ATP-competitive Kinase Inhibitors Research Articles

Therapy for patients with chronic phase-chronic myeloid leukemia previously treated with ⩾2 tyrosine kinase inhibitors: a systematic literature review.

ATP-competitive tyrosine kinase inhibitors (TKIs) are the current standard of care for patients with chronic phase-chronic myeloid leukemia (CP-CML) in the first-line and second-line (2 L) setting. Treatment after 2 L is not clearly established. The objective of this study was to summarize the available evidence to compare the efficacy and safety of interventions in the treatment of CP-CML patients who had received ⩾2 prior TKIs. A systematic literature review was performed. A systematic literature review (SLR) of studies published until May 2021, reporting clinical outcomes in adult patients with CP-CML who had received ⩾ 2 prior TKIs was performed. Studies were identified through the database searches via Ovid platform (Embase, MEDLINE Epub Ahead of Print, In-Process and Other Non-Indexed Citations, and Cochrane Central Register of Controlled Trials), the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP), bibliographic search of relevant reviews, and proceedings from the previous 3 years of the key conferences in the field of oncology. Our search identified 38 relevant studies. Among the identified studies of the current third-line treatments, the major molecular response (MMR) rate for ponatinib was 19.0-66.7%, 23.3-25.5% for asciminib, 19.2% for omacetaxine, and 13.2% for bosutinib at 6 months. The complete cytogenetic response (CCyR) rate was 21.4-64.8% for ponatinib, 38.7-40.8% for asciminib, 18-24.2% for bosutinib, and 16.1% for omacetaxine at 6 months. The findings from current SLR demonstrated the lack of data for patients with CML treated with ⩾2 TKIs. TKIs such as asciminib, ponatinib, and bosutinib are valid options for those patients. Further research is needed to identify the best treatment option for patients with CML receiving later lines of therapy.

A cell-based chemical-genetic screen for amino acid stress response inhibitors reveals torins reverse stress kinase GCN2 signaling

mTORC1 and GCN2 are serine/threonine kinases that control how cells adapt to amino acid availability. mTORC1 responds to amino acids to promote translation and cell growth while GCN2 senses limiting amino acids to hinder translation via eIF2α phosphorylation. GCN2 is an appealing target for cancer therapies because malignant cells can harness the GCN2 pathway to temper the rate of translation during rapid amino acid consumption. To isolate new GCN2 inhibitors, we created cell-based, amino acid limitation reporters via genetic manipulation of Ddit3 (encoding the transcription factor CHOP). CHOP is strongly induced by limiting amino acids and in this context, GCN2-dependent. Using leucine starvation as a model for essential amino acid sensing, we unexpectedly discovered ATP-competitive PI3 kinase-related kinase inhibitors, including ATR and mTOR inhibitors like torins, completely reversed GCN2 activation in a time-dependent way. Mechanistically, via inhibiting mTORC1-dependent translation, torins increased intracellular leucine, which was sufficient to reverse GCN2 activation and the downstream integrated stress response including stress-induced transcriptional factor ATF4 expression. Strikingly, we found that general translation inhibitors mirrored the effects of torins. Therefore, we propose that mTOR kinase inhibitors concurrently inhibit different branches of amino acid sensing by a dual mechanism involving direct inhibition of mTOR and indirect suppression of GCN2 that are connected by effects on the translation machinery. Collectively, our results highlight distinct ways of regulating GCN2 activity.

CML-466 Asciminib Provides Durable Molecular Responses in Patients With Chronic Myeloid Leukemia in Chronic Phase (CML-CP) With the T315I Mutation: Updated Efficacy and Safety Data From a Phase I Trial

