Hyperpolarized α-keto[1-C]isocaproate as a C magnetic resonance spectroscopic agent for profiling branched chain amino acid metabolism in tumors

Magnetic resonance spectroscopy (MRS) is a technique that allows us to non-invasively detect multiple small metabolites within cells or extracellular spaces (1, 2). The clinical use of MRS as an adjunct to magnetic resonance imaging (MRI), particularly in cancer detection and treatment response evaluation, has expanded dramatically over the past several years. Although MRS is theoretically applicable to any nucleus possessing spin, the most important and more frequently investigated applications are in proton (H) and carbon-13 (C) (1, 3, 4). C MRS is superior to H MRS in many respects (5-7). C MRS can provide specific information about the identity and structure of biologically important compounds. The chemical shift range for carbon (250 ppm) is much larger than that for proton (15 ppm), allowing for improved resolution of metabolites. In addition, the T relaxation time of C in small molecules is much longer than that of H (0.1–2.0 s in a magnetic field of 0.1–3.0 T), allowing the generation of hyperpolarized C-labeled tracers outside the subject and the MRI scanner. Some of these tracers are endogenous and therefore have lower toxicity than drugs and exogenous contrast media (2). However, C MRS is limited by the low natural abundance of C (1.1%) and its low magnetogyric ratio (γ of C is one quarter that of H) (1, 6). Several techniques have been used to overcome the C MRS limitation through enhancing the polarization of nuclear spins. One technique is proton decoupling, which eliminates the coupling of H with C by irradiating the entire H resonance absorption range and consequently collapsing C resonances to singlets (8, 9). The signal/noise ratio of C resonances has been shown to be significantly increased with proton decoupling. Another technique is known as dynamic nuclear polarization (DNP), which introduces one or more C molecules into a metabolic substrate (3, 10, 11). DNP transfers high electron spin polarization to nuclear spins microwave irradiation. Nearly 100% nuclear polarization for H and 50% for C can be achieved in various organic molecules when DNP is performed in a strong magnetic field and at cryogenic temperatures. Hyperpolarization of protons ex vivo is less interesting for medical applications, because most of the hyperpolarization would have vanished before the molecule reaches the target organ. However, the T of C in small molecules, in general, is much longer than the T of protons. A large number of low molecular weight substances with C T in excess of 10 s are available. Replacing the C isotope (98.9% natural abundance) with the C isotope at a specific carbon or carbons in a metabolic substrate does not affect the substrate's biochemistry. With C MRS, the body tissues are virtually invisible, and only regions where the hyperpolarized C-labeled substance is present will appear in the generated images. Thus, C-labeled substrates can provide >10,000-fold enhancement of the C MRS signals from the substrate and its subsequent metabolic products, allowing the assessment of changes in metabolic fluxes through glycolysis, citric acid cycle, and fatty acid synthesis (2, 11, 12). Vascular and perfusion imaging can also be performed without background signal from surrounding tissues (6, 13). Karlsson et al. generated a hyperpolarized small molecule, α-keto[1-C]isocaproate ([1-C]KIC) as a C MRS agent for imaging the molecular signature of branched chain amino acid metabolism, which is regulated by the branched chain amino acid transferase (BCAT) (5). There are two isoforms, BCAT1 and BCAT2, that code for mitochondrial and cytosolic BCAT, respectively (5, 14). BCAT is highly expressed in early embryogenesis, and it is also a target for MYC activity during oncogenesis. BCAT has been shown to be a useful marker for grading and genetic characterization of tumors. KIC is a substrate of BCAT and is metabolized to leucine . Karlsson et al. demonstrated that metabolism of the hyperpolarized [1-C]KIC yielded unprecedented MRI contrast between EL4 murine lymphoma and surrounding healthy tissue, but yielded no contrast between R3230AC rat mammary adenocarcinoma and its surrounding tissue. The [1-C]leucine signal detected in the two tumor models correlated well with measurements of the BCAT activity in the two tumor tissues. The investigators concluded that the understanding of metabolic differences between tumors could be advanced with use of the hyperpolarized [1-C]KIC biomarker in assessing tissue BCAT activity by means of the [1-C]leucine signal (5).