Phosphocyclocreatine is the dominant form of cyclocreatine in control and creatine transporter deficiency patient fibroblasts

Abstract

Creatine transporter deficiency (CTD) is a metabolic disorder resulting in cognitive, motor, and behavioral deficits. Cyclocreatine (cCr), a creatine analog, has been explored as a therapeutic strategy for the treatment of CTD. We developed a rapid, selective, and accurate HILIC ultra‐performance liquid chromatography‐tandem mass spectrometry (UPLC‐MS/MS) method to simultaneously quantify the intracellular concentrations of cCr, creatine (Cr), creatine‐d3 (Cr‐d3), phosphocyclocreatine (pcCr), and phosphocreatine (pCr). Using HILIC‐UPLC‐MS/MS, we measured cCr and Cr‐d3 uptake and their conversion to the phosphorylated forms in primary human control and CTD fibroblasts. Altogether, the data demonstrate that cCr enters cells and its dominant intracellular form is pcCr in both control and CTD patient cells. Therefore, cCr may replace creatine as a therapeutic strategy for the treatment of CTD.

Impedimetric Sensors for Cyclocreatine Phosphate Determination in Plasma Based on Electropolymerized Poly(o-phenylenediamine) Molecularly Imprinted Polymers

“…LOD and LOQ were 2.47 × 10 –10 and 8.25 × 10 –10 mol/L, respectively. As a result, it is clear from comparing the proposed method with the published ones for CCrP determination that our method provides a linearity range (22.9–0.23 ng/mL) close to those of reported tandem mass spectroscopy methods. , Figure S6 provides the EIS experiment result with different CCrP concentrations.…”

“…Due to the crucial role that CCrP plays, we inspected the analytical methodologies used for its monitoring closely. , CCrP detection is difficult to analyze using conventional high-performance liquid chromatography (HPLC) as it is a highly polar molecule that is nonelectroactive and nonfluorescent and has a weak UV absorption signal. Moreover, the presence of structural analogue compounds in biological fluids at high concentration, such as creatine (Cr), CCr, creatine phosphate (CrP), and adenosine triphosphate (ATP) makes the task even more challenging.…”

“…Moreover, the presence of structural analogue compounds in biological fluids at high concentration, such as creatine (Cr), CCr, creatine phosphate (CrP), and adenosine triphosphate (ATP) makes the task even more challenging. A literature survey revealed that CCrP has only reported to be monitored with methods based on hydrophilic interaction liquid chromatography (HILIC), which can analyze highly hydrophilic small molecules. , From the analytical chemistry standpoint, HPLC is considered the benchmark standard for pharmaceutical analysis; however, it has various limitations related to its high cost, long cycle time, and high organic solvent consumption. , Alternatively, impedimetric sensors have numerous advantages such as low energy usage, ability to detect nonelectroactive molecules, high sensitivity, relatively simple, and eco-friendly process. Furthermore, such sensors can be highly selective based on modifying the electrode surface with suitable surface recognition elements, such as antibodies, aptamers, or molecularly imprinted polymers (MIPs). − Therefore, establishing a sensor based upon MIP for CCrP measurement provides a promising and advantageous alternative to determine small polar molecules in plasma.…”

Cyclocreatine treatment ameliorates the cognitive, autistic and epileptic phenotype in a mouse model of Creatine Transporter Deficiency

“…It has been reported that lipophilic analogs and other derivatives of Cr can enter cells independently of CrT and could represent a promising approach for CTD treatment 10 , 16 – 19 . In particular, cCr is a nearly planar Cr analog that can be phospho/dephosphorylated by Cr kinase, thus mimicking the metabolic function of Cr 10 , 20 , 21 . However, the evaluation of cCr and other treatment options in CTD has been limited to biochemical measurements, behavioral outcomes and in vitro assays of neuronal function.…”

The Role of Preclinical Models in Creatine Transporter Deficiency: Neurobiological Mechanisms, Biomarkers and Therapeutic Development

“…Cyclocreatine (cCr) is an artificial analog of Cr, which can enter cells independently of CRT [135,136] and can be phospho/dephosphorylated by CK [136,137], mimicking the metabolic function of Cr. This compound has been proposed as a possible replacement of endogenous Cr.…”