In CML-CP, the BCR::ABL1 T315I mutation confers resistance to previously approved ATP-competitive tyrosine kinase inhibitors (TKIs), except ponatinib and olverembatinib. In a previous analysis of the phase I, dose-escalation trial X2101, asciminib-a BCR::ABL1 inhibitor that binds to the ABL myristoyl pocket-demonstrated efficacy and a favorable safety profile in heavily pretreated patients with T315I-mutated CML. We report updated efficacy and safety data in patients with CML-CP with the T315I mutation (data cutoff: January 6, 2021). Provide updated safety and efficacy data for patients with T315I-mutated CML-CP after added exposure. Patients with T315I-mutated CML-CP and treated with ≥1 prior TKI were enrolled and received asciminib 200mg twice daily (BID). 48 patients were included; 25 patients (52.1%) received ≥3 prior TKIs. At data cutoff, treatment was ongoing in 27 patients (56.3%). 45 of 48 patients were evaluable (BCR::ABL1IS >0.1% at baseline) for major molecular response (MMR); 3 were excluded for BCR::ABL1 atypical transcripts. Among evaluable patients, 19 (42.2%) achieved MMR by week 24 and 22 (48.9%) by week 96. Evaluable patients included 26 ponatinib-pretreated and 19 ponatinib-naive patients; 34.6% and 68.4%, respectively, achieved MMR by week 96. The probability of maintaining MMR for ≥96 weeks was 84% (95% CI, 68.1%-100.0%). 23 of 37 patients (62.2%) with BCR::ABL1IS >1% at baseline achieved BCR::ABL1IS ≤1% by week 96. The safety/tolerability profile of asciminib remained favorable after ≈9 months of added exposure (median duration of exposure, 2.08 years; range, 0.04-4.13 years). The most common (≥10%) grade ≥3 adverse events (AEs) were lipase increase (18.8%, all asymptomatic elevations) and thrombocytopenia (14.6%). Arterial occlusive events occurred in 4 patients (8.3%); none led to dose adjustment/interruption/discontinuation. AEs leading to discontinuation occurred in 5 patients (10.4%). Only 2 study deaths, both due to COVID-19, occurred in this patient population. After a median exposure of >2 years, asciminib monotherapy 200mg BID exhibited a sustained, favorable safety profile and clinical efficacy in patients with T315I-mutated CML-CP-a population with high unmet medical need. This updated analysis confirms asciminib as a treatment option for patients with T315I-mutated CML-CP, including those previously treated with ponatinib.

Advances in RIPK1 kinase inhibitors.

Programmed necrosis is a new modulated cell death mode with necrotizing morphological characteristics. Receptor interacting protein 1 (RIPK1) is a critical mediator of the programmed necrosis pathway that is involved in stroke, myocardial infarction, fatal systemic inflammatory response syndrome, Alzheimer’s disease, and malignancy. At present, the reported inhibitors are divided into four categories. The first category is the type I ATP-competitive kinase inhibitors that targets the area occupied by the ATP adenylate ring; The second category is type Ⅱ ATP competitive kinase inhibitors targeting the DLG-out conformation of RIPK1; The third category is type Ⅲ kinase inhibitors that compete for binding to allosteric sites near ATP pockets; The last category is others. This paper reviews the structure, biological function, and recent research progress of receptor interaction protein-1 kinase inhibitors.

Protein kinase inhibitor ceritinib blocks ectonucleotidase CD39 – a promising target for cancer immunotherapy

BackgroundAn important mechanism, by which cancer cells achieve immune escape, is the release of extracellular adenosine into their microenvironment. Adenosine activates adenosine A2A and A2B receptors on immune cells constituting...

English

We set up six light conditions to investigate the changes in the development of Arabidopsis thaliana hypocotyls and roots. Seedlings grown for 96 h under darkness were scored with shorter roots and longer hypocotyls. In shoot-shaded conditions, seedlings were unable to carry out photosynthesis, resulting in insufficient stored nutrients for root development. In the three groups of different light intensities applied to the roots, total light caused stress in the entire seedlings and the length of roots and hypocotyls were shorter than in conditions when roots were growing within light-dark gradients. Importantly, root lengths were higher within light-dark gradients than in total light. Different light treatments did significantly affect root growth and hypocotyl growth. The addition of ATP-competitive mTOR kinase inhibitor (AZD), drastically reduced root, however, this did not occur with hypocotyl length.   Key words: Total light, total dark, gradient light, shoot dark with light blocker, light blocker, shoot dark.

