Discovery and Preclinical Pharmacology of NX-2127, an Orally Bioavailable Degrader of Bruton's Tyrosine Kinase with Immunomodulatory Activity for the Treatment of Patients with B Cell Malignancies.

Bruton's tyrosine kinase (BTK), a member of the TEC family of kinases, is an essential effector of B-cell receptor (BCR) signaling. Chronic activation of BTK-mediated BCR signaling is a hallmark of many hematological malignancies, which makes it an attractive therapeutic target. Pharmacological inhibition of BTK enzymatic function is now a well-proven strategy for the treatment of patients with these malignancies. We report the discovery and characterization of NX-2127, a BTK degrader with concomitant immunomodulatory activity. By design, NX-2127 mediates the degradation of transcription factors IKZF1 and IKZF3 through molecular glue interactions with the cereblon E3 ubiquitin ligase complex. NX-2127 degrades common BTK resistance mutants, including BTKC481S. NX-2127 is orally bioavailable, exhibits in vivo degradation across species, and demonstrates efficacy in preclinical oncology models. NX-2127 has advanced into first-in-human clinical trials and achieves deep and sustained degradation of BTK following daily oral dosing at 100 mg.

NRX-0492 degrades wild-type and C481 mutant BTK and demonstrates in vivo activity in CLL patient-derived xenografts.

Bruton tyrosine kinase (BTK) is essential for B-cell receptor (BCR) signaling, a driver of chronic lymphocytic leukemia (CLL). Covalent inhibitors bind C481 in the active site of BTK and have become a preferred CLL therapy. Disease progression on covalent BTK inhibitors is commonly associated with C481 mutations. Here, we investigated a targeted protein degrader, NRX-0492, that links a noncovalent BTK-binding domain to cereblon, an adaptor protein of the E3 ubiquitin ligase complex. NRX-0492 selectively catalyzes ubiquitylation and proteasomal degradation of BTK. In primary CLL cells, NRX-0492 induced rapid and sustained degradation of both wild-type and C481 mutant BTK at half maximal degradation concentration (DC50) of ≤0.2 nM and DC90 of ≤0.5 nM, respectively. Sustained degrader activity was maintained for at least 24 hours after washout and was equally observed in high-risk (deletion 17p) and standard-risk (deletion 13q only) CLL subtypes. In in vitro testing against treatment-naïve CLL samples, NRX-0492 was as effective as ibrutinib at inhibiting BCR-mediated signaling, transcriptional programs, and chemokine secretion. In patient-derived xenografts, orally administered NRX-0492 induced BTK degradation and inhibited activation and proliferation of CLL cells in blood and spleen and remained efficacious against primary C481S mutant CLL cells collected from a patient progressing on ibrutinib. Oral bioavailability, >90% degradation of BTK at subnanomolar concentrations, and sustained pharmacodynamic effects after drug clearance make this class of targeted protein degraders uniquely suitable for clinical translation, in particular as a strategy to overcome BTK inhibitor resistance. Clinical studies testing this approach have been initiated (NCT04830137, NCT05131022).

Discovery of Mcl-1-specific inhibitor AZD5991 and preclinical activity in multiple myeloma and acute myeloid leukemia.

Mcl-1 is a member of the Bcl-2 family of proteins that promotes cell survival by preventing induction of apoptosis in many cancers. High expression of Mcl-1 causes tumorigenesis and resistance to anticancer therapies highlighting the potential of Mcl-1 inhibitors as anticancer drugs. Here, we describe AZD5991, a rationally designed macrocyclic molecule with high selectivity and affinity for Mcl-1 currently in clinical development. Our studies demonstrate that AZD5991 binds directly to Mcl-1 and induces rapid apoptosis in cancer cells, most notably myeloma and acute myeloid leukemia, by activating the Bak-dependent mitochondrial apoptotic pathway. AZD5991 shows potent antitumor activity in vivo with complete tumor regression in several models of multiple myeloma and acute myeloid leukemia after a single tolerated dose as monotherapy or in combination with bortezomib or venetoclax. Based on these promising data, a Phase I clinical trial has been launched for evaluation of AZD5991 in patients with hematological malignancies (NCT03218683).

Structure-Based Design of Selective Noncovalent CDK12 Inhibitors.

