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###### Department of Health and Human Services

###### National Institutes of Health

# AGENCY:

National Institutes of Health, Public Health Service, DHHS.

# ACTION:

Notice.

# SUMMARY:

The inventions listed below are owned by agencies of the U.S. Government and are available for licensing in the U.S. in accordance with 35 U.S.C. 207 to achieve expeditious commercialization of results of federally-funded research and development. Foreign patent applications are filed on selected inventions to extend market coverage for companies and may also be available for licensing.

# ADDRESSES:

Licensing information and copies of the U.S. patent applications listed below may be obtained by writing to the indicated licensing contact at the Office of Technology Transfer, National Institutes of Health, 6011 Executive Boulevard, Suite 325, Rockville, Maryland 20852-3804; telephone: 301/496-7057; fax: 301/402-0220. A signed Confidential Disclosure Agreement will be required to receive copies of the patent applications.

## Enhanced Homologous Recombination Mediated by Lambda Recombination Proteins

Drs. E. Lee, N. Copeland, N. Jenkins, and D. Court (NCI)

DHHS Reference No. E-077-01/0 filed Feb 26, 2001

Licensing Contact: John Rambosek; 301/496-7056 ext. 270; e-mail: rambosej@od.nih.gov

The present invention concerns a method to enhance homologous recombination in bacteria using the Red recombination system derived from a defective lambda prophage. This lambda system, like the RecET system, uses homologous recombination proteins to protect and recombine the electroporated linear DNA. However, the lambda system is at least 50 to 100 times more efficient than the RecET system. The high recombination efficiency offered by this system makes it possible to manipulate DNA without drug selection. Point mutations, deletions, or insertions can be engineered into any gene on plasmids or bacterial artificial chromosomes (BACs) for gene functional analysis. This recombination system also can be used to subclone DNA fragments as large as 80 kb from BACs by gap repair. Targeting vectors for embryonic stem cells or transgenic constructs by BAC engineering can now be subcloned with ease, and virtually any region of the engineered BAC may be included in the final subclone. The ability to efficiently and precisely modify genes or regulatory sequences on BACs, combined with the ability to include or exclude them during the subcloning process, should make it possible to dissect the function of these sequences in the whole animal at a high-through-put level not previously possible.

This lambda recombination system has been used to introduce a Cre recombinase gene into the coding region of the mouse neural-specific enolase gene carried on a 250 kb mouse BAC after transfer of the mouse BAC into DY380 E. coli cells which carry the lambda recombination system. Transgenic mice that were subsequently generated which carry this modified BAC specifically expressed Cre in all mature neurons and Cre expression mirrored that of the mouse neural-specific enolase gene.

                    This abstract modifies an abstract for this technology published in the                     **Federal Register**                     on Thursday, April 5, 2001 (66 FR 18098).                

## Use of Endogenous Vertebrate Phytase to Increase Capacity To Utilize Phytic Acid in Livestock Feed

Stephen Shears (NIEHS), Paul Reynolds, Jim Petitte

DHHS Reference No. E-139-00/0 filed Aug 11, 2000

Licensing Contact: John Rambosek; 301/496-7056 ext. 270; e-mail: rambosej@od.nih.gov

This invention discloses the concept of creating transgenic farm animals that secrete a native phytase enzyme into their digestive tracts. It has long been recognized that monogastric animals (e.g. pigs and chickens) do not utilize dietary phosphorus as efficiently as possible. This is because a high percentage of total phosphorus (70% in cereals, 50% in legume seeds) is present as phytic acid and its salts—phytate. Monogastric animals utilize phytate inefficiently because they lack the enzyme phytase in their digestive systems. Phytase liberates the phosphorus from phytate, thereby making dietary phosphorus available to the animals. This has the dual effect of both promoting more efficient growth of the animals, as well as imposing less of an environmental burden in the form of excess phosphorus in water streams.

Use of phytase as a growth feed supplement is well known. However, in the past the focus has always been on adding exogenous phytase to animal feed, or to increase the level of phytase expression in the seeds making up the feed. The inventors' novel concept is to redirect expression of a naturally occurring phytase gene so that the enzyme will be secreted into the intestinal lumen. This will create farm animals that can more efficiently utilize unsupplemented feeds. Another problem with existing phytases that the present invention overcomes is that phytase tends to be unstable during the heat treatment used to process feed. This invention overcomes this limitation because the phytase does not have to incorporated into feed at all.

## Cloning of the Human Nuclear Receptor Co-Repressor Gene

Dr. Johnson M. Liu (NHLBI)

DHHS Reference Number E-088-99/0 filed Aug 3, 1999

Licensing Contact: John Rambosek; 301/496-7056 ext. 270; e-mail: rambosej@od.nih.gov

Alteration in the expression of human genes is critical to the development and progression of many diseases. These include, among others, cancer, inflammation, cardiovascular disease, hypercholesterolemia, blood pressure, and diabetes. The Human Nuclear Receptor Co-Repressor (HuN-Cor) gene represents a technology that may be used to alter the transcription of genes. It provides a general mechanism by which many genes may be modulated throughout the entire range of being turned on to being completely turned off. The HuN-Cor gene encodes for a ubiquitously expressed protein that silences other genes. It does this by specifically recruiting an enzyme complex that causes local folding of chromatin, not allowing other transcription factors to access the DNA. HuN-Cor represents a powerful research tool that can be used to study gene expression and characterization of many different genes. It may ultimately have great utility in controlling gene expression via gene therapy technology, and may also be useful as a target for the isolation of pharmaceutical compounds that enhance or inhibit expression of genes. For example, it may be possible to engineer mutations of the HuN-Cor gene that dominantly inhibit its function; these mutants could then be expressed in appropriate target tissues or cells in order to control gene expression. Finally, the gene product may have utility in the discovery of therapeutic compounds that modulate gene expression via HuN-Cor.

## Antibodies That Selectively Detect the Human Nestin Protein

                    Conrad Messam et al. (NINDS)                          ( print page 21406)    

DHHS Reference Nos. E-145-99/0 and E-009-01/0

Licensing Contact: Norbert Pontzer; 301/496-7736, ext. 284; e-mail: pontzern@od.nih.gov

Nestin is an intermediate filament protein first described in early embryonic neuroepithelial stem cells. Although not found in most cells of the mature CNS, nestin is the predominant marker used to detect the small population of undifferentiated cells. The presence of nestin identifies stem, progenitor and some tumor cells in the CNS, and also labels areas of reactive gliosis in the CNS. Available methods to detect nestin use antibodies generated against rat nestin protein. Since rat and human nestin have only about fifty percent sequence homology, these antibodies may not be optimal for detecting nestin in human cells.

NIH scientists used a novel human nestin immunogen to generate polyclonal and monoclonal antibodies that bind with high affinity and specificity to human nestin. The immunogen was expressed from a 450 base-pair segment of human nestin mRNA, which has 11 nucleotide differences from previously published human nestin. These antibodies increase the specificity to accurately detect human nestin in all stages of brain development and will increase our understanding of glial differentiation. In addition, this technology may be useful for detecting glioblastomas or other early stage neuroectodermal tumors and for following transplanted stem cells.

Dated: April 20, 2001.

Jack Spiegel,

Director, Division of Technology Development and Transfer, Office of Technology Transfer, National Institutes of Health.

[FR Doc. 01-10580 Filed 4-27-01; 8:45 am]

BILLING CODE 4140-01-P