P704: ASCIMINIB PROVIDES DURABLE MOLECULAR RESPONSES IN PATIENTS (PTS) WITH CHRONIC MYELOID LEUKEMIA IN CHRONIC PHASE (CML-CP) WITH THE T315I MUTATION: UPDATED EFFICACY AND SAFETY DATA FROM A PHASE I TRIAL

Background: In pts with CML, the BCR::ABL1 T315I mutation is associated with poor clinical outcomes and confers resistance to previously approved ATP-competitive tyrosine kinase inhibitors (TKIs). Until recently, ponatinib (PON) was the only TKI available for these pts, but its use may be limited by associated cardiovascular events. In the primary analysis of the phase I trial X2101, asciminib—the 1st BCR::ABL1 inhibitor to Specifically Target the ABL Myristoyl Pocket (STAMP)—demonstrated efficacy and a favorable safety profile in heavily pretreated pts with CML with T315I. These results supported the FDA approval of asciminib as a new treatment option for pts with CML-CP with T315I (NCCN 2021). We report updated efficacy and safety data in these pts (data cutoff: January 6, 2021). Aims: Provide updated safety and efficacy data for pts with CML-CP with T315I treated with asciminib monotherapy 200 mg twice daily (BID) after added exposure. Methods: Pts with CML-CP with T315I were enrolled if treated with ≥1 prior TKI and no other effective therapy was available, provided informed consent, and received asciminib 200 mg BID. Results: 48 pts with T315I were included; 2 (4.2%) pts had additional BCR::ABL1 mutations at baseline. Eight (16.7%), 15 (31.3%) and 25 (52.1%) pts received 1, 2, and ≥3 prior TKIs, respectively. At data cutoff, treatment was ongoing in more than half (27 [56.3%]) of pts; the predominant reason for treatment discontinuation was physician’s decision (11 [22.9%]), mainly due to lack of efficacy. Of the 48 pts, 45 were evaluable (BCR::ABL1IS >0.1% at baseline) for major molecular response (MMR); 3 were excluded for BCR::ABL1 atypical transcripts. Among evaluable pts, 19 (42.2%) achieved MMR by wk 24 and 22 (48.9%) by wk 96; 19 were still in MMR at the cutoff date. Evaluable pts included 26 PON-pretreated and 19 PON-naive pts; 34.6% and 68.4%, respectively, achieved MMR by the cutoff date (Table). The probability of pts maintaining MMR for ≥96 wks was 84% (95% CI, 68.1-100.0). Thirteen (28.9%) and 11 (24.4%) pts achieved MR4 and MR4.5, respectively. Twenty (54.1%) and 23 (62.2%) of 37 pts with BCR::ABL1IS >1% at baseline achieved BCR::ABL1IS ≤1% by wk 48 and 96, respectively. The median duration of exposure was 2.08 (range, 0.04-4.13) yrs with more than half (27 [56.3%]) of pts receiving treatment for ≥96 wks; the median daily dose intensity was 398.3 (range, 179-400) mg/day. The safety/tolerability profile of asciminib remained favorable after ≈9 months of added follow-up (Table). The most common (≥5%) grade ≥3 adverse events (AEs) were lipase increase (18.8%, all asymptomatic elevations), thrombocytopenia (14.6%), and vomiting, ALT increase, abdominal pain, hypertension, anemia, neutropenia, and neutrophil count decrease (6.3% each). Arterial occlusive events occurred in 4 (8.3%) pts; none led to dose adjustment/interruption/discontinuation. AEs leading to discontinuation were reported in 2 new pts since the previous data cutoff; both pts discontinued and died due to COVID-19. These were the only study deaths reported in this pt population. Image:Summary/Conclusion: Asciminib monotherapy 200 mg BID exhibited a sustained, favorable safety profile after added exposure with no new safety signals in pts with CML-CP with T315I—a population with high unmet medical need. The clinical efficacy of asciminib is demonstrated by the high proportion of pts achieving durable MMR and BCR::ABL1IS ≤ 1%. The updated analysis confirms asciminib as a treatment option for pts with CML-CP with T315I, including those for whom treatment with PON has failed.

