Microbial natural products, a key source of blockbuster drugs and new drug leads, are highly complex molecules, each evolved for specific function and fine-tuned to interact with and modulate an array of central cellular targets. Within this context, the natural products produced by microbes found in the environment have led to major therapeutic breakthroughs for a variety of conditions ranging from infectious disease to cancer. A key to reducing the rate of rediscovery, and thus improving the chances of finding new microbial natural products, is exploration in untapped or previously unexplored environments.
The University of Kentucky’s Natural Products Discovery Program, part of the UK College of Pharmacy’s Center for Pharmaceutical Research and Innovation (CPRI), is exploring untapped environments in our own back yard.
Appalachia, as one of the largest contiguous biodiversity “hot spots” in the nation, is expected to serve as fertile ground for the discovery of new microbes.
Specifically, Kentucky’s mines provide unprecedented access to biodiverse environments, particularly unique locations that may present access to unexplored terrain containing bacteria or microbes that have yet to be discovered. Subterranean environments represent the ocean beds of many centuries ago, where microbial spores have also been exposed to dramatic extremes in salinity, heavy metal exposure, temperature, and pressure.
In short, microbes from such unexplored environments are expected to be unprecedented and thereby present a greater chance for the discovery of new natural products.
To access unique environments across the state, CPRI initially teamed up with the UK’s Center for Applied Energy Research. This connection helped establish additional collaborations with the Kentucky Division of Abandoned Mines, the Kentucky Geological Survey, and a number of mining operators across the commonwealth.
These cumulative relationships provide access to a wide array of unusual environments from the microbial natural products discovery perspective, including underground and surface coal mines, thermal vents from underground coal mine fires, mining reclamation sites, and deep-well core drilling operations.
Collaboration is also key to determining the function and therapeutic relevance of the natural products discovered. The pure natural products isolated as a result of UK’s program are cataloged and stored in CPRI’s Natural Products Repository as a unique resource.
Within this context, CPRI also works with UK investigators to develop and validate bioactivity assays relevant to their scientific or therapeutic areas of interest and subsequently employs these assays to screen the repository for compounds that display the desired activity. CPRI also has established strategic external collaborations, including with the National Cancer Institute (NCI).
This early collaborative work sets the stage for downstream studies. Should the compounds display sufficient activity with tolerable toxicity or perceived therapeutic index, potential preclinical development studies could be pursued.
Natural Product Discovery Process
Once soil samples are collected, the CPRI team extracts bacteria and bacterial spores from a small amount of each soil sample and deposits portions of these extracts on media plates to begin the painstaking process of purifying and growing each individual strain of bacteria. Each recovered strain is phenotypically and genotypically characterized and then grown in liquid media to prepare extracts for metabolomic analysis. Strains are prioritized based on their potential for novel metabolite production with a bias toward bacteria genera/species underrepresented in natural products discovery.
High-priority strains are then grown on a large scale from which all accessible metabolites are purified via a range of selective extractions and chromatographic techniques, and then fully characterized structurally using mass spectrometry, nuclear magnetic resonance, and other spectrophotometric methods.