NOTICES 
49605 
nant DNA Molecule Program Advisory Com- 
mittee advisee the Secretary. Department of 
Health. Education, and Welfare; the Assist- 
ant Secretary for Health. Department of 
Health. Education, and Welfare; and the 
Director. National Institutes of Health, on 
a program for the evaluation of potential 
biological and ecological hazards of recom- 
binant DNAs. on the development of pro- 
cedures which are designed to prevent the 
spread of such molecules within human and 
other populations, and on guidelines to be 
followed by investigators working with po- 
tentially hazardous recombinants. 
The NTH Recombinant DNA Molecule Pro- 
gram Advisory Committee has responsibility 
for; (1) Recommending revisions of the 
Ouldellnes to be followed by Investigators 
working with DNA recombinants. (U) re- 
ceiving Information on proposed HV1. HV3 
< EK2 ) , and HV3 i EX3) host- vector systems, 
determining whether the proposed systems 
meet the established criteria, and recom- 
mending to the Director. NTH. the certlflca- 
tion of systems which meet the criteria, and 
(Ul) resolving questions concerning poten- 
tial biohazards and adequacy of contain- 
ment If the NTH Office of Recombinant DNA 
Activities so requests. 
The Committee shall have the authority 
to recommend to the Director. NTH. exemp- 
tions from specific requirements of the 
Ouldellnes. when such exemptions are Jus- 
tified by exceptional circumstances. For ex- 
ample. the committee may recommend ex- 
emptions from the containment require- 
ments of the Ouldellnes for committee-ap- 
proved rlak- assessment studies. Each request 
for such an exemption must be accompanied 
by adequate documentation, which shall In- 
clude clear and convincing evidence of the 
need for the particular risk -assessment 
study The committee may also recommend 
exemptions for specific experiment^ which 
fall within the six categories of "experiments 
that are not to be performed " m making 
such recommendations for exceptions, 
weight will be glyen both to scientific and 
societal benefits and to potential risks. 
E NIH NTH staff has responsibility for: 
(1) Assuring that no NIH grants or contracts 
are awarded for recombinant DNA research 
unless they (a) conform to these Ouldellnes. 
(b) have been properly reviewed and recom- 
mended for approval, and (e) Include a prop- 
erly executed Memorandum of Understand- 
ing and Agreement. (11) reviewing and re- 
sponding to questions or problems or reports 
submitted by Institutional biohazards com- 
mittees or principal Investigators, and dis- 
seminating findings, as appropriate. (Ul) re- 
ceiving and reviewing applications for ap- 
proval to lower containment levels when a 
cloned DNA recombinant derived from a 
shotgun experiment has been rigorously 
characterised and there Is sufficient evidence 
that It Is free of harmful genes. (It) referring 
Items covered under (U) and (111) above to 
the NIH Recombinant DNA Molecule Pro- 
gram Advisory Committee, as deemed neces- 
sary. and (v) performing site Inspections of 
all P* physical containment facilities en- 
gaged In recombinant DNA research, and site 
Inspections at other facilities as deemed 
necessary 
The Director. NTH. Is responsible for over- 
seeing Implementation and for Judicious and 
flexible Interpretation of these Ouldellnes. 
V. Foothotzs 
' In general, recombinant DNA formed be- 
tween segments of eukaryotic viral DNA and 
any eukaryotic DNA (viral, evtrachromo- 
somal) shall not be excluded from these 
Ouldellnes until such time as there Is more 
Information about the extent of naturally 
occurring recomblnatlonal events between 
these DNAs 
• Biological safety cabinets referred to In 
this section are classified as Class I. Class II. 
or Class III cabinets. A Class I is a ventilated 
cabinet for personnel protection having an 
Inward flow of air away from the operator. 
The exhaust air from this cabinet Is filtered 
through a high efficiency or high efficiency 
particular air ( HEP A) filter before being 
discharged to the outside atmosphere. This 
cabinet Is used In three operational modes; 
(1) with an 8-lnch high full width open 
front. (3) with an Installed front closure 
panel (having four 8-lnch diameter open- 
ings) without gloves, and (3) with an In- 
stalled front closure panel equipped with 
arm length rubber gloves. The face velocity 
of the Inward flow of air through the full 
width open front Is 73 feet per minute or 
greater. A Class II cabinet Is a ventilated 
cabinet for personnel and product protec- 
tion having an open front with Inward air 
flow for personnel protection, and HEPA 
filtered mass recirculated air flow for product 
protection. The cabinet exhaust air Is filtered 
through a HEPA filter. The face velocity of 
the Inward flow of air through the full width 
open front is 76 feet per minute or greater. 
Design and performance specifications for 
Class II cabinets have been adopted by the 
National Sanitation Foundation. Ann Arbor. 
Mich. A Class ni cabinet Is a closed front 
ventilated cabinet of gas tight construction 
which provides the highest level of personnel 
protection of all biohazard safety cabinets. 
The Interior of the cabinet Is protected from 
contaminants exterior to the cabinet. The 
cabinet Is fitted with arm length rubber 
gloves and Is operated under a negative pres- 
sure of at least 0.5 Inches water gauge. All 
supply air Is filtered through HEPA filters. 
Exhaust air Is filtered through HEPA filters 
or incinerated before being discharged to the 
outside environment. 