Abstract 5357: mTOR kinase inhibitor AZD8055 reverses acquired rapamycin resistance in prostate cancer cells

Abstract The mTOR pathway is frequently over-activated in human cancers. However, classic allosteric mTOR inhibitors rapamycin only exerts limited clinical benefits. It has been shown that prostate cancer are able to acquire resistance after rapamycin treatment for a period of time. Such secondary resistance remains as major concerns in rapamycin-based anti-cancer therapies that may inevitably lead to therapeutic failure. In the current study, we evaluated treatment effects of ATP-competitive dual mTOR kinase inhibitor AZD8055 on cancer cells that have acquired resistance to rapamycin, and underpinning molecular mechanisms. To establish cells with acquired rapamycin resistance, PTEN-mt prostate cancer PC-3 cells were gradually treated with increasing concentrations of rapamycin (up to 10 μM) until the cells were completely unresponsive to rapamycin (PC-3RR). Cell proliferation was determined by MTT assay and clonogenic survival was tested by colony formation assay. m7G pull-down assay was employed to detect the cap-dependent translation. mTOR downstream pathways, including p-S6K (T389)/p-S6 (S235/236), p-4EBP1 (T37/S46) and p-Akt (S473) were detected by Western Blotting. PC-3RR cells essentially failed to respond to rapamycin, with IC50 values greater than 1 μM. However, AZD8055 suppressed PC-3RR proliferation in a dose dependent manner, with IC50 < 30 nM. Similarly, clonogenic capacity was impaired by AZD8055 to a greater extent than was the case for rapamycin. At the molecular level, AZD8055, but not rapamycin, strongly prevents the dissociation of 4E-BP1 from eIF4E. Further, while the p-S6K and p-S6 pathway was potently inhibited by both agents, AZD8055 displayed more striking inhibition on p-4E-BP1 and p-Akt than rapamycin. Interestingly, similar to rapamycin, AZD8055 slightly increased p-Akt (T308), suggesting the existence of the PI3K-mediated feedback loop. These findings indicate that AZD8055 significantly inhibits malignant functions in cancer cells that are highly resistant to rapamycin. Such effects are likely to be associated with the deactivation of rapamycin-insensitive mTOR downstream pathways. These data provide preliminary evidence for future evaluation of ATP-competitive mTOR inhibitors in cancer models that are resistant to rapamycin analogs. Citation Format: Yao Dai, Dietmar Siemann. mTOR kinase inhibitor AZD8055 reverses acquired rapamycin resistance in prostate cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5357.

C5-Iminosugar modification of casein kinase 1δ lead 3-(4-fluorophenyl)-5-isopropyl-4-(pyridin-4-yl)isoxazole promotes enhanced inhibitor affinity and selectivity.

The quest for isoform-selective and specific ATP-competitive protein kinase inhibitors is of great interest, as inhibitors with these qualities will come with reduced toxicity and improved efficacy. However, creating such inhibitors is very challenging due to the high molecular similarity of kinases ATP active sites. To achieve selectivity for our casein kinase (CK) 1 inhibitor series, we elected to endow our previous CK1δ-hit, 3-(4-fluorophenyl)-5-isopropyl-4-(pyridin-4-yl)isoxazole (1), with chiral iminosugar scaffolds. These scaffolds were attached to C5 of the isoxazole ring, a position deemed favorable to facilitate binding interactions with the ribose pocket/solvent-open area of the ATP binding pocket of CK1δ. Here, we describe the synthesis of analogs of 1 ((-)-/(+)-34, (-)-/(+)-48), which were prepared in 13 steps from enantiomerically pure ethyl (3R,4S)- and ethyl (3S,4R)-1-benzyl-4-[(tert-butyldimethylsilyl)oxy]-5-oxopyrrolidine-3-carboxylate ((-)-11 and (+)-11), respectively. The synthesis involved the coupling of Weinreb amide-activated chiral pyrrolidine scaffolds with 4- and 2-fluoro-4-picoline and reaction of the resulting 4-picolyl ketone intermediates ((-)-/(+)-40 and (-)-/(+)-44) with 4-fluoro-N-hydroxybenzenecarboximidoyl chloride to form the desired isoxazole ring. The activity of the compounds against human CK1δ, -ε, and -α was assessed in recently optimized in vitro assays. Compound (-)-34 was the most active compound with IC50 values (CK1δ/ε) of 1/8 µM and displayed enhanced selectivity toward CK1δ.

GCN2 kinase activation by ATP-competitive kinase inhibitors.

