a b s t r a c tWe have studied the naturally split a subunit of the DNA polymerase III (DnaE) intein from Nostoc punctiforme PCC73102 (Npu) using purified proteins and determined an apparent first-order rate constant of ð1:1 AE 0:2Þ Â 10 À2 s À1 at 37°C. This represents the highest rate reported for the protein trans-splicing reaction so far (t 1=2 of $ 60 s). Furthermore, the reaction was very robust and highyielding with respect to different extein sequences, temperatures from 6 to 37°C, and the presence of up to 6 M urea. Given these outstanding properties, the Npu DnaE intein appears to be the intein of choice for many applications in protein and cellular chemistry.

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Phytosanitary irradiation (PI) treatments are promising measures to overcome quarantine barriers to trade and are currently used in several countries. Although PI has advantages compared with other treatments one disadvantage bedevils research, approval, and application: organisms may remain alive after importation. Although this does not preclude their use as a phytosanitary treatment, it does leave the treatment without an independent verification of efficacy and places a greater burden for assuring quarantine security on the research supporting the treatment. This article analyses several factors that have been hypothesized to affect PI efficacy: low oxygen, pest stage, host, dose rate, and temperature. Of these factors, the first is known to affect efficacy, whereas host and dose rate probably need more research. The International Plant Protection Convention considered several PI treatments for its international standard on phytosanitary treatments and did not approve some at first because of perceived problems with the research or the presence of live adults after irradiation. Based on these concerns recommendations for research and dealing with the issue of live adults postirradiation are given. Generic PI treatments are suggested.

TransCon CNP is a C-type natriuretic peptide (CNP-38) conjugated via a cleavable linker to a polyethylene glycol carrier molecule, designed to provide sustained systemic CNP levels upon weekly subcutaneous administration. TransCon CNP is in clinical development for the treatment of comorbidities associated with achondroplasia. In both mice and cynomolgus monkeys, sustained exposure to CNP via TransCon CNP was more efficacious in stimulating bone growth than intermittent CNP exposure. TransCon CNP was well tolerated with no adverse cardiovascular effects observed at exposure levels exceeding the expected clinical therapeutic exposure. At equivalent dose levels, reductions in blood pressure and/or an increase in heart rate were seen following single subcutaneous injections of the unconjugated CNP-38 molecule or a daily CNP-39 molecule (same amino acid sequence as Vosoritide, USAN:INN). The half-life of the daily CNP-39 molecule in cynomolgus monkey was estimated to be 20 minutes, compared with 90 hours for CNP-38, released from TransCon CNP. C max for the CNP-39 molecule (20 mg/kg) was approximately 100-fold higher, compared with the peak CNP level associated with administration of 100 mg/kg CNP as TransCon CNP. Furthermore, CNP exposure for the daily CNP-39 molecule was only evident for up to 2 hours postdose (lower limit of quantification 37 pmol/l), whereas TransCon CNP gave rise to systemic exposure to CNP-38 for at least 7 days postdose. The prolonged CNP exposure and associated hemodynamically safe peak serum concentrations associated with TransCon CNP administration are suggested to improve efficacy, compared with short-lived CNP molecules, due to better therapeutic drug coverage and decreased risk of hypotension. SIGNIFICANCE STATEMENT The hormone C-type natriuretic peptide (CNP) is in clinical development for the treatment of comorbidities associated with achondroplasia, the most common form of human dwarfism. The TransCon Technology was used to design TransCon CNP, a prodrug that slowly releases active CNP in the body over several days. Preclinical data show great promise for TransCon CNP to be an effective and well-tolerated drug that provides sustained levels of CNP in a convenient once-weekly dose, while avoiding high systemic CNP bolus concentrations that can induce cardiovascular side effects.

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Hypoparathyroidism (HP) is a condition of parathyroid hormone (PTH) deficiency leading to abnormal calcium and phosphate metabolism. The mainstay of therapy consists of vitamin D and calcium supplements, as well as adjunct Natpara (PTH(1‐84)). However, neither therapy optimally controls urinary calcium (uCa) or significantly reduces the incidence of hypercalcemia and hypocalcemia. TransCon PTH, a sustained‐release prodrug of PTH(1‐34) in development for the treatment of HP, was designed to overcome these limitations. To determine the pharmacokinetics and pharmacodynamics of TransCon PTH, single and repeat s.c. dose studies were performed in rats and monkeys. TransCon PTH demonstrated a half‐life of 28 and 34 hours in rats and monkeys, respectively. After repeated dosing, an infusion‐like profile of the released PTH, characterized by low peak‐to‐trough levels, was obtained in both species. In intact rats and monkeys, daily subcutaneous administration of TransCon PTH was associated with increases in serum calcium (sCa) levels and decreases in serum phosphate levels (sP). In monkeys, at a single dose of TransCon PTH that increased sCa levels within the normal range, a concurrent decrease in uCa excretion was observed. In 4‐week repeat‐dose studies in intact rats and monkeys, uCa excretion was comparable to controls across all dose levels despite increases in sCa levels. Further, in a rat model of HP, TransCon PTH normalized sCa and sP levels 24 hours per day. This was in contrast to only transient trends toward normalization of sCa and sP levels with an up to 6‐fold higher molar dose of PTH(1‐84). After repeated dosing to HP rats, uCa excretion transiently increased, corresponding to increases in sCa above normal range, but at the end of the treatment period, uCa excretion was generally comparable to sham controls. TransCon PTH was well tolerated and the observed pharmacokinetics and pharmacodynamics were in line with the expected action of physiological replacement of PTH. © 2019 American Society for Bone and Mineral Research.