Cyclin-dependent kinase (CDK) 12 knockdown via siRNA decreases the transcription of DNA-damage-response genes and sensitizes BRCA wild-type cells to poly(ADP-ribose) polymerase (PARP) inhibition. To recapitulate this effect with a small molecule, we sought a potent, selective CDK12 inhibitor. Crystal structures and modeling informed hybridization between dinaciclib and SR-3029, resulting in lead compound 5 [(S)-2-(1-(6-(((6,7-difluoro-1H-benzo[d]imidazol-2-yl)methyl)amino)-9-ethyl-9H-purin-2-yl)piperidin-2-yl)ethan-1-ol]. Further structure-guided optimization delivered a series of selective CDK12 inhibitors, including compound 7 [(S)-2-(1-(6-(((6,7-difluoro-1H-benzo[d]imidazol-2-yl)methyl)amino)-9-isopropyl-9H-purin-2-yl)piperidin-2-yl)ethan-1-ol]. Profiling of this compound across CDK9, 7, 2, and 1 at high ATP concentration, single-point kinase panel screening against 352 targets at 0.1 µm, and proteomics via kinase affinity matrix technology demonstrated the selectivity. This series of compounds inhibits phosphorylation of Ser2 on the C-terminal repeat domain of RNA polymerase II, consistent with CDK12 inhibition. These selective compounds were also acutely toxic to OV90 as well as THP1 cells.

Correction to "Structure Based Design of Non-Natural Peptidic Macrocyclic Mcl-1 Inhibitors".

[This corrects the article DOI: 10.1021/acsmedchemlett.6b00464.].

Volume Load and Neuromuscular Fatigue During an Acute Bout of Agonist-Antagonist Paired-Set vs. Traditional-Set Training.

The purpose of this study was to investigate the acute effects of performing paired-set (PS) vs. traditional-set (TS) training over 3 consecutive sets, on volume load and electromyographic fatigue parameters of the latissimus dorsi, biceps brachii, pectoralis major, and triceps brachii muscles. Fifteen trained men performed 2 testing protocols (TS and PS) using 10 repetition maximum loads. The TS protocol consisted of 3 sets of bench press (BP) followed by 3 sets of wide-grip seated row (SR). The PS consisted of 3 sets of BP and 3 sets of SR performed in an alternating manner. Volume load was calculated as load × repetitions. The electromyographic signal, time (CRMS) and frequency (Cf5) domain, parameters were recorded during SR. Under the PS protocol, sets of SR were performed immediately after the sets of BP. A 2-minute rest interval between the completion of the set of SR and the subsequent set of BP was implemented (e.g., between PSs). Under the TS protocol, 2-minute rest intervals were implemented between all sets. BP and SR volume loads decreased significantly from set 1 to set 2 and from set 2 to set 3 under both conditions. Volume load was greater for all sets of both exercises under PS as compared with TS. Muscle fatigue indices were greater under PS as compared with TS. In general, these results indicate that as compared with TS, PS produced a greater training volume in less time and may induce greater fatigue and thereby provide an enhanced training stimulus.

Structure Based Design of Non-Natural Peptidic Macrocyclic Mcl-1 Inhibitors.

Mcl-1 is a pro-apoptotic BH3 protein family member similar to Bcl-2 and Bcl-xL. Overexpression of Mcl-1 is often seen in various tumors and allows cancer cells to evade apoptosis. Here we report the discovery and optimization of a series of non-natural peptide Mcl-1 inhibitors. Screening of DNA-encoded libraries resulted in hit compound 1, a 1.5 µM Mcl-1 inhibitor. A subsequent crystal structure demonstrated that compound 1 bound to Mcl-1 in a ß-turn conformation, such that the two ends of the peptide were close together. This proximity allowed for the linking of the two ends of the peptide to form a macrocycle. Macrocyclization resulted in an approximately 10-fold improvement in binding potency. Further exploration of a key hydrophobic interaction with Mcl-1 protein and also with the moiety that engages Arg256 led to additional potency improvements. The use of protein-ligand crystal structures and binding kinetics contributed to the design and understanding of the potency gains. Optimized compound 26 is a <3 nM Mcl-1 inhibitor, while inhibiting Bcl-2 at only 5 µM and Bcl-xL at >99 µM, and induces cleaved caspase-3 in MV4-11 cells with an IC50 of 3 µM after 6 h.

