L 006235

IC50: 0.25 nM

L-006235 is a potent and selective inhibitor of Cathepsin K.

In vitro: After dilution of L-006235 to 0.05 nM, the cathepsin K enzyme activity was initially inhibited, but slowly recovered with a first-order rate constant of 0.023 s-1. The final steady-state enzyme activity was 80-90% that of control, suggesting the complete reversibility of the L-006235-cathepsin K complex. L-006235 was found to be not a substrate for the nitrilase activity of Cat K [1].

In vivo: L-006235 was orally bioavailable in rats, with a terminal half-life of over 3 h. L-006235 was orally dosed in ovariectomized rhesus monkeys once per day for 7 days. Results showed that collagen breakdown products were dose-dependently reduced by up to 76%. Plasma concentrations of L-006235 above the bone resorption IC50 after 24 h indicated a correlation between functional cellular and in vivo assays. These findings suggested that the inhibition of collagen breakdown by cathepsin K inhibitors, such as L-006235, might be useful in osteoporosis and other indications involving bone resorption [1].

Clinical trial: N/A

Reference:
[1] Palmer JT,Bryant C,Wang DX et al.  Design and synthesis of tri-ring P3 benzamide-containing aminonitriles as potent, selective, orally effective inhibitors of cathepsin K. J Med Chem.2005 Dec 1;48(24):7520-34.

Response of the rhizosphere prokaryotic community of barley (Hordeum vulgare L.) to elevated atmospheric CO2 concentration in open-top chambers.[Pubmed:28371280]

Microbiologyopen. 2017 Aug;6(4).

The effect of elevated atmospheric CO2 concentration [CO2 ] on the diversity and composition of the prokaryotic community inhabiting the rhizosphere of winter barley (Hordeum vulgare L.) was investigated in a field experiment, using open-top chambers. Rhizosphere samples were collected at anthesis (flowering stage) from six chambers with ambient [CO2 ] (approximately 400 ppm) and six chambers with elevated [CO2 ] (700 ppm). The V4 region of the 16S rRNA gene was PCR-amplified from the extracted DNA and sequenced on an Illumina MiSeq instrument. Above-ground plant biomass was not affected by elevated [CO2 ] at anthesis, but plants exposed to elevated [CO2 ] had significantly higher grain yield. The composition of the rhizosphere prokaryotic communities was very similar under ambient and elevated [CO2 ]. The dominant taxa were Bacteroidetes, Actinobacteria, Alpha-, Gamma-, and Betaproteobacteria. Elevated [CO2 ] resulted in lower prokaryotic diversity in the rhizosphere, but did not cause a significant difference in community structure.

A protective role for nitric oxide and salicylic acid for arsenite phytotoxicity in rice (Oryza sativa L.).[Pubmed:28371690]

Plant Physiol Biochem. 2017 Jun;115:163-173.

Nitric oxide (NO) and salicylic acid (SA) are important signaling molecules in plant system. In the present study both NO and SA showed a protective role against arsenite (As(III)) stress in rice plants when supplied exogenously. The application of NO and SA alleviated the negative impact of As(III) on plant growth. Nitric oxide supplementation to As(III) treated plants greatly decreased arsenic (As) accumulation in the roots as well as shoots/roots translocation factor. Arsenite exposure in plants decreased the endogenous levels of NO and SA. Exogenous supplementation of SA not only enhanced endogenous level of SA but also the level of NO through enhanced nitrate reductase (NR) activity, whether As(III) was present or not. Exogenously supplied NO decreased the NR activity and level of endogenous NO. Arsenic accumulation was positively correlated with the expression level of OsLsi1, a transporter responsible for As(III) uptake. The endogenous level of NO and SA were positively correlated to each other either when As(III) was present or not. This close relationship indicates that NO and SA work in harmony to modulate the signaling response in As(III) stressed plants.

Nitric oxide inhibits aluminum-induced programmed cell death in peanut (Arachis hypoganea L.) root tips.[Pubmed:28371714]

J Hazard Mater. 2017 Jul 5;333:285-292.

It had been reported that Aluminum (Al) stress altered nitric oxide (NO) concentration and induced programmed cell death (PCD) in plants. However, the relationship between NO and PCD occurrence under Al stress is unclear. The results showed that cell death induced by Al was significant negative correlation with the inhibition of Al on root elongation growth in peanut. AlCl3 at 100mumolL(-1) induced DNA ladder, chromatin condensation, typical apoptotic chromatin condensation staining with DAPI, apoptosis related gene Hrs203j expression and caspase3-like protease activation in peanut root tip cells, and showed that Al-induced cell death in peanut root tip cells was a typical PCD. Exogenous NO donor sodium nitroprusside (SNP) at 200mumolL(-1) inhibited Al-induced PCD occurrence, but NO specific scavenger cPTIO aggravated PCD production. It suggests that NO is a negative regulator of Al-induced PCD in peanut root tips.

Changes in ABA, IAA and JA levels during calyx, fruit and leaves development in cape gooseberry plants (Physalis peruviana L.).[Pubmed:28371691]

Plant Physiol Biochem. 2017 Jun;115:174-182.

