Molecular imaging and chemistry: defining the future

There are around 200 Nuclear Medicine Centers in India. PET based product mainly 18F-FDG is localized around big metropolitan cities where there is a centralized cyclotron facility for producing 18F and which can be transported quickly. At present around 18 cyclotrons are operational in India, which fulfill the need of 18F-FDG to greater part of the country. The first Medical cyclotron (16 MeV) was commissioned in the year 2002 at Radiation Medicine Centre, Mumbai. Since then a number of private set ups have installed cyclotron all over the country. The market for 18F-FDG is far superior to any other diagnostic product in India. Other than 18F-FDG, several other 18F based radiopharmaceuticals such as 18F-sodium fluoride for bone cancer detection (used since 2007), 18F-MISO for tumor hypoxia imaging and 18F-FLT for cancer detection (used since 2010) at the cellular level are now being used at limited nuclear medicine centers in India.

FDG is the only PET or SPECT radiopharmaceutical being supplied commercially to many centers. Today in our country we have 98 Running  PET/CT Centers and 70 centers are going be added in the next couple of years. Every center if doing average 5 cases a day and running 20 days then total FDG doses are 9800/month and 117600/year.

Another promising PET radionuclide i.e. 68Ga which is available through a generator is gaining wide acceptance among the clinics. 68Ga labelled peptide such as DOTA-NOC, DOTA-TATE for neuroendocrine tumor imaging is used in number of Nuclear medicine centers in India. There is a small percentage of use of 11C-CO2, 13N-NH3, 15O-H2O and 82Rb which are in use for myocardial perfusion imaging.

Positron emission tomography has deep roots in neuroscience stemming from its first application in brain tumour and brain metabolism imaging. PET neuroimaging has evolved considerably since its birth and continues to play a prominent role in the study of neurochemistry in the living human brain-addressing research questions related to, among many others, protein density, drug occupancy, and endogenous neurochemical release.  New radiotracers combined with new imaging protocol and equipment (e.g. simultaneous MR-PET) position PET neuroimaging for major breakthroughs in our understanding of human brain function and disease.  Using recent studies of neuroinflammation and neuroepigenetics, I will attempt to highlight emerging opportunities with PET and the future needs of the field.  Additionally, I will highlight the use of simultaneous MR-PET and dissecting temporal information from neuroimaging in an attempt to understand and validate models of neurochemical transmission.

Like other MRI contrast agents, CEST contrast from an endogenous species or an exogenous agent depends upon agent concentration, water or proton exchange rates, the relaxation rate of water protons, and other factors.  CEST is governed by the exchange condition, k_ex <Dw, so if Dw is large, then faster exchanging systems can be used to initiate CEST contrast.  It is relatively easy to modify Dw with paramagnetic complexes but manipulating k_ex is generally more difficult.  Many types of paraCEST complexes have been reported over the past few years including those that can be CEST activated by irradiation at the frequency of a bound water molecule and others that are activated at the frequency of an exchanging ligand –NH or –OH proton.     Even though water exchange rates in paramagnetic lanthanide complexes can be modified quite dramatically, water exchange in most paraCEST complexes remains too fast to achieve optimal CEST contrast.  Conversely, many diaCEST agents that use an –OH or –NH resonance for CEST activation have proton exchange rates that are too slow for optimal CEST contrast.  Proton exchange rates can of course be modified by temperature but simple chemical principles can also be used.  For example, when a ligand –OH group coordinates directly to a metal ion, proton exchange from that bound –OH group is considerably faster. Using this principle, we have been exploring the possibility of using lanthanide shift reagents (LSR) to alter the frequency of slowing exchanging protons in endogenous metabolites such as glucose and lactate. The exchanging lactate –OH proton can potentially be used for CEST detection but the chemical shift of the lactate –OH proton is very close to water itself.  Hence, it is quite difficult to activate the exchanging lactate –OH proton without substantially altering the water signal.  Lactate is overproduced by most tumors even in the presence of abundant oxygen (the Warburg effect) so a method for direct imaging of lactate production by tumors could become a useful tool for monitoring tumor aggressiveness.  To test this idea, we have used EuDO3A and other derivatives as aqueous shift reagents (SR) to shift the –OH resonance of lactate far away from water protons and used CEST to quantify the amount of lactate produced by tumor cells growing in tissue culture and, more recently, in vivo. Interestingly, the intensity of the lactate CEST signal is more intense in a slightly acidic environment and is more intense at 37°C than at 25°C.  This new SR approach provides a framework for enabling metabolite-selective imaging of endogenous molecules by MRI.