Small-molecule kinase inhibitors represent a major group of cancer therapeutics, but tumor responses are often incomplete. To identify pathways that modulate kinase inhibitor response, we conducted a genome-wide knockout (KO) screen in glioblastoma cells treated with the pan-ErbB inhibitor neratinib. Loss of general control nonderepressible 2 (GCN2) kinase rendered cells resistant to neratinib, whereas depletion of the GADD34 phosphatase increased neratinib sensitivity. Loss of GCN2 conferred neratinib resistance by preventing binding and activation of GCN2 by neratinib. Several other Food and Drug Administration (FDA)-approved inhibitors, such erlotinib and sunitinib, also bound and activated GCN2. Our results highlight the utility of genome-wide functional screens to uncover novel mechanisms of drug action and document the role of the integrated stress response (ISR) in modulating the response to inhibitors of oncogenic kinases.

Elucidating the role of the kinase activity of endothelial cell focal adhesion kinase in angiocrine signalling and tumour growth.

A common limitation of cancer treatments is chemotherapy resistance. We have previously identified that endothelial cell (EC)-specific deletion of focal adhesion kinase (FAK) sensitises tumour cells to DNA-damaging therapies, reducing tumour growth in mice. The present study addressed the kinase activity dependency of EC FAK sensitisation to the DNA-damaging chemotherapeutic drug, doxorubicin. FAK is recognised as a therapeutic target in tumour cells, leading to the development of a range of inhibitors, the majority being ATP competitive kinase inhibitors. We demonstrate that inactivation of EC FAK kinase domain (kinase dead; EC FAK-KD) in established subcutaneous B16F0 tumours improves melanoma cell sensitisation to doxorubicin. Doxorubicin treatment in EC FAK-KD mice reduced the percentage change in exponential B16F0 tumour growth further than in wild-type mice. There was no difference in tumour blood vessel numbers, vessel perfusion or doxorubicin delivery between genotypes, suggesting a possible angiocrine effect on the regulation of tumour growth. Doxorubicin reduced perivascular malignant cell proliferation, while enhancing perivascular tumour cell apoptosis and DNA damage in tumours grown in EC FAK-KD mice 48 h after doxorubicin injection. Human pulmonary microvascular ECs treated with the pharmacological FAK kinase inhibitors defactinib, PF-562,271 or PF-573,228 in combination with doxorubicin also reduced cytokine expression levels. Together, these data suggest that targeting EC FAK kinase activity may alter angiocrine signals that correlate with improved acute tumour cell chemosensitisation. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

Targeting anaplastic lymphoma kinase (ALK) gene alterations in neuroblastoma by using alkylating pyrrole-imidazole polyamides.

Anaplastic lymphoma kinase (ALK) aberration is related to high-risk neuroblastomas and is an important therapeutic target. As acquired resistance to ALK tyrosine kinase inhibitors is inevitable, novel anti-ALK drug development is necessary in order to overcome potential drug resistance against ATP-competitive kinase inhibitors. In this study, to overcome ALK inhibitor resistance, we examined the growth inhibition effects of newly developed ALK-targeting pyrrole-imidazole polyamide CCC-003, which was designed to directly bind and alkylate DNA within the F1174L-mutated ALK gene. CCC-003 suppressed cell proliferation in ALK-mutated neuroblastoma cells. The expression of total and phosphorylated ALK was downregulated by CCC-003 treatment but not by treatment with a mismatch polyamide without any binding motif within the ALK gene region. CCC-003 preferentially bound to the DNA sequence with the F1174L mutation and significantly suppressed tumor progression in a human neuroblastoma xenograft mouse model. Our data suggest that the specific binding of CCC-003 to mutated DNA within the ALK gene exerts its anti-tumor activity through a mode of action that is distinct from those of other ALK inhibitors. In summary, our current study provides evidence for the potential of pyrrole-imidazole polyamide ALK inhibitor CCC-003 for the treatment of neuroblastoma thus offering a possible solution to the problem of tyrosine kinase inhibitor resistance.

ATP-competitive partial antagonists of the IRE1α RNase segregate outputs of the UPR.

