The previous entry in this blog described U.S. Environmental Protection Agency (EPA) regulations under the Toxic Substances Control Act (TSCA) that may cover the use of certain genetically modified microorganisms in biofuel production. In this posting I’ll go into a little more detail about these regulations and discuss how companies can successfully manage possible EPA oversight under these rules.
As described in the previous entry, the TSCA Biotech Rule covers only certain microorganisms, those containing deliberate combinations of coding nucleic acids derived from more than one taxonomic genus, when used for commercial purposes not governed by other federal agencies. The use of microorganisms or algae to produce ethanol, butanol, biodiesel or other biofuels is an industrial application that would almost certainly fall under TSCA jurisdiction in most cases. For those modified microorganisms falling under the rules, applicants must notify EPA at least 90 days before any proposed commercial use of the strain, using a procedure called the Microbial Commercial Activity Notice (MCAN). However, the applicability of the regulations to any modified strain will depend on the make-up of the microorganism in question. Specifically, some genetically modified microbes will fall completely outside the regulations and will thus not be subject to these provisions at all, while others may be subject to the rule but may qualify for one of the exemptions the regulations provide. So it is important to first assess, either on one’s own or in consultation with EPA staff, the status of the production microorganism under these regulations.
(Please note that presubmission consultation with Agency staff is always recommended and is often quite helpful, and this will be discussed below. In addition, the discussion in this blog is of a general nature and should not be construed as legal or business advice, and each potential applicant should discuss your situation with a knowledgeable consultant or attorney to make determinations of how these regulations would apply to your particular circumstance).
Companies planning use of modified microorganisms for biofuel production should begin by considering the following questions.
1. Is the proposed industrial use one that is subject to TSCA jurisdiction? For biofuel organisms, the answer to this question will most likely be “yes”, but this should be the threshold question, because microorganisms used for a purpose regulated by other federal agencies like the FDA or EPA’s pesticide branch would not be subject to the TSCA Biotech Rule. However, grey areas could arise if the microorganism was also intended for industrial uses other than biofuel production.
2. Does the microorganism fall within the scope of the regulations? The Biotech Rule defines a “new microorganism” as an “intergeneric microorganism”, which is defined to mean “a microorganism that is formed by the deliberate combination of genetic material originally isolated from organisms of different taxonomic genera”. Not all genetically engineered microorganisms will meet the definition of “new microorganism”, and those that do not will not be subject to regulation under the TSCA Biotech Rule. The most obvious class of microbes that may fall outside the scope of the regulations are those where the artificial modifications result only in the deletion of genes native to the host organism, or the enhancement of function or activity of genes and gene products naturally found in that host. Most such organisms would not meet the definition of “new organism” because they would contain no coding DNA of any genus other than that of the host organism, and such microbes would therefore not at all be subject to the regulations. However, care must be taken to analyze whether any foreign genetic elements may be retained in the final organism as a result of the genetic manipulations, for example, “scars” resulting from the use of the cre-lox system to delete endogenous genes. However, the presence of such non-coding DNA would likely not itself cause such a microorganism to be subject to the rule. One should also not forget that the presence of a marker gene, such as one encoding antibiotic resistance, could subject the organism to the regulations, if the origin of the coding sequences of the marker gene was from a genus other than the genus of the host organism. (Note that the presence of an antibiotic resistance marker is not itself a barrier to commercial use of a genetically modified strain, although it may possibly lead to some additional issues in EPA’s review, for those microbes covered by the rule).
3. Does the microorganism qualify for an exemption under the regulations? For those microbes that are judged to include coding DNA from different genera and are thus considered “intergeneric”, the next analysis would be whether the modified strain might qualify for one of the exemptions under the regulations. The Biotech Rule provides exemptions from EPA reporting for specific organisms that meet the qualifications established in the rule. The initial determination of whether an exemption might be available is based solely on the biology and make-up of the modified commercial microorganism, and can be made relatively easily by the applicant alone or in consultation with EPA. But as discussed below, it is important to remember that for neither of the two tiers of exemptions is the applicant completely removed from EPA oversight, and in fact the availability of one of the exemptions requires stricter adherence to certain procedural controls than the other exemption or even for microbes subject to full EPA notification and review.
What I call the “biological criteria” for exemption consists of two prongs. First, the host, or recipient, organism must be one that is included on the following list, found in Section 725.420 of the regulations, that includes many well-studied species with long histories of use for industrial purposes.
