Thomas J. McIntosh, Ph.D. (Carnegie-Mellon University)
Professor, Department of Cell Biology
CMB, Structural Biology and Biophysics and Biological Chemistry Programs
443 Sands Bldg., Box 3011
Duke University Medical Center
Durham, NC 27710
Our research primarily focuses on understanding the mechanisms by which membrane lipids govern the function of key membrane channels. Such lipid modulation has been proposed to be caused either by direct lipid-protein interactions or indirectly by collective properties of the lipid matrix such as bilayer width or elasticity.
A particularly critical lipid molecule is cholesterol, which is found in relatively large concentrations (>30 mol% lipid) in most cell plasma membranes. We are currently analyzing the role of cholesterol on the function of two classes of physiologically important plasma membrane channels, aquaporin (AQP) water channels and transient receptor potential (TRP) cation channels. For the AQPs, we are focusing on AQP-0 of the eye lens and AQP-4 of glial cells in the brain. For the case of TRP channels we are initially analyzing the TRP vanilloid 1 channel (TRPV1), a key signal transduction component in nociceptors.
We have found that the water permeability of both AQP-0 and AQP-4 channels depends strongly on the composition of the membrane lipid bilayer. Specifically, there is a marked reduction in AQP permeability with increasing bilayer cholesterol content, with this reduction quantitatively correlating with increasing width of the bilayer.
We have also found that increasing cholesterol reduces the TRPV1 current in cell plasma membranes. Here this effect is due to a direct interaction of cholesterol with an amino acid sequence in the fifth TRPV1 transmembrane helix that is consistent with cholesterol recognition amino acid residue (CRAC) motifs found in other proteins that bind cholesterol. We have shown that cholesterol docks to this sequence and that a single point mutation in this sequence makes TRPV1 insensitive to cholesterol addition or removal.
Thus, for two different channel types, AQP water channels and TRPV1 ion channels, membrane lipid cholesterol significantly modifies channel function. However, the molecular mechanisms are quite different for these two channels, with an indirect bilayer effect for AQP and a specific cholesterol interaction in the case of TRPV1.
Our future plans are determining whether AQP acylation, which promotes the partitioning of AQP0 and AQP4 into membrane microdomains with high cholesterol content, decreases channel permeability and therefore provides channel regulation. A second project involves the effects of cholesterol on key membrane channels in sperm that contain the CRAC motif. In sperm a necessary event in the fertilization process is the removal of cholesterol from the plasma membrane resulting in an increase of calcium flux across the membrane. Our hypothesis is that this increased permeability is caused by cholesterol removal from CRAC sequences in sperm channels.
A second interest, in collaboration with Dr. Mark Grinstaff of Boston University, is improving lipid-amphiphile complexes for DNA and gene delivery to cells for biological and medical applications. Dr. Grinstaff has designed lipids that bind and then release DNA based on changes in electrostatic interactions due to enzymatic reactions, and we are involved in determining the structure of these DNA-lipid complexes. Specific complexes have been found to be effective for in vitro DNA transfections.
Tong J, Wu Z, Briggs MM, Schulten K, McIntosh TJ. (2016) The Water Permeability and Pore Entrance Structure of Aquaporin-4 Depend on Lipid Bilayer Thickness. Biophys J. 12;111(1):90-9.
Wang W, McIntosh TJ, Jiang X, Doddareddy R, Dell EC, Zhou H. (2016) Deciphering the In Vivo Performance of a Monoclonal Antibody to Neutralize Its Soluble Target at the Site of Action in a Mouse Collagen-Induced Arthritis Model. Pharm Res. 33(4):1040-9.
Zheng S, McIntosh T, Wang W. (2015) Utility of free and total target measurements as target engagement and efficacy biomarkers in biotherapeutic development--opportunities and challenges. J Clin Pharmacol. 55 Suppl 3:S75-84.
McIntosh TJ. Stepping between membrane microdomains. Biophys J. 17;108(4):783-4.
Wang W, Wang X, Doddareddy R, Fink D, McIntosh T, Davis HM, Zhou H. (2014) Mechanistic pharmacokinetic/target engagement/pharmacodynamic (PK/TE/PD) modeling in deciphering interplay between a monoclonal antibody and its soluble target in
cynomolgus monkeys. AAPS J. 16(1):129-39.
Han C, McIntosh TS, Geist BJ, Jiao T, Puchalski TA, Goldberg KM, Yang TY, Pendley CE, Zhou H, Davis HM. (2014) A novel approach to evaluate the pharmacokinetic biocomparability of a monoclonal antibody derived from two different cell lines using simultaneous crossover design. AAPS J. 16(1):125-8.
Zhang XX, Lamanna CM, Kohman RE, McIntosh TJ, Han X, Grinstaff MW. (2013) Lipid-mediated DNA and siRNA Transfection Efficiency Depends on Peptide Headgroup. Soft Matter. 5;9(17).
Fetterly GJ, Aras U, Meholick PD, Takimoto C, Seetharam S, McIntosh T, de Bono JS, Sandhu SK, Tolcher A, Davis HM, Zhou H, Puchalski TA. (2013) Utilizing pharmacokinetics/pharmacodynamics modeling to simultaneously examine free CCL2, total CCL2 and carlumab (CNTO 888) concentration time data. J Clin Pharmacol. 53(10):1020-7.
Kelley M, Ahene AB, Gorovits B, Kamerud J, King LE, McIntosh T, Yang J. (2013) Theoretical considerations and practical approaches to address the effect of anti-drug antibody (ADA) on quantification of biotherapeutics in circulation. AAPS J. 15(3):646-58. Review.
Geist BJ, Davis D, McIntosh T, Yang TY, Goldberg K, Han C, Pendley C, Davis HM. (2013) A novel approach for the simultaneous quantification of a therapeutic monoclonal antibody in serum produced from two distinct host cell lines. MAbs. 5(1):150-61.