In vivo ³¹P magnetic resonance spectroscopic imaging (MRSI) for metabolic profiling of human breast cancer xenografts.

PURPOSE: To study cancer associated with abnormal metabolism of phospholipids, of which several have been proposed as biomarkers for malignancy or to monitor response to anticancer therapy. We explored 3D (31) P magnetic resonance spectroscopic imaging (MRSI) at high magnetic field for in vivo assessment of individual phospholipids in two patient-derived breast cancer xenografts representing good and poor prognosis (luminal- and basal-like tumors). MATERIALS AND METHODS: Metabolic profiles from luminal-like and basal-like xenograft tumors were obtained in vivo using 3D (31) P MRSI at 11.7T and from tissue extracts in vitro at 14.1T. Gene expression analysis was performed in order to support metabolic differences between the two xenografts. RESULTS: In vivo (31) P MR spectra were obtained in which the prominent resonances from phospholipid metabolites were detected at a high signal-to-noise ratio (SNR >7.5). Metabolic profiles obtained in vivo were in agreement with those obtained in vitro and could be used to discriminate between the two xenograft models, based on the levels of phosphocholine, phosphoethanolamine, glycerophosphocholine, and glycerophosphoethanolamine. The differences in phospholipid metabolite concentration could partly be explained by gene expression profiles. CONCLUSION: Noninvasive metabolic profiling by 3D (31) P MRSI can discriminate between subtypes of breast cancer based on different concentrations of choline- and ethanolamine-containing phospholipids.

IDH1 R132H mutation generates a distinct phospholipid metabolite profile in glioma.

Many patients with glioma harbor specific mutations in the isocitrate dehydrogenase gene IDH1 that associate with a relatively better prognosis. IDH1-mutated tumors produce the oncometabolite 2-hydroxyglutarate. Because IDH1 also regulates several pathways leading to lipid synthesis, we hypothesized that IDH1-mutant tumors have an altered phospholipid metabolite profile that would impinge on tumor pathobiology. To investigate this hypothesis, we performed (31)P-MRS imaging in mouse xenograft models of four human gliomas, one of which harbored the IDH1-R132H mutation. (31)P-MR spectra from the IDH1-mutant tumor displayed a pattern distinct from that of the three IDH1 wild-type tumors, characterized by decreased levels of phosphoethanolamine and increased levels of glycerophosphocholine. This spectral profile was confirmed by ex vivo analysis of tumor extracts, and it was also observed in human surgical biopsies of IDH1-mutated tumors by (31)P high-resolution magic angle spinning spectroscopy. The specificity of this profile for the IDH1-R132H mutation was established by in vitro (31)P-NMR of extracts of cells overexpressing IDH1 or IDH1-R132H. Overall, our results provide evidence that the IDH1-R132H mutation alters phospholipid metabolism in gliomas involving phosphoethanolamine and glycerophosphocholine. These new noninvasive biomarkers can assist in the identification of the mutation and in research toward novel treatments that target aberrant metabolism in IDH1-mutant glioma.

Metabolic imaging of multiple x-nucleus resonances.

This study describes a technique for fast imaging of x-nuclei metabolites. Due to increased sensitivity and larger chemical shift dispersion at high magnetic fields, images of multiple metabolites can be obtained simultaneously by selective excitation of their resonances with a multifrequency selective radiofrequency pulse at any desired flip angle. This aim is achieved by combining a three-dimensional gradient echo imaging sequence with a Shinnar-LeRoux optimized excitation pulse. A proper choice of bandwidth, imaging matrix size, and field of view allows using the chemical shift dispersion of the different resonances to completely separate their images within one large field of view. The method of fast metabolic imaging is illustrated with (13)C measurements of a phantom containing a solution of (13)C labeled glucose, lactate, and sodium octanoate and by dynamic measurements of the (31)P metabolites phosphocreatine and ß-adenosine triphosphate in human femoral muscle in vivo, both at 7T. With dynamic selective (31)P imaging of the larger part of the upper leg, phosphocreatine signal intensity changes of specific muscles can be studied simultaneously by analyzing the sum of phosphocreatine signals within arbitrarily shaped regions of interest following the muscles' contours. This concept of dynamic metabolic imaging can be applied to other organs and further expanded to other MR-detectable nuclei and metabolites.

NMR at earth's magnetic field using para-hydrogen induced polarization.

