has never been greater. Meeting this goal of producing recombinant proteins at even higher levels requires engineering (science) and insight (art). Bottlenecks arise frequently because functional recombinant proteins are difficult to produce, and protein engineers are often forced to return to the drawing board. This usually requires designing new cloning schemes, including lengthy verification and sequence analysis of the gene or protein of interest, moving a gene from one vector to another, transfecting the vector in an alternative host, re-characterizing the expressed protein or any of the above – an inefficient, time-consuming and expensive process. Cambridge Healthtech Institute’s Sixth Annual Engineering Genes, Vectors, Constructs and Clones meeting continues the tradition of applying effective engineering strategies for protein discovery research that leads to functional biotherapeutic products. Learn from seasoned, savvy researchers as they share their real-world experiences, applications and results.

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4:00-5:00 pm Short Course Registration

5:00-8:00 Dinner Short Courses (SC1-SC7) More Details >> 

4:00-8:00 Main Conference Registration


7:30 am Conference Registration and Morning Coffee

Gene Engineering 

9:00 Chairperson’s Opening Remarks

James L. Hartley, Ph.D., Senior Scientist, Protein Expression Laboratory, Advanced Technology Program, Frederick National Laboratory for Cancer Research

Keynote Presentation

9:10 Next-Generation DNA Synthesis to Optimize Gene Expression

Sriram KosuriSriram Kosuri, Ph.D., Assistant Professor, Chemistry and Biochemistry, University of California, Los Angeles

New technologies for constructing DNA are beginning to revolutionize our understanding and engineering of biology. Here, I will present new large-scale methods for the construction of synthetic gene libraries, and discuss how we apply them to understand gene expression in both bacterial and mammalian systems using multiplexed characterization.

9:50 Tools and Processes to Design, Write and Assemble DNA Sequences
JamesKaysenJames Kaysen, Ph.D., Staff Scientist, Life Technologies 

The newly emerging field of synthetic biology has created an increasing demand for low-cost custom-designed DNA sequences for a variety of applications. This presentation will describe the current state-of-the-art tools, products and services available from Life Technologies to synthesize DNA fragments and then assemble these fragments into larger genes and ultimately complex genetic circuits. These products and services offer an alternative to the time and expense of gene cloning and allow for codon optimization and custom protein design.

10:20 Coffee Break

10:45 High-Throughput Recombinant Antibody Discovery Platform: From Antigen Production to Binder Validation

 Marcin PaduchMarcin Paduch, Ph.D., Technical Director, Recombinant Antibody Network, Biochemistry and Molecular Biology, The University of Chicago

We have developed a powerful recombinant-based strategy for generating customized synthetic antibodies by integrating a set of wholly in vitro techniques into an automated high-throughput pipeline. The task involved designing novel phage display libraries, improved biopanning techniques, optimizing small-scale screening and purification techniques all networked together by modern lab automation systems. Developed recombinant antibody generation process requires efficient production of highly purified proteins and streamlined quality control which will be discussed in more detail.

11:15 Complete Knockout of the Lactate Dehydrogenase-A Gene Is Lethal in Pyruvate Dehydrogenase Kinase 1, 2, 3 Down-Regulated CHO Cells

Yongping CrawfordYongping Crawford, Ph.D., Scientist, Early Stage Cell Culture, Genentech, Inc.

We utilized zinc finger nuclease technology to disrupt the LDHA gene in a PDHK1, 2, 3 knockdown CHO cell line. After screening 3000 clones, only heterozygous mutants were recovered. Second attempt to knockout the remaining wild type allele yielded two mutants that retained LDH activity: one had a 3bp insertion and the other possessed a rare duplication with one wild type and two frameshifted alleles. Our data indicate that complete LDHA knockout is lethal despite PDHK1, 2, 3 down-regulation.

11:45 “Reverse Engineering” of Clone Cell Line Developments for Effective and Efficient Biomanufacturing

Wei ChenWei Chen, Ph.D., Managing Director, BioPharmaneer, Inc.

Conventional cell line development typically starts in a research lab and focuses more on optimal cell growth based on biology principles. As biotech is emerging from R&D to a more mature industry, Engineering Principles for efficient “Mass Production” get critical and optimal product production becomes the goal. Examples of cell lines-coupled upstream designs and downstream outcomes of various biologic products including MAbs, therapeutic proteins and vaccines will be discussed.