Catalytic enantioselective addition of organoboron reagents to fluoroketones controlled by electrostatic interactions.

Organofluorine compounds are central to modern chemistry, and broadly applicable transformations that generate them efficiently and enantioselectively are in much demand. Here we introduce efficient catalytic methods for the addition of allyl and allenyl organoboron reagents to fluorine-substituted ketones. These reactions are facilitated by readily and inexpensively available catalysts and deliver versatile and otherwise difficult-to-access tertiary homoallylic alcohols in up to 98% yield and >99:1 enantiomeric ratio. Utility is highlighted by a concise enantioselective approach to the synthesis of the antiparasitic drug fluralaner (Bravecto, presently sold as the racemate). Different forms of ammonium-organofluorine interactions play a key role in the control of enantioselectivity. The greater understanding of various non-bonding interactions afforded by these studies should facilitate the future development of transformations that involve fluoroorganic entities.

Highly Chemoselective Iridium Photoredox and Nickel Catalysis for the Cross-Coupling of Primary Aryl Amines with Aryl Halides.

A visible-light-promoted iridium photoredox and nickel dual-catalyzed cross-coupling procedure for the formation C-N bonds has been developed. With this method, various aryl amines were chemoselectively cross-coupled with electronically and sterically diverse aryl iodides and bromides to forge the corresponding C-N bonds, which are of high interest to the pharmaceutical industries. Aryl iodides were found to be a more efficient electrophilic coupling partner. The coupling reactions were carried out at room temperature without the rigorous exclusion of molecular oxygen, thus making this newly developed Ir-photoredox/Ni dual-catalyzed procedure very mild and operationally simple.

Practical and Broadly Applicable Catalytic Enantioselective Additions of Allyl-B(pin) Compounds to Ketones and α-Ketoesters.

A set of broadly applicable methods for efficient catalytic additions of easy-to-handle allyl-B(pin) (pin=pinacolato) compounds to ketones and acyclic α-ketoesters was developed. Accordingly, a large array of tertiary alcohols can be obtained in 60 to >98 % yield and up to 99:1 enantiomeric ratio. At the heart of this development is rational alteration of the structures of the small-molecule aminophenol-based catalysts. Notably, with ketones, increasing the size of a catalyst moiety (tBu to SiPh3 ) results in much higher enantioselectivity. With α-ketoesters, on the other hand, not only does the opposite hold true, since Me substitution leads to substantially higher enantioselectivity, but the sense of the selectivity is reversed as well.

Photoredox Mediated Nickel Catalyzed Cross-Coupling of Thiols With Aryl and Heteroaryl Iodides via Thiyl Radicals.

Ni-catalyzed cross-couplings of aryl, benzyl, and alkyl thiols with aryl and heteroaryl iodides were accomplished in the presence of an Ir-photoredox catalyst. Highly chemoselective C-S cross-coupling was achieved versus competitive C-O and C-N cross-couplings. This C-S cross-coupling method exhibits remarkable functional group tolerance, and the reactions can be carried out in the presence of molecular oxygen. Mechanistic investigations indicated that the reaction proceeded through transient Ni(I)-species and thiyl radicals. Distinct from nickel-catalyzed cross-coupling reactions involving carbon-centered radicals, control experiments and spectroscopic studies suggest that this C-S cross-coupling reaction does not involve a Ni(0)-species.

Effects of Molecular Oxygen, Solvent, and Light on Iridium-Photoredox/Nickel Dual-Catalyzed Cross-Coupling Reactions.

In order to achieve reproducibility during iridium-photoredox and nickel dual-catalyzed sp(3)-sp(2) carbon-carbon bond-forming reactions, we investigated the role that molecular oxygen (O2), solvent and light-source (CF lamp or blue LED) play in a variety of Ir-photoredox mediated transformations. The presence of O2 was discovered to be important for catalyst activation when air-stable Ni(II) precatalysts were used in DMF under CF lamp irradiation; however, O2 was not required for catalysis when conducted with Ni(COD)2 in the same reaction system. O2 is believed to promote rapid reduction of the Ni(II) precatalyst by Ir(II) to Ni(0). In addition to O2, the effects that solvent and light-source have on the dual-catalyzed decarboxylative cross-coupling reactions will be discussed. These findings have enabled us to develop a more robust dual-catalyzed decarboxylative cross-coupling protocol.

Changes in the athletic profile of elite college American football players.

