The study of IL-12, its receptors and subunits offers promising avenues for therapeutic research and for elucidation of mechanisms of immunity. As the initiator of cell mediated immunity, it has been suggested that IL-12 has therapeutic potential as a stimulator of cell mediated immune responses to microbial pathogens, metastatic cancers, and viral infections such as AIDS.
Researchers at the Trudeau Institute have shown that a splice variant of IL-12Rβ1 (IL12Rβ1ΔTM) augments the chemotactic activity of motile cells and augments the signaling mediated by ligands of the IL-12 receptor complex.
Since IL12Rβ1 mediates the activity of the cytokines Il-12, IL-12p70, IL-23 and IL-12(p40)2 it impacts many pathways that effectuate immune responses. It also plays an important role in regulation of damaging inflammatory responses associated with autoimmune pathologies (i.e. Arthritis, Psoriasis, and MS). Absence of IL-12Rβ1 function is a factor that results in predisposition to increased susceptibility to tuberculosis and other pathogens.
This novel assay is designed for use by scientists to determine whether the mRNA splice variant of the IL-12 gene (IL12Rβ1ΔTM) is present within their experimental systems.
Current real time PCR protocols cannot distinguish between IL12Rβ1 and IL12Rβ1ΔTM due to the relatively small difference in nucleotide sequence between them.
Interleukin-12 (IL-12) is a pro-inflammatory cytokine that has regulatory effects on T and natural killer (NK) cells; it is expressed by activated macrophages that serve as an essential inducer of Th1 cell development. This cytokine has been found to be important for sustaining a sufficient number of memory/effector Th1 cells to mediate long-term protection to an intracellular pathogen.
Researchers at the Trudeau Institute have determined that both the IL12 Receptor Beta 1 (IL12Rβ1) gene as well as an IL12Rβ1 alternative splice variant (IL12Rβ1ΔTM) are induced in a mouse model of human disease. This splice variant had been previously described but no function had been assigned to it. Careful examination of IL12Rβ1 mRNA expression in dendritic cells led to detection of an IL-12Rβ1 gene product.
The Trudeau research shows that this splice variant is induced in cells that are exposed to Mycobacterium tuberculosis and that it is expressed in vivo in the lungs of mice infected with these pathogens.
Typical polyclonal anti-IL12Rβ1 antibody reagents bind to the extracellular portion of the IL-12Rβ1 molecule making it likely that it will not distinguish between the intact and spliced molecules. Therefore novel primer design was undertaken and combined with Spectratype analysis.
Quantifying the relative ratios of IL12Rβ1ΔTM using Spectratype analysis is performed as follows; cDNA is amplified with primers that flank the transmembrane-encoding region. The resulting amplicons are then carboxyfluorescein (FAM)-labeled via a run-off PCR reaction with a single FAM-conjugated primer. Since FAM-labeled IL12Rβ1 and IL12Rβ1ΔTM amplicons are different sizes, their relative abundance can be quantified using fluorescent capillary electrophoresis.
The assay described above is a simple and informative way to measure the relative levels of the regular IL-12Rβ1 molecule and the alternatively spliced molecule that is induced in response to immune challenge from cancer, pathogens, allergens and viral infection.
Potential Areas of Application:
- Cytokine research
- Cancer research
- Autoimmune research
- Vaccine adjuvant research
- Provides the basis for a PCR reagent kit for researchers interested in cytokines IL12, IL-12p70, IL-23 and IL- 12(p40)2
- Accurately distinguishes between IL12Rβ1 and IL12Rβ1ΔTM – no existing products address this need
Provisional patent no. 61/304,025
Health Research, Inc. on behalf of the Trudeau Institute is seeking partners to assist in the further development of this novel discovery into therapeutic products. Partnership opportunities exist in the form of licensing and/or sponsored research with intellectual property rights.
Available for license
Andrea M. Cooper, Ph.D.
Richard T. Robinson, Ph.D.
Robinson, Richard T., Khader, Shabaana A., Martino, Cynthia A., Fountain, Jeffrey J., Teixeira-Coelho, Maria, Pearl, John E., Smiley, Stephen T., Winslow, Gary M., Woodland, David L., Walter, Michael J., Conejo-Garcia, Jose R., Gubler, Ueli, and Cooper, Andrea M. Mycobacterium tuberculosis infection induces ilrb1 to generate a novel IL12Rβ1 isoform that enhances DC migration. Journal of Experimental Medicine 2010 Mar 15;207(3):591-605. Epub 2010 Mar 8.
Robinson, Richard and Cooper, Andrea Non-hematopoietic IL12p40-responsiveness is required for tubercular granuloma formation. The Journal of Immunology, 2010, 184, (Meeting Abstract Supplement) 42.19 http://www.jimmunol.org/cgi/content/meeting_abstract/184/1_MeetingAbstracts/42.19
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