• The terms "characterized" and "free of 
harmful genes" are unavoidably vague. The 
following types of data should be consid- 
ered: (a) The absence of potentially harm- 
ful genes (eg., sequences contained In In- 
digenous tumor viruses or sequences that 
code for toxins. In vaslns. virulence factors, 
etc., that might potentiate the pathogenicity 
or communicability of the vector and or host 
or be detrimental to humans, animals or 
plants); (b) the type(a) of genetic Infor- 
mation on the cloned segment and the nature 
of transcriptional and translation gene prod- 
ucts specified; (c) the relationship between 
the recovered and desired segment (eg., hy- 
bridization and restriction endonuclease 
fragmentation analysts where applicable); 
(d) the genetic stability of the cloned frag- 
ment: and (e). any alterations In the biologi- 
cal properties of the vector and host. 
• We are specifically concerned with po- 
tent toxins which could be produced by 
acquiring a single gene or cluster of genes. 
■The containment requirements specified 
for DNA derived from viruses of eukaryotes 
were designed chiefly with two hypothetical 
risk mechanisms In mind. The first Is the 
concern lest a bacterium carrying viral nu- 
cleic acid be able to produce virus Infec- 
tions; this Is most easily envisioned as oc- 
curring from delivery of the viral nucleic 
acid Into proximity with host cells. (Pro- 
duction of Infectious virus within the host 
bacterium appears to be a most unlikely 
event, except possibly In the case of the 
smaller lcosahedral viruses. However, fur- 
ther data are needed.) Obviously, this mech- 
anism applies only when the complete viral 
genome Is present, and containment condi- 
tions are highest when there Is a possibility 
that the whole genome Is present. 
The second hypothectlcal risk mechanism 
Is focused on viral oncogenesis. The chief 
component of the model Is again delivery 
of the viral nucleic acid to host cells, with 
subsequent Integration of the viral DNA 
Into the genome of h 06 t cells In some DNA 
virus systems, cell transformation can be 
produced by a segment of the viral genome; 
such segments are from the early region 
of the viral genome, and presumably code 
for products which affect regulation of DNA 
synthesis or function. Genes coding for the 
synthesis of virus capsid components have 
not been associated with oncogenesis. Con- 
sequently. containment conditions for sub- 
viral segments from DNA viruses are set 
higher for gene’ regions coding for nonstruc- 
tural proteins, which are chiefly Involved with 
DNA synthesis, than for genes for virion 
structural components. 
With the exception of the Retroviruses 
(l.e., the family of viruses whose members 
possess the reverse transcriptase enzyme, and 
which includes a number of oncogenic viruses 
(Fenner. F.. Classification and Nomenclature 
of Viruses; Intervirology. 7:1-116. 1976.)). 
RNA viruses have not been Implicated In 
tumortgenesls. Further, the genes of RNA 
viruses analogous to those of DNA viruses 
which specify early functions and aonstruc- 
t ural protein synthesis tend to be specific 
for the RNA of that virus, and are not 
thought to affect cellular processes. 
The concept of production by the host 
bacterium of a viral protein which would 
act from outside the cells as a carcinogenic 
substance la without experimental basis. 
Also, there la no Instance of a highly toxic 
viral product Production of disease by a viral 
Infection requires the functioning of the en- 
tire virus; there la no basis for concern that 
a eukaryotic viral DNA segment might cause 
the host bacterium to produce a toxin. 
It should be pointed out that the concern 
about risk from a microorganism carrying 
a viral Insert la directed at those species that 
may come In contact with the host micro- 
organism. and is not particularly directed at 
those species that the virus Infects In nature. 
• The DNA preparation Is defined as "puri- 
fied" If the desired DNA represents at least 
99 percent ( w w) of the total DNA In the 
preparation, provided that It was verified by 
more than one procedure. 
r Baculovtruses which have been registered 
to date Include the Heliothis tea baculovlrus. 
registration number 11373-17 (Jan. 1976) 
and the Orgyia pseudotsugata baculovlrus. 
Registration number 37586-1 (Aug. 1976). 
* The reason for lowering the containment 
level when this degree of purification has 
been obtained Is based on the fact that <he 
total number of clones that must be ex- 
amined to obtain the desired clone Is 
markedly reduced. Thus, the probability of 
cloning a harmful gene could, for example, 
be reduced by more than HP-fold when a 
nonrepetltive gene from mammals was being 
sought. Furthermore, the level of purity 
specified here makes It easier to establish 
that the desired DNA does not contain harm- 
ful genes. 
•In special circumstances. In consultation 
with the NTH Office of Recombinant DNA 
Activities, an area biohazards committee may 
be formed, composed of members from the 
Institution and/or other organizations be- 
yond Its own staff, as an alternative when 
additional expertise outside the Institution 
Is needed for the Indicated reviews. 
VI. Refebxnczs 
1. Laboratory Safety at the Center for Dis- 
ease Control (Sept., 1974). D.S. Department 
of Health. Education and Welfare Publica- 
tion No. CDC 75-8118. 
2. Classification of Etiologic Agents on the 
Basis of Hazard. (4th Edition. July. 1974). 
US. Department of Health, Education and 
Welfare Public Health Service. Center tor 
Disease Control. Office of Biosafety. Atlanta. 
Georgia 30333. 
3. National Cancer Institute Safety Stand- 
ards for Research Involving Oncogenic Vir- 
FEDERAl REGISTER, VOL 42, NO. 117 — TUESDAY, SEPTEMBER 27, 1977 
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