The purpose of this study was to compare positional anthropometric and National Football League (NFL) Combine performance levels in elite college American football players over the 3-year period from 1999 to 2001 to the 3-year period from 2008 to 2010. The sample included 15 offensive and defensive positions, and only those players invited to the combine and subsequently drafted in the same year (n = 1,712) were included in the study. Data from 10 combine physical tests were examined, including weight; height; the 9.1-, 18.3-, and 36.6-m sprints; the vertical and horizontal jumps; the 18.3-m shuttle run; the 3-cone drill; and the 102.1-kg bench press for maximum repetitions. Independent samples t-tests detected differences for each of the 15 positions (p < 0.05). There were no discernible trends in height and weight over the period in question, whereas players in the more recent group significantly improved performance in straight sprinting, the 3-cone drill, and the horizontal jump. Findings suggest that these tests better reflect characteristics such as explosiveness and first-step quickness as compared with the 18.3-m shuttle and the vertical jump, and that such characteristics have become more highly sought after by NFL coaches and scouts. The results of the present research suggest that the position-specific profiles changed over a relatively short period of time. Coaches and practitioners will be able to use the findings of this research to better prepare athletes for entry into the NFL.

A C-H borylation approach to Suzuki-Miyaura coupling of typically unstable 2-heteroaryl and polyfluorophenyl boronates.

A method for the synthesis of biaryls and heterobiaryls from arenes and haloarenes without the intermediacy of unstable boronic acids is described. Pinacol boronate esters that are analogous to unstable boronic acids are formed in high yield by iridium-catalyzed C-H borylation of heteroarenes and fluoroarenes. These boronates are stable in the solid state or in solution and can be generated and used in situ. They couple with aryl halides in the presence of simple palladium catalysts, providing a convenient route to biaryl and heteroaryl products that have been challenging to prepare via boronic acids.

The normalization of explosive functional movements in a diverse population of elite American football players.

The objective of this study was to investigate the need to normalize, for body mass, explosive functional tasks in a population exhibiting diverse body masses. Measures investigated in elite college American football players attending the National Football League's annual combine (n = 1,136) were the 9.1-, 18.3-, and 36.6-m sprints, vertical and horizontal jumps, 18.3-m shuttle, and 3-cone drill. To determine the relationship between body mass and performance outcomes, Pearson's correlation coefficients (r) were generated using log-transformed data. Task-specific allometric exponents, accounting for body mass, were also determined. The strength of the correlations suggests that sprint and jump abilities are associated with body mass, whereas change-of-direction ability is not. The determined allometric exponents range between 0.296 and -0.463 for the sprint and jump tasks and are -0.022 and -0.006 for the 18.3-m shuttle and the 3-cone drill, respectively. In populations exhibiting relatively large variations in body mass, normalization of sprint and jump abilities is recommended, whereas normalization of change-of-direction ability is unwarranted. Novel suggestions derived from the present research are that sprint and jump abilities in diverse populations warrant normalization and that physical attributes associated with explosive functional movements deserve attribute-specific consideration when contemplating normalization.

Movement demands in Australian rules football as indicators of muscle damage.

The purpose of this study was to determine if there is an association between variables that describe movements in an Australian Rules football (ARF) game with muscle damage. Fourteen elite junior ARF players were monitored with a global positioning system (GPS) during a match, and muscle damage was estimated by determining creatine kinase (CK) 24 hours postmatch. The players were median split based on CK levels, into a high and low CK group, and the groups were compared with independent t-tests. The primary finding was that the group that experienced greater muscle damage (high CK group) generally covered significantly (p < 0.05) greater distances. This was the case for running speeds between 4 and 7 m·s(-1) and, with the exception of high acceleration, all intensities of acceleration and deceleration. The high, as compared with the low, CK group also produced a significantly greater (42%) "player load." All of these significant differences were accompanied by large effect sizes. Group-specific Pearson (r) correlations between CK level and GPS variables suggest that a certain volume of movement is required before the elicitation of a positive relationship beyond trivial or small. Correlations between CK and running speeds >4 m·s(-1) and moderate-high acceleration and deceleration were negative in the low CK (lesser volumes) group. With the exception of low-intensity acceleration/deceleration, the same relationships were positive and generally of a moderate-to-large magnitude in the high CK (greater volumes) group. It may be that a certain volume of movement is required for that movement to be strongly associated with CK levels. It was concluded that selected GPS variables obtained from ARF games can be used as indicators of muscle damage, and this information may be used to individualize recovery strategies after games.