(a) Acetobacter aceti
(b) Aspergillus niger
(c) Aspergillus oryzae
(d) Bacillus licheniformis
(e) Bacillus subtilis
(f) Clostridium acetobutylicum
(g) Escherichia coli K-12
(h) Penicillium roqueforti
(i) Saccharomyces cerevisiae
(j) Saccharomyces uvarum
Second, as specified in Section 725.421, the introduced genetic material in the microorganism must be well characterized (that is, the function of all introduced DNA is known); must be limited in size to the minimal genetic information needed; must be poorly mobilizable, that is, that the ability of the genetic material to be transferred and mobilized has been inactivated and that the frequency of transfer is less than 10-8 transfer events per recipient; and must be known to be free of harmful sequences. meet several criteria. Given the precision possible in today’s recombinant DNA techniques, these conditions should be easy to meet. Again, for applicants intending to use a modified strain of one of these listed host organisms, it should be easy to ascertain whether these “biological criteria” for the exemption have been met, although consultation with EPA will often be desired.
To qualify for one of the exemptions, organisms meeting the first two sets of criteria (the “biological criteria”) must be used in accordance with the guidance of Section 725.422 of the regulations, which specifies specific containment and control procedures to minimize the possibility that the engineered microorganism might inadvertently be released from the facility. If an organism meets the biological criteria, and the applicant can use the microbe in strict compliance with these Section 422 procedures, the process is eligible for a “Tier I” exemption and can be used commercially merely upon 10 days advance notice to EPA. For microbes meeting the biological criteria but which are intended for use under conditions less strict than the Section 422 procedures, the applicant can submit a petition for a “Tier II” exemption 45 days before intended manufacture. EPA would approve the Tier II request if it felt that the proposed containment and control procedures, although not identical to the Section 422 procedures, were sufficient for the organism in question. Note that the Section 422 procedures are also recommended for use with microorganisms subject to MCAN reporting.
The Section 422 procedures and containment conditions can be found within the regulations on the EPA website. It would go beyond the scope of this blog entry to describe them in detail, but they pertain to the use of commonly-accepted procedures to inactivate the microorganisms after their commercial utility is completed and to otherwise minimize the chance that living organisms can escape the fermentation vessel or a closed production system; and to procedures for ensuring that all workers take appropriate precautions to avoid being exposed to large quantities of living microorganisms; that access to the facility is controlled, and to put in place appropriate emergency procedures.
At this writing it does not appear that any microorganism modified for a biofuel-related purpose has made use of the Tier I or Tier II exemptions; in fact it seems that no applicant has taken advantage of these exemptions for any microbe for any purpose. This may be because when proposing use of a modified microbe in an MCAN, the applicant has leeway to justify to EPA’s satisfaction that procedures differing from (i.e., less stringent than) the Section 422 procedures are adequate for the potential risk of the microorganism, whereas under a Tier I exemption the Section 422 procedures must be followed to the letter.
4. Is the microorganism intended to be used for a commercial purpose? As noted in the previous entry, many R&D uses of new microorganisms are exempt from MCAN reporting. A use of a new microorganism would qualify for the “small quantities” exemption under TSCA if it is intended to be used solely for purposes of research and development and is used in a suitably contained structure. The rule specifically contemplates that this exemption would apply broadly to many types of structures, including greenhouses, fermenters and process stream bioreactors, and applicants are given fairly broad leeway in designing and operating facilities to meet the “contained structure” definition. Under this definition, it is likely that most laboratory research in biofuels would be exempt from commercial reporting. In addition, many uses of engineered microorganisms in biofuel pilot or demonstration plants could also qualify for this exemption, as long as the microbe were used “solely” for R&D purposes. This is a question many companies may need to consider, depending on the disposition of the fuel and other byproducts that may arise from use of the process at pilot or demonstration scale.
MCAN Reporting Requirements. If, after asking these questions, the proposed use of a modified microorganism meeting the “new microorganism” definition that is to be used for a commercial purpose subject to TSCA’s jurisdiction, and which does not qualify for an exemption, would likely require the filing of a Microbial Commercial Activity Notification (MCAN) 90 days in advance of the commercial activity. The information and other data that applicants need to submit in the MCAN are listed in Section 725.155 of the regulations, Much of the required information has to do with the biological characterization of the modified microorganism and a detailed description of how it was constructed, but information is also to be submitted on the proposed use of the microbe, the proposed production process, the containment and control procedures to be used, the likelihood for worker exposure and the steps taken to control exposure, and an assessment of the potential environmental effects of the microorganism should it be released from the facility. It is important to note that, in MCANs or other submissions to EPA under the Biotech Rule, the applicant can claim much or all of the submitted information as “confidential business information”, which the agency must keep confidential and which cannot be released to the public, but the applicant must provide EPA with the justification for the confidentiality claim (in fact, this justification must be included within the MCAN filing). EPA has published a detailed “Points to Consider” document summarizing the required data and the format for submission, which, together with guidance from the publicly-available versions of previously-filed MCANs (i.e. the parts of prior MCANs not claimed by the applicant as confidential), can be used to help applicants prepare MCAN submissions. (Note that EPA has recently issued new regulations for electronic submission of chemical premanufacture notices that is also applicable to MCAN submissions. This requirement is being phased in from April 2010 to April 2011).