The unfolded protein response (UPR) homeostatically matches endoplasmic reticulum (ER) protein-folding capacity to cellular secretory needs. But under high/chronic ER stress, the UPR triggers apoptosis. This dichotomy is promoted by differential activation of the ER transmembrane kinase/endoribonuclease (RNase) IRE1α. We previously found that IRE1α’s RNase can be either fully activated or inactivated by ATP-competitive kinase inhibitors. Here we developed kinase inhibitors—’PAIR’s—Partial Antagonists of IRE1α RNase—that partially antagonize IRE1α’s RNase at full occupancy. Biochemical and structural studies show that PAIRs promote partial RNase antagonism by intermediately displacing the αC helix in IRE1α’s kinase domain. In insulin-producing β-cells, PAIRs permit adaptive XBP1 mRNA splicing, while quelling destructive ER mRNA endonucleolytic decay and apoptosis. By preserving XBP1 mRNA splicing, PAIRs allow B-lymphocytes to differentiate into immunoglobulin-producing plasma cells. Thus, an intermediate RNase-inhibitory “sweet spot”, achieved by PAIR-bound IRE1α captures a desirable conformation for drugging this master UPR sensor/effector.

Identification of unprecedented ATP-competitive choline kinase inhibitors

In this article we describe the identification of unprecedented ATP-competitive ChoKα inhibitors starting from initial hit NMS-P830 that binds to ChoKα in an ATP concentration-dependent manner. This result is confirmed by the co-crystal structure of NMS-P830 in complex with Δ75-ChoKα. NMS-P830 is able to inhibit ChoKα in cells resulting in the reduction of intracellular phosphocholine formation. A structure-based medicinal chemistry program resulted in the identification of selective compounds that have good biochemical activity, solubility and metabolic stability and are suitable for further optimization. The ChoKα inhibitors disclosed in this article demonstrate for the first time the possibility to inhibit ChoKα with ATP-competitive compounds.

BSCI-05. ABL2-HSF1-E2F signaling axis promotes lung adenocarcinoma brain metastasis

Brain metastases are the most common intracranial tumors in adults and are associated with increased patient morbidity and mortality. Limited therapeutic options are currently available for the treatment of brain metastasis. We have identified an actionable signaling pathway utilized by metastatic tumor cells whereby the transcriptional regulator Heat Shock Factor 1 (HSF1) drives a transcriptional program, divergent from its canonical role as the master regulator of the heat shock response, leading to enhanced expression of a subset of E2F transcription factor family gene targets. We showed that HSF1 is required for survival and outgrowth by metastatic lung cancer cells in the brain parenchyma. Unexpectedly, we identified the ABL2 tyrosine kinase as an upstream regulator of HSF1 protein expression, and showed that the Src-homology 3 (SH3) domain of ABL2 directly interacts with HSF1 protein at a non-canonical, proline-independent SH3 interaction motif. Importantly, knockdown of ABL2 impairs expression of HSF1 protein and HSF1-E2F transcriptional gene targets. Notably, we found that pharmacologic inhibition of the ABL kinases using selective ABL allosteric inhibitors, but not ATP-competitive inhibitors, ablates the physical interaction between ABL2 and HSF1, leading to markedly decreased expression of HSF1, E2F1 and E2F8 proteins in brain-metastatic lung cancer cells, and depletion of HSF1-E2F transcriptional targets. These findings highlight potential differences affecting intra- and inter-molecular protein-protein interactions induced by allosteric versus ATP-competitive kinase inhibitors that have important therapeutic implications. Importantly, the targetable nature of the ABL2-HSF1-E2F signaling network identifies ABL allosteric inhibitors as a potentially effective therapy for the treatment of metastatic lung cancers characterized by high expression of HSF1.

Chemical and Structural Strategies to Selectively Target mTOR Kinase.

Dysregulation of the mechanistic target of rapamycin (mTOR) pathway is implicated in cancer and neurological disorder, which identifies mTOR inhibition as promising strategy for the treatment of a variety of human disorders. First‐generation mTOR inhibitors include rapamycin and its analogues (rapalogs) which act as allosteric inhibitors of TORC1. Structurally unrelated, ATP‐competitive inhibitors that directly target the mTOR catalytic site inhibit both TORC1 and TORC2. Here, we review investigations of chemical scaffolds explored for the development of highly selective ATP‐competitive mTOR kinase inhibitors (TORKi). Extensive medicinal chemistry campaigns allowed to overcome challenges related to structural similarity between mTOR and the phosphoinositide 3‐kinase (PI3K) family. A broad region of chemical space is covered by TORKi. Here, the investigation of chemical substitutions and physicochemical properties has shed light on the compounds’ ability to cross the blood brain barrier (BBB). This work provides insights supporting the optimization of TORKi for the treatment of cancer and central nervous system disorders.