Positional relationships between various sprint and jump abilities in elite American football players.

The purpose of this study was to investigate positional relationships between sprint and jump abilities and body mass in elite college American football players (n = 1,136). Data from the annual National Football League combine over the years 2005-2009 were examined. The measures included for examination were the 9.1-, 18.3-, 36.6-, and flying 18.3-m sprints and the vertical and horizontal jumps. Pearson's correlation coefficients (r) were calculated to determine the relationships between the tests, and coefficients of determination (r2) were used to determine common variance. With the exception of the relationship between the 9.1-m and the flying 18.3-m sprints, the relationships between all sprints are very strong. Vertical jump ability is more strongly associated with maximum speed, as compared with acceleration. Horizontal jump ability is similarly associated with maximum speed and acceleration. The 9.1-, 18.3-, and flying 18.3-m sprints and the jump tests would appear to measure independent skills. Stationary start sprints up to 36.6 m appear to be heavily influenced by acceleration and may thus measure similar characteristics. The flying 18.3-m sprint is recommended as a measure of maximum speed. Body mass was most strongly associated with performance in the lineman group. When body mass was controlled for, correlations weakened across all the groups. The role of body mass remains unclear. Regardless of sport, the present research supports the notion that the relationships between various sprint and jump abilities warrant positional consideration. Coaches and practitioners will be able to use the findings of this research to better test and monitor athletes requiring different skills.

Relationships between National Football League combine performance measures.

The purpose of this study was to investigate the relationships between the athletic skills measured at the National Football League (NFL) combine. The combine comprises the following tests: 36.6-m sprint with split times at 9.1 and 18.3 m, vertical and horizontal jumps, 18.3-m shuttle run, 3-cone drill, and 102.1-kg bench press. Draftees to the NFL who participated in the annual combine from 2005 to 2009 were included in the study (n = 1,136). Pearson's (r) correlations were calculated to determine the relationships between the tests, and coefficients of determination (r) were used to determine common variance. The 9.1-, 18.3-, and 36.6-m sprint times are nearly perfectly correlated (r ranges from 0.900 to 0.967) as are the change-of-direction ability tests, 18.3-m shuttle run, and 3-cone drill (r = 0.948), suggesting similar skills are being measured. Performance in both jumping tasks is more strongly associated with longer sprint distances, suggesting mechanisms such as the stretch-shortening cycle may be more important at maximal, or near-maximal, speeds. The correlations between change-of-direction ability and sprinting and jumping are generally much weaker (r ranges from 0.250 to -0.653), suggesting less association and independent motor skills. Although not particularly large correlation coefficients, bench press performance is positively correlated with outcomes in all running drills and inversely correlated with jump abilities, suggesting that in the observed cohort, upper body strength may be of little benefit to these tasks. Incorporation of a nonacceleration influenced (i.e., moving start) measure of maximal speed may be preferred if the intention of a test battery is to measure independent motor skills. Further, when constructing test batteries, either the 18.3-m shuttle or 3-cone drill is likely sufficient as a measure of change-of-direction ability. Test batteries should be constructed to measure independent motor skills.

The effect of training volume on lower-body strength.

The objective of this study was to examine the chronic effects on lower-body strength in resistance trained men of performing varying training volumes over 6 weeks. A pretest and posttest design was used to investigate the effects on 1-repetition maximum (1RM) squat strength. Also, 1RM testing was performed at 3 weeks. Participants were randomly assigned to an intensity-matched (80% of 1RM) low (1-SET), moderate (4-SET), or high (8-SET) volume condition. In addition to significant strength increases in all groups at the end of the 6-week period, increases were observed at 3 weeks under the 4- and 8-SET conditions, which were greater than the improvement under the 1-SET condition. At 6 weeks, the magnitude of improvement was significantly greater for the 8-SET, as compared with that of the 1-SET group. The magnitude of improvement elicited in the 4-SET group was not different from that of the 1-SET or 8-SET groups. The results suggest that "high" volumes (i.e., >4 sets) are associated with enhanced strength development but that "moderate" volumes offer no advantage. Practitioners should be aware that strength development may be dependent on appropriate volume doses and training duration.