EPA review of MCANs can be expected to be fairly straightforward, and will focus on the potential risks and benefits of the commercial use of the modified microorganism. Most of EPA’s prior reviews of MCANs have taken place within the 90-day period specified in the regulations, although EPA has the power to unilaterally extend the review period by an additional 90 days, or to ask the applicant to voluntarily suspend the review period, if the Agency decides it needs more time. MCANs for the “contained” use of new microorganisms in bio-based manufacturing have generally not caused any concerns or significant issues in EPA’s review, and most have been routinely cleared for commercial use without any delays or difficulties. MCANs (like chemical PMNs) are not “approved” per se but if no issues emerge they are cleared for commercialization by EPA taking no action against the application. The applicant must then file a Notice of Commencement within thirty days of beginning commercial use or importation of the microorganism, a notice that requires submission only of minimal information.
Planning an MCAN or other submission. As is the case with almost all interactions with regulatory agencies, applicants are strongly advised to consult with the appropriate EPA staff well in advance of making any formal submission. In my experience, the EPA TSCA biotech group is happy to meet with and provide guidance to any company or institution that is contemplating a submission under the regulations. The purpose of these meetings is usually for the applicant to briefly introduce the agency to the technology and the make-up of the organism(s) that would be subject to the submission, describe the proposed use, and begin a dialogue about the kinds of data EPA would want to see in support of the application, as well as to obtain the agency’s input on possible applicability of exemptions and/or the type of application that best suits the circumstances. EPA can usually conduct presubmission meetings under confidential conditions, so that not only the substance of the meetings but also the fact that the meetings are taking place can be protected as confidential and would not be releasable to the public under the Freedom of Information Act.
Potential applicants should start the process well in advance of the proposed commercial start date. Speaking very generally, a good time to approach the agency would be at least 6 months, perhaps 6-9 months, before the anticipated start of commercial activity. After first setting up and conducting a meeting with agency staff, it should then require perhaps 2-3 months after the meeting to develop the needed data (in consultation with EPA), and to draft and finalize the MCAN. Naturally, it would be desirable to file the MCAN farther in advance of the intended commercial start date than the 90 days specified in the regulations, because of the small chance that the review period could be extended or suspended if more data needs to be submitted. Such EPA action is rare, particularly under the biotech regulations, but it would seem wise to give oneself at least 1-2 months leeway before the start date to allow for unexpected delays.
The following is a summary of the activities that would likely be carried out in the preparation of any needed EPA submission.
- Contact EPA staff to arrange a presubmission meeting.
- Meet with EPA, discuss and obtain answers to any specific questions that may arise (it may take several weeks or even longer for EPA to answer any specific requests in writing).
- Begin to assemble the needed information and data, including biological information on the strain, description of the process and containment procedures, and any available information or data on possible environmental effects of the microorganism.
- Allow time for several iterations of internal review of the draft MCAN, and for outside legal review, if so desired.
- Allow time to compile and finalize all the information into the final MCAN, to prepare the information required to justify confidentiality claims (additional time may be needed to come up to speed on the new electronic submission procedures as well).
EPA has been receiving PMNs and other notifications of biotechnology products under its interim TSCA policy since 1987 and under the current rules since 1997. Most of the notifications received have been for contained applications: uses of intergeneric microorganisms for manufacturing products for commercial purposes not regulated by other federal agencies, primarily including industrial enzymes . Several of these enzyme products appear to have applicability to the production of cellulosic ethanol or other biofuels. In recent years, EPA has received at least the following three MCANs for genetically modified production organisms for use in cellulosic ethanol production:
- MCAN No. J08-0001, received 05/14/08 from Verenium: Escherichia coli strain BD26981.
- MCAN No. J08-0002, received 05/14/08 from Verenium: Klebsiella oxytoca strain BD26985.
- MCAN No. J09-0003, received 09/02/09 from DuPont: Zymomonas mobilis strain ZW801.
I will review these MCAN submissions in a future entry in the blog, in which I’ll also more specifically discuss how biofuels programs may be affected by the TSCA biotech rule, including the possible applicability of the rule in ways perhaps not originally contemplated by the drafters of the regulation.
D. Glass Associates, Inc. is a consulting company specializing in several fields of biotechnology. David Glass, Ph.D. is a veteran of nearly thirty years in the biotech industry, with expertise in industrial biotechnology regulatory affairs, patents, technology licensing, and market and technology assessments. This blog provides back-up and expanded content to complement a presentation Dr. Glass made at the EUEC 2010 conference on February 2, 2010 entitled “Prospects for the Use of Genetic Engineering in Biofuel Production.” The slides from that presentation, along with more information on D. Glass Associates’ regulatory affairs consulting capabilities, are available at www.slideshare.net/djglass99 or at www.dglassassociates.com.