Among imaging techniques, MRI has great potential to enable mapping of neural events with excellent specificity, spatiotemporal resolution and unlimited tissue penetration depth. Molecular imaging approaches using neurotransmitter-sensitive MRI agents have appeared recently to study neuronal activity, along with the first successful in vivo MRI studies. 

The design of bioresponsive MRI agents for molecular targets such as neurotransmitters is highly challenging. So far, two approaches have been explored to tackle this problem, one involving host molecules developed by nature (i.e. proteins), and the second one involving supramolecular chemistry and artificial host molecules. In this second approach, our objective was to create a synthetic molecular platform, smaller than the robust proteins, with appropriate recognition moieties for neurotransmitters that generate an MR signal change upon neurotransmitter binding. We targeted zwitterionic amino acid neurotransmitters without necessarily seeking for selectivity to individual neurotransmitters. Our chemical design offers ditopic binding via interactions between a positively charged metal complex and the carboxylate and between the crown ether moiety and the amine of amino acids. The probes show selectivity towards zwitterionic over non-zwitterionic neurotransmitters. Neurotransmitter binding leads to a remarkable relaxivity decrease, due to a decrease in the hydration number. One of the probes was successfully used to monitor neural activity in acute mouse brain slices by MRI.

Since functional magnetic resonance imaging (fMRI) was first demonstrated in 1991, the potential of the technique in cognitive neuroscience, neurology and psychiatry was apparent. Technical improvements in magnet and gradient hardware, combined with the work of many researchers to develop robust methods for experimental design, data acquisition and analysis have led to the method now being the pre-eminent technique in cognitive neuroscience research, with applications also in clinical neurology and in the understanding of psychiatric disease. The use of the technique has moved beyond the stage of relating simple functional tasks to anatomy, to investigating brain networks, neural connectivity and the anatomical substrates of complex neural processing. This work is nearly all based on exploiting the endogenous MRI contrast associated with deoxyhaemoglobin and the effect that local changes in neural activity have on blood oxygenation and consequently the local concentration of deoxyhaemoglobin (BOLD effect – blood oxygenation level dependent contrast).  In addition to investigating cognitive and sensory challenges, fMRI can also be used to explore the brain’s reaction to neuro-active drugs by recording the BOLD response to pharmacological challenge (phMRI).  The principles and application of phMRI will be discussed in this presentation, with examples of both human and animal implementations and the use of double challenges to elucidate neurotransmitter pathways and neurovascular coupling. There will be a focus on the serotonergic and glutamatergic neurotransmitter systems in the context of depression and schizophrenia.

Most imaging applied within clinical drug development relies upon downstream markers of disease response (e.g. CT measurement of tumour size). Whilst such techniques remain important there is no doubt that molecular imaging has the potential to fill critical knowledge gaps in the early development of a drug. Important questions include- has the drug reached and engaged with the intended target and has there been some pharmacological response? Too often large, costly clinical trials proceed without these questions being adequately addressed. Whilst there are good examples of molecular imaging in pharmaceutical product development what are the impediments to a broader and more impactful use? In some cases molecular imaging may not be tractable based on current chemistry, it may be perceived as too costly or there may be insufficient time to develop a method. To ensure drug research benefits from molecular imaging tools, drug R&D organisations must develop strong multi-disciplinary partnerships with academic and commercial imaging groups. Through close collaboration we must share challenges and promote early scientific engagement with a focus on translation to the clinical experiment. If successful we will kill off poor drug candidates early, accelerate promising medicines towards clinical use and reduce the cost of drug development. All of these will improve industry productivity and deliver much needed medicines to patients. This talk will highlight the opportunities for molecular imaging and begin to identify the impediments towards broader use.