In Silico Dreams by Brian S. Hilbush

Author:Brian S. Hilbush [Hilbush, Brian]
Language: eng
Format: epub
ISBN: 9781119745631
Publisher: Wiley
Published: 2021-08-31T00:00:00+00:00

Genetics, Gene Discovery, and Drugs for Rare Human Diseases

Molecular biology continued to expand the recombinant DNA toolkit with technical advances to analyze nucleic acid sequences in the late 1970s. The pioneers of the molecular biology era were at work developing two distinct approaches to “read” a DNA sequence. Fred Sanger, who had earlier won a Nobel prize in Chemistry for techniques to sequence proteins, developed what was ultimately to become the mainstay procedure. His ingenious approach was to use chain terminating inhibitors of each of the four DNA bases, such that DNA strands could be synthesized by DNA polymerase reading off of its template until it incorporated a nucleotide with a dideoxy base, which prevented chemical linkage of the growing DNA copy. Images in the popular media today of X-ray films contain the archetypal DNA banding pattern are products of Sanger reactions (and iconography of antiquated technology). Allan Maxam and Walter Gilbert developed an alternative DNA sequencing method based on chemical modification of DNA in one step and subsequent cleavage at specific bases in a second step. Maxam-Gilbert sequencing had an initial accuracy edge over the Sanger method and the advantage of allowing purified samples of DNA to be used without further cloning. However, this was a technically difficult method and laboratories eventually favored Sanger sequencing after refinements to dideoxy sequencing had been implemented in the early 1980s. The development of DNA sequencing technology was a gateway to retrieving and recording the information in genomes. What remained was the automation of these arduous laboratory processes to enable widespread use for basic research, diagnostics, and therapeutic development.

Although decades away from gene-based therapies, advocates for rare diseases together with geneticists were about to launch monumental disease gene discovery projects that could shed light on paths to new treatments. The mapping of disease genes began in parallel with DNA sequencing technology and utilized the tools of genetics—linkage analysis and fine mapping in large, well-characterized pedigrees. By tracking the inheritance of DNA markers that traveled along with affected individuals of a particular disease, scientists could begin the hunt for a chromosomal location and eventually determine the DNA sequence of the disease-causing allele. Establishing that the underlying genetic variants were causal required evidence that the risk genotype was absent in healthy members among unrelated pedigrees with a familial history of disease. More laborious work was needed to determine the functional consequences of the genetic mutations, relying on animal models and biochemistry that tended to grind on for years of investigation to reach an answer—or remain a mystery. Huntington's disease exemplified the latter.

Nancy Wexler at the Hereditary Disease Foundation and Columbia University in New York led a large, decade-long collaborative effort that culminated in the discovery of the Huntington's disease gene on chromosome 4. The journey began with Wexler's heroic efforts to obtain pedigree information and blood samples for DNA marker analysis from an extensive set of families in Venezuela living around Lake Maracaibo, all of whom descended from a woman with the disease living in the 19th century.

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