Changes in abscisic acid (ABA), indole-3-acetic acid (IAA) and jasmonic acid (JA) content in developing calyx, fruits and leaves of Physalis peruviana L. plants were analysed. Plant hormones have been widely studied for their roles in the regulation of various aspects related to plant development and, in particular, into their action during development and ripening of fleshly fruits. The obtained evidences suggest that the functions of these hormones are no restricted to a particular development stage, and more than one hormone is involved in controlling various aspects of plant development. Our results will contribute to understand the role of these hormones during growth and development of calyx, fruits and leaves in cape gooseberry plants. This work offers a good, quickly and efficiently protocol to extract and quantify simultaneously ABA, IAA and JA in different tissues of cape gooseberry plants.

Effect of cathepsin k inhibitor basicity on in vivo off-target activities.[Pubmed:17940194]

Mol Pharmacol. 2008 Jan;73(1):147-56.

Cathepsin K is a lysosomal cysteine protease that is a pharmacological target for the treatment of osteoporosis. Previous studies showed that basic, lipophilic cathepsin K inhibitors are lysosomotropic and have greater activities in cell-based assays against cathepsin K, as well as the physiologically important lysosomal cysteine cathepsins B, L, and S, than expected based on their potencies against these isolated enzymes. Long-term administration of the basic cathepsin K inhibitors N-(1-(((cyanomethyl)amino)carbonyl)cyclohexyl)-4-(2-(4-methyl-piperazin-1-yl)-1,3 -thiazol-4-yl)benzamide (L-006235) and balicatib to rats at a supratherapeutic dose of 500 mg/kg/day for 4 weeks resulted in increased tissue protein levels of cathepsin B and L but had no effect on cathepsin B and L message. This is attributed to the inhibitor engagement of these off-target enzymes and their stabilization to proteolytic degradation. No such increase in these tissue cathepsins was detected at the same dose of N-(cyanomethyl)-N(2)-{(1S)-2,2,2-trifluoro-1-[4'-methylsulfonyl)biphenyl-4-yl]eth yl}-l-leucinamide (L-873724), a potent nonbasic cathepsin K inhibitor with a similar off-target profile, although all three inhibitors provided similar plasma exposures. Using an activity-based probe, (125)I-BIL-DMK, in vivo inhibition of cathepsins B, L, and S was detected in tissues of mice given a single oral dose of L-006235 and balicatib, but not in mice given L-873724. In each case, similar tissue levels were achieved by all three compounds, thereby demonstrating the in vivo cathepsin selectivity of L-873724. In conclusion, basic cathepsin K inhibitors demonstrate increased off-target cysteine cathepsin activities than their nonbasic analogs and potentially have a greater risk of adverse effects associated with inhibition of these cathepsins.

Lysosomotropism of basic cathepsin K inhibitors contributes to increased cellular potencies against off-target cathepsins and reduced functional selectivity.[Pubmed:16302795]

J Med Chem. 2005 Dec 1;48(24):7535-43.

The lysosomal cysteine protease cathepsin K is a target for osteoporosis therapy. The aryl-piperazine-containing cathepsin K inhibitor CRA-013783/L-006235 (1) displays greater than 4000-fold selectivity against the lysosomal/endosomal antitargets cathepsin B, L, and S. However, 1 and other aryl-piperazine-containing analogues, including balicatib (10), are approximately 10-100-fold more potent in cell-based enzyme occupancy assays than against each purified enzyme. This phenomenon arises from their basic, lipophilic nature, which results in lysosomal trapping. Consistent with its lysosomotropic nature, 1 accumulates in cells and in rat tissues of high lysosome content. In contrast, nonbasic aryl-morpholino-containing analogues do not exhibit lysosomotropic properties. Increased off-target activities of basic cathepsin K inhibitors were observed in a cell-based cathepsin S antigen presentation assay. No potency increases of basic inhibitors in a functional cathepsin K bone resorption whole cell assay were detected. Therefore, basic cathepsin K inhibitors, such as 1, suffer from reduced functional selectivities compared to those predicted using purified enzyme assays.

Design and synthesis of tri-ring P3 benzamide-containing aminonitriles as potent, selective, orally effective inhibitors of cathepsin K.[Pubmed:16302794]

J Med Chem. 2005 Dec 1;48(24):7520-34.

We have prepared a series of achiral aminoacetonitriles, bearing tri-ring benzamide moieties and an aminocyclohexanecarboxylate residue at P2. This combination of binding elements resulted in sub-250 pM, reversible, selective, and orally bioavailable cathepsin K inhibitors. Lead compounds displayed single digit nanomolar inhibition in vitro (of rabbit osteoclast-mediated degradation of bovine bone). The best compound in this series, 39n (CRA-013783/L-006235), was orally bioavailable in rats, with a terminal half-life of over 3 h. 39n was dosed orally in ovariectomized rhesus monkeys once per day for 7 days. Collagen breakdown products were reduced by up to 76% dose-dependently. Plasma concentrations of 39n above the bone resorption IC50 after 24 h indicated a correlation between functional cellular and in vivo assays. Inhibition of collagen breakdown by cathepsin K inhibitors suggests this mechanism of action may be useful in osteoporosis and other indications involving bone resorption.