A method to achieve NMR of dilute samples in the earth's magnetic field by applying para-hydrogen induced polarization is presented. Maximum achievable polarization enhancements were calculated by numerically simulating the experiment and compared to the experimental results and to the thermal equilibrium in the earth's magnetic field. Simultaneous 19F and 1H NMR detection on a sub-milliliter sample of a fluorinated alkyne at millimolar concentration (∼10(18) nuclear spins) was realized with just one single scan. A highly resolved spectrum with a signal/noise ratio higher than 50:1 was obtained without using an auxiliary magnet or any form of radio frequency shielding.

Effects of targeting the VEGF and PDGF pathways in diffuse orthotopic glioma models.

Currently available compounds that interfere with VEGF-A signalling effectively inhibit angiogenesis in gliomas, but influence diffuse infiltrative growth to a much lesser extent. Development of a functional tumour vascular bed not only involves VEGF-A but also requires platelet-derived growth factor receptor-ß (PDGFRß), which induces maturation of tumour blood vessels. Therefore, we tested whether combined inhibition of VEGFR and PDGFRß increases therapeutic benefit in the orthotopic glioma xenograft models E98 and E473, both displaying the diffuse infiltrative growth that is characteristically observed in most human gliomas. We used bevacizumab and vandetanib as VEGF(R) inhibitors, and sunitinib to additionally target PDGFRß. We show that combination therapy of sunitinib and vandetanib does not improve therapeutic efficacy compared to treatment with sunitinib, vandetanib or bevacizumab alone. Furthermore, all compounds induced reduction of vessel leakage in compact E98 tumour areas, resulting in decreased detectability of these mostly infiltrative xenografts in Gd-DTPA-enhanced MRI scans. These data show that inhibition of VEGF signalling cannot be optimized by additional PDGFR inhibition and support the concept that diffuse infiltrative areas in gliomas are resistant to anti-angiogenic therapy.

Rapid and robust transgenic high-grade glioma mouse models for therapy intervention studies.

PURPOSE: To develop a transgenic mouse model of glioma that can be conveniently used for testing therapy intervention strategies. High-grade glioma is a devastating and uniformly fatal disease for which better therapy is urgently needed. Typical for high-grade glioma is that glioma cells infiltrate extensively into surrounding pivotal brain structures, thereby rendering current treatments largely ineffective. Evaluation of novel therapies requires the availability of appropriate glioma mouse models. EXPERIMENTAL DESIGN: High-grade gliomas were induced by stereotactic intracranial injection of lentiviral GFAP-Cre or CMV-Cre vectors into compound LoxP-conditional mice, resulting in K-Ras(v12) expression and loss of p16(Ink4a)/p19(Arf) with or without concomitant loss of p53 or Pten. RESULTS: Tumors reproduced many of the features that are characteristic for human high-grade gliomas, including invasiveness and blood-brain barrier functionality. Especially, CMV-Cre injection into p53;Ink4a/Arf;K-Ras(v12) mice resulted in high-grade glioma with a short tumor latency (2-3 weeks) and full penetrance. Early detection and follow-up was accomplished by noninvasive bioluminescence imaging, and the practical utility for therapy intervention was shown in a study with temozolomide. CONCLUSION: We have developed a realistic high-grade glioma model that can be used with almost the same convenience as traditional xenograft models, thus allowing its implementation at the forefront of preclinical evaluation of new treatments.

Contrast enhanced susceptibility weighted imaging (CE-SWI) of the mouse brain using ultrasmall superparamagnetic ironoxide particles (USPIO).

Susceptibility weighted imaging (SWI) has been introduced as a novel approach to visualize the venous vasculature in the human brain. With SWI, small veins in the brain are depicted based on the susceptibility difference between deoxyhaemoglobin in the veins and surrounding tissue, which is further enhanced by the use of MR phase information. In this study we applied SWI in the mouse brain using an exogenous iron-based blood-pool contrast agent, with the aims of further enhancing the susceptibility effect and allowing the visualization of individual veins and arteries. Contrast enhanced (CE-) SWI of the brain was performed on healthy mice and mice carrying intracerebral glioma xenografts. This study demonstrates that detailed vascular information in the mouse brain can be obtained by using CE-SWI and is substantially enhanced compared to native SWI (i.e. without contrast agent). CE-SWI images of tumour-bearing mice were directly compared to histology, confirming that CE-SWI depicts the vessels supplying and draining the tumour. We propose that CE-SWI is a very promising tool for the characterization of tumour vasculature.