Aromatic Rings as Molecular Determinants for the Molecular Recognition of Protein Kinase Inhibitors.

Protein kinases are key enzymes in many signal transduction pathways, and play a crucial role in cellular proliferation, differentiation, and various cell regulatory processes. However, aberrant function of kinases has been associated with cancers and many other diseases. Consequently, competitive inhibition of the ATP binding site of protein kinases has emerged as an effective means of curing these diseases. Over the past three decades, thousands of protein kinase inhibitors (PKIs) with varying molecular frames have been developed. Large-scale data mining of the Protein Data Bank resulted in a database of 2139 non-redundant high-resolution X-ray crystal structures of PKIs bound to protein kinases. This provided us with a unique opportunity to study molecular determinants for the molecular recognition of PKIs. A chemoinformatic analysis of 2139 PKIs resulted in findings that PKIs are “flat” molecules with high aromatic ring counts and low fractions of sp3 carbon. All but one PKI possessed one or more aromatic rings. More importantly, it was found that the average weighted hydrogen bond count is inversely proportional to the number of aromatic rings. Based on this linear relationship, we put forward the exchange rule of hydrogen bonding interactions and non-bonded π-interactions. Specifically, a loss of binding affinity caused by a decrease in hydrogen bonding interactions is compensated by a gain in binding affinity acquired by an increase in aromatic ring-originated non-bonded interactions (i.e., π–π stacking interactions, CH–π interactions, cation–π interactions, etc.), and vice versa. The very existence of this inverse relationship strongly suggests that both hydrogen bonding and aromatic ring-originated non-bonded interactions are responsible for the molecular recognition of PKIs. As an illustration, two representative PKI–kinase complexes were employed to examine the relative importance of different modes of non-bonded interactions for the molecular recognition of PKIs. For this purpose, two FDA-approved PKI drugs, ibrutinib and lenvatinib, were chosen. The binding pockets of both PKIs were thoroughly examined to identify all non-bonded intermolecular interactions. Subsequently, the strengths of interaction energies between ibrutinib and its interacting residues in tyrosine kinase BTK were quantified by means of the double hybrid DFT method B2PLYP. The resulting energetics for the binding of ibrutinib in tyrosine kinase BTK showed that CH–π interactions and π–π stacking interactions between aromatic rings of the drug and hydrophobic residues in its binding pocket dominate the binding interactions. Thus, this work establishes that, in addition to hydrogen bonding, aromatic rings function as important molecular determinants for the molecular recognition of PKIs. In conclusion, our findings support the following pharmacophore model for ATP-competitive kinase inhibitors: a small molecule features a scaffold of one or more aromatic rings which is linked with one or more hydrophilic functional groups. The former has the structural role of acting as a scaffold and the functional role of participating in aromatic ring-originated non-bonded interactions with multiple hydrophobic regions in the ATP binding pocket of kinases. The latter ensure water solubility and form hydrogen bonds with the hinge region and other hydrophilic residues of the ATP binding pocket.

Marine-Derived Natural Products as ATP-Competitive mTOR Kinase Inhibitors for Cancer Therapeutics.

The mammalian target of rapamycin (mTOR) is a serine/threonine kinase portraying a quintessential role in cellular proliferation and survival. Aberrations in the mTOR signaling pathway have been reported in numerous cancers including thyroid, lung, gastric and ovarian cancer, thus making it a therapeutic target. To attain this objective, an in silico investigation was designed, employing a pharmacophore modeling approach. A structure-based pharmacophore (SBP) model exploiting the key features of a selective mTOR inhibitor, Torkinib directed at the ATP-binding pocket was generated. A Marine Natural Products (MNP) library was screened using SBP model as a query. The retrieved compounds after consequent drug-likeness filtration were subjected to molecular docking with mTOR, thus revealing four MNPs with better scores than Torkinib. Successive refinement via molecular dynamics simulations demonstrated that the hits formed crucial interactions with key residues of the pocket. Furthermore, the four identified hits exhibited good binding free energy scores through MM-PBSA calculations and the subsequent in silico toxicity assessments displayed three hits deemed essentially non-carcinogenic and non-mutagenic. The hits presented in this investigation could act as potent ATP-competitive mTOR inhibitors, representing a platform for the future discovery of drugs from marine natural origin.

