Sebelipase alfa over 52 weeks reduces serum transaminases, liver volume and improves serum lipids in patients with lysosomal acid lipase deficiency.

BACKGROUND & AIMS: Lysosomal acid lipase deficiency is an autosomal recessive enzyme deficiency resulting in lysosomal accumulation of cholesteryl esters and triglycerides. LAL-CL04, an ongoing extension study, investigates the long-term effects of sebelipase alfa, a recombinant human lysosomal acid lipase. METHODS: Sebelipase alfa (1mg/kg or 3mg/kg) was infused every-other-week to eligible subjects. Safety and tolerability assessments, including liver function, lipid profiles and liver volume assessment, were carried out at regular intervals. RESULTS: 216 infusions were administered to eight adult subjects through week 52 during LAL-CL04. At week 52, mean alanine aminotransferase and aspartate aminotransferase levels were normal with mean change from baseline of -58% and -40%. Mean changes for low-density lipoprotein, total cholesterol, triglyceride and high-density lipoprotein were -60%, -39%, -36%, and +29%, respectively. Mean liver volume by magnetic resonance imaging and hepatic proton density fat fraction decreased (12% and 55%, respectively). Adverse events were mainly mild and unrelated to sebelipase alfa. Infusion-related reactions were uncommon: three events of moderate severity were reported in two subjects; one patient's event was suggestive of a hypersensitivity-like reaction, but additional testing did not confirm this, and the subject has successfully re-started sebelipase alfa. Of samples tested to date, no anti-drug antibodies have been detected. CONCLUSIONS: Long-term dosing with sebelipase alfa in lysosomal acid lipase-deficient patients is well tolerated and produces sustained reductions in transaminases, improvements in serum lipid profile and reduction in the hepatic fat fraction. A randomized, placebo-controlled phase 3 trial in children and adults is underway (ARISE: NCT01757184).

At least four distinct circadian regulatory mechanisms are required for all phases of rhythms in mRNA amount.

Since the advent of techniques to investigate gene expression on a large scale, numerous circadian rhythms in mRNA amount have been reported. These rhythms generally differ in amplitude and phase. The authors investigated how far a parameter not regulated by the circadian clock can influence the phase of a rhythm in RNA amount arising from a circadian rhythm of transcription. Using a discrete-time approach, they modeled a sinusoidal rhythm in transcription with various constant exponential RNA decay rates. They found that the slower the RNA degradation, the later the phase of the RNA amount rhythm compared with the phase of the transcriptional rhythm. However, they also found that the phase of the RNA amount rhythm is limited to a timeframe spanning the first quarter of the period following the phase of the transcriptional rhythm. This finding is independent of the amplitude and vertical shift of the transcriptional rhythm or even of the way constant RNA degradation is modeled. The authors confirmed their results with a continuous-time model, which allowed them to derive a simple formula relating the phase of the RNA amount rhythm solely to the phase and period of its sinusoidal transcriptional rhythm and its constant RNA half-life. This simple formula even holds true for the best sinusoidal approximations of a nonsinusoidal rhythm of transcription and RNA amount. When expanding the model to include additional events with constant exponential kinetics, such as RNA processing, they found that each event expands the phase limit by another quarter of the period when it occurs in sequence but not when it occurs as a competing process. However, the limit expansion comes at the price of minuscule amplitudes. When using a discrete-time approach to model constant rates of transcription with a sinusoidal RNA half-life, the authors found that the phase of the RNA amount rhythm is unaffected by changes in the constant rate of transcription. In summary, their data show that at least 4 distinct circadian regulatory mechanisms are required to allow for all phases in rhythms of RNA amount, one for each quarter of the period.

Chlamydomonas reinhardtii strain CC-124 is highly sensitive to blue light in addition to green and red light in resetting its circadian clock, with the blue-light photoreceptor plant cryptochrome likely acting as negative modulator.

The unicellular green alga Chlamydomonas reinhardtii has long served as model organism for studies on the circadian clock. This clock is present in all eukaryotes and some prokaryotes allowing them to anticipate and take advantage of the daily oscillations in the environment. Although much is known about the circadian clock in C. reinhardtii, the photoreceptors mediating entrainment of the clock to the daily changes of light remain obscure. Based on its circadian rhythm of phototaxis as a reporter of the clock's phase, we show here that C. reinhardtii strain CC-124 is highly sensitive to blue light of 440 nm when resetting its circadian clock upon light pulses. Thus, CC-124 differs in this respect from what was previously reported for a cell wall-deficient strain. An action spectrum analysis revealed that CC-124 also responds with high sensitivity to green (540 nm), red (640-660 nm), and possibly UV-A (≤400 nm) light, and therefore shows similarities as well to what has been reported for the cell wall-deficient strain. We also investigated two RNA interference strains with reductions in the level of the blue light photoreceptor plant cryptochrome (CPH1). One of them, the strain with the greater reduction, surprisingly showed an increased sensitivity in clock resetting upon blue light pulses of 440 nm. This increase in sensitivity reverted to wild-type levels when the RNA interference strain reverted to wild-type protein levels. It suggests that plant cryptochrome in C. reinhardtii could function as negative rather than positive modulator of circadian clock resetting.

Improved automated monitoring and new analysis algorithm for circadian phototaxis rhythms in Chlamydomonas.

Automated monitoring of circadian rhythms is an efficient way of gaining insight into oscillation parameters like period and phase for the underlying pacemaker of the circadian clock. Measurement of the circadian rhythm of phototaxis (swimming towards light) exhibited by the green alga Chlamydomonas reinhardtii has been automated by directing a narrow and dim light beam through a culture at regular intervals and determining the decrease in light transmittance due to the accumulation of cells in the beam. In this study, the monitoring process was optimized by constructing a new computer-controlled measuring machine that limits the test beam to wavelengths reported to be specific for phototaxis and by choosing an algal strain, which does not need background illumination between test light cycles for proper expression of the rhythm. As a result, period and phase of the rhythm are now unaffected by the time a culture is placed into the machine. Analysis of the rhythm data was also optimized through a new algorithm, whose robustness was demonstrated using virtual rhythms with various noises. The algorithm differs in particular from other reported algorithms by maximizing the fit of the data to a sinusoidal curve that dampens exponentially. The algorithm was also used to confirm the reproducibility of rhythm monitoring by the machine. Machine and algorithm can now be used for a multitude of circadian clock studies that require unambiguous period and phase determinations such as light pulse experiments to identify the photoreceptor(s) that reset the circadian clock in C. reinhardtii.