VTU Technology GmbH12:15 pm Uniform GlcNAc2Man5-Decorated Proteins by Pichia pastoris: Achievements in High-Level Production and Characterization

Weis_RolandRoland Weis, Ph.D., Head, Operations, VTU Technology GmbH

Heterogeneous N-linked glycan profiles on therapeutic proteins represent a regulatory disadvantage paired with an elevated workload to identify the respective glycan distribution from batch to batch. Next to its extraordinary secretion capacity, the established recombinant production host Pichia pastoris now features homogeneous N-glycan structures (GlcNAc2Man5) for reliable high-level production of demanding proteins. Case studies of the biological implications of these glycans on human serum proteins will be presented.

12:45 Luncheon Presentation (Sponsorship Opportunity Available) or Lunch on Your Own


2:00 Chairperson’s Remarks

Wei Chen, Ph.D., Managing Director, BioPharmaneer, Inc.

2:05 Ventures into Synonymous Codon Space

James L. HartleyJames L. Hartley, Ph.D., Senior Scientist, Protein Expression Laboratory, Advanced Technology Program, Frederick National Laboratory for Cancer Research

There are 61 amino acid codons for 20 amino acids, so it follows that there are about 3 exp 100 = 10 exp 47 different nucleotide sequences for a specific protein comprising 100 amino acids. While this is an impossibly large number to even remotely sample, we have constructed libraries of genes in which the codons for each amino acid are random. I will describe some construction and expression characteristics of these unusual gene libraries.

2:35 Bacterial Chromosomal Engineering for Optimization of Protein Expression and Function

Joseph KittleJoseph Kittle, Jr., Ph.D., Assistant Professor, Chemistry and Biochemistry, Ohio University and CSO, Molecular Technologies Laboratories, LLC

We have used chromosomal engineering to develop advanced bacterial strains and methods for improved expression of therapeutic and research proteins. Modified expression of bacterial host genes alters bacterial metabolism and improves fermentation. Rapid chromosomal engineering methods for generating gene variants facilitate some protein variants with improved function and/or stability. We will document the performance of engineered strains and proteins compared to traditional plasmid-based systems and show how gene variant screening can help solve problems in antibody fragment- based protein expression.

3:05 Molecular Evolution of Human Butyrylcholinesterase

JohnCashmanJohn Cashman, Ph.D., President and Founder, Human BioMolecular
Research Institute

From random cDNA libraries, using a medium throughput screen, molecular evolution of human butyrylcholinesterase was accomplished. Several promising organophosphate detoxication catalysts were identified.

Lucigen logo3:35 New Endotoxin-Free E. coli Cell Strains for Plasmid and Protein Production without Endotoxin Removal 

Knox_CurtisCurtis Knox, Vice President, Marketing & Sales, Lucigen Corp.

Lucigen will present novel competent E. coli cell strains lacking lipopolysaccharide (LPS) for endotoxin-free protein and DNA production. Examples of endotoxin reduction levels achieved with Limulus Amoebocyte Lysate (LAL) and human cell line-based assays will be demonstrated, as well as protein expression and plasmid production experimental data.

3:50 Refreshment Break

4:15 Screening Antibody Phage Libraries in Product Format

Partha Chowdhury, Ph.D., Principal Scientist, Antibody Discovery and Protein Engineering, MedImmune, Inc.

Despite being a powerful technology for antibody discovery, phage libraries are severely limited because they cannot be directly screened in a relevant product format which is typically IgG. This talk will focus on the development and validation of a new technology that enables batch conversion of scFvs from phage libraries and high-throughput screening as IgGs.

Featured Presentation

4:45 Genetic Engineering and Preclinical Testing of Salmonella Live Vaccines

James E. GalenJames E. Galen, Ph.D., Professor, Medicine, Chief, Salmonella Live Vector Vaccine Section, University of Maryland School of Medicine

The genetic engineering of attenuated Salmonella Typhi live vector vaccines, capable of delivering protective antigens from unrelated human pathogens, will be described, with emphasis on the development of live vaccines against plague. The immunogenicity and protective efficacy in mice of our novel vaccines will be also be presented.

5:30 Close of Session

Protein Simple5:45-7:00 Welcome Reception in the Exhibit Hall With Poster Viewing

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