SAPROCAN: Saracatinib (AZD0530) and docetaxel in metastatic,castrate-refractory prostate cancer (mCRPC)—A phase I/randomized phase II study by the United Kingdom National Cancer Research Institute Prostate Group.

107 Background: Saracatinib is an orally-available, highly selective inhibitor of Abl and Src family members. It is an ATP-competitive tyrosine kinase inhibitor. Preclinical data suggested that the combination of a Src kinase inhibitor and docetaxel is synergistic, and Src kinase activity was also implicated in the bone’s metabolic response to cancer metastases. Methods: Patients with mCRPC were initially enrolled in an open-label, dose escalation phase I trial of oral saracatinib (cohorts of 50mg, 125mg and 175mg daily) with docetaxel (75mg/m2) in a 3+3 design. Subsequent patients were randomised 1:1 between saracatinib 175mg and placebo once daily. Pharmacokinetics (PK) of docetaxel were explored in phase I to exclude significant drug-drug interaction. The primary endpoint of phase II was biochemical or radiographic progression free survival (PFS). Secondary endpoints included overall survival (OS), safety and tolerability. Changes in circulating tumour cell (CTC) counts were also measured. The phase II was designed with a 1-sided alpha of 0.2 with 90% power to detect a hazard ratio (HR) for PFS of 0.67. Results: 10 patients were enrolled in phase I and 142 in the randomised phase II. No dose limiting toxicities or PK interactions were observed and the recommended dose for phase II was 175mg saracatinib daily and 75mg/m2docetaxel every 21 days. In phase II, the HR for PFS was 1.35 (80% confidence interval (CI) 1.07 to 1.70). The HR for OS was 1.42 (1.08 – 1.81). 41/71 and 29/71 experienced treatment related toxicities of grade 3 or above in the saracatinib and placebo arms respectively. 10/19 (53%) and 14/27 (52%) evaluable patients demonstrated a reduction in CTCs from ≥5 to < 5 /7.5ml blood at 6 weeks after starting saracatinib and placebo respectively. Conclusions: Saracatinib, in combination with docetaxel, adds toxicity and not efficacy in mCRPC. This combination should not be developed further in combination with docetaxel in the treatment of mCRPC. Clinical trial information: ISRCTN22566729.

A cell-based drug delivery platform for treating central nervous system inflammation.

Mesenchymal stem cells (MSCs) are promising candidates for the development of cell-based drug delivery systems for autoimmune inflammatory diseases, such as multiple sclerosis (MS). Here, we investigated the effect of Ro-31-8425, an ATP-competitive kinase inhibitor, on the therapeutic properties of MSCs. Upon a simple pretreatment procedure, MSCs spontaneously took up and then gradually released significant amounts of Ro-31-8425. Ro-31-8425 (free or released by MSCs) suppressed the proliferation of CD4+ T cells in vitro following polyclonal and antigen-specific stimulation. Systemic administration of Ro-31-8425-loaded MSCs ameliorated the clinical course of experimental autoimmune encephalomyelitis (EAE), a murine model of MS, displaying a stronger suppressive effect on EAE than control MSCs or free Ro-31-8425. Ro-31-8425-MSC administration resulted in sustained levels of Ro-31-8425 in the serum of EAE mice, modulating immune cell trafficking and the autoimmune response during EAE. Collectively, these results identify MSC-based drug delivery as a potential therapeutic strategy for the treatment of autoimmune diseases. KEY MESSAGES: MSCs can spontaneously take up the ATP-competitive kinase inhibitor Ro-31-8425. Ro-31-8425-loaded MSCs gradually release Ro-31-8425 and exhibit sustained suppression of T cells. Ro-31-8425-loaded MSCs have more sustained serum levels of Ro-31-8425 than free Ro-31-8425. Ro-31-8425-loaded MSCs are more effective than MSCs and free Ro-31-8425 for EAE therapy.