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ME (Emmy) M Dolman1, Stefan Harmsen1, Ebel HE Pieters1, Rolf W Sparidans2, Marie Lacombe3, Bálint Szokol4, László Orfi4, György Kéri4, Gert Storm1, Wim E Hennink1, Robbert J Kok11Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands; 2Faculty of Science, Department of Pharmaceutical Sciences, Division of Pharmacoepidemiology and Clinical Pharmacology, Utrecht University, Utrecht, The Netherlands; 3Kreatech Biotechnology BV, Amsterdam, The Netherlands; 4Vichem Chemie Ltd, Budapest, HungaryBackground: Activated proximal tubular cells play an important role in renal fibrosis. We investigated whether sunitinib and a kidney-targeted conjugate of sunitinib were capable of attenuating fibrogenic events in tubulointerstitial fibrosis.Methods: A kidney-targeted conjugate was prepared by linkage of a sunitinib analog (named 17864) via a platinum-based linker to the kidney-specific carrier lysozyme. Pharmacological activity of 17864-lysozyme was evaluated in human kidney proximal tubular cells (HK-2); the capability of the kidney-directed conjugate to accumulate in the kidneys was studied in mice. Potential antifibrotic effects of a single-dose treatment were evaluated in the unilateral ureteral obstruction (UUO) model in mice.Results: The 17864-lysozyme conjugate and its metabolites strongly inhibited tyrosine kinase activity. Upon intravenous injection, 17864-lysozyme rapidly accumulated in the kidneys and provided sustained renal drug levels for up to 3 days after a single dose. Renal drug level area under the curve was increased 28-fold versus an equimolar dose of sunitinib malate. Daily treatment of UUO mice with a high dose of sunitinib malate (50 mg/kg) resulted in antifibrotic responses, but also induced drug-related toxicity. A single dose of 17864-lysozyme (equivalent to 1.8 mg/kg sunitinib) was safe but showed no antifibrotic effects.Conclusion: Multikinase inhibitors like sunitinib can be of benefit in the treatment of fibrotic diseases, provided that their safety can be improved by strategies as presented in this paper, and sustained renal levels can be achieved.Keywords: drug delivery, sunitinib, fibrosis, platinum linker

Selective inhibition of the "false" proliferative signals via targeting tyrosine kinases resulting in the induction of apoptosis by depletion of the "survival factors" is one of the most studied and widely accepted concepts of modern chemotherapy. We have synthesized a series of potent tyrosine kinase inhibitors and tested these compounds for apoptosis induction. Some of the tyrosine kinase inhibitors caused either apoptotic or cytoplasmic vacuolar cell death in various tumor cell cultures. The somatostatin analogue oligopeptide TT-232, which indirectly inhibits tyrosine kinases, exerted a dose-dependent apoptosis-inducing effect. The tumor growth-inhibitory effect of TT-232 and some tyrosine kinase inhibitors has also been proven by in vivo experiments, using human tumor xenografts. On the other hand, a dose-dependent pro- or anti-apoptotic activity of --deprenyl has been shown in melanoma cell cultures, the lower doses inhibiting and the higher doses inducing apoptosis. Various metabolites of --deprenyl are responsible for these actions. The effect of --deprenyl is connected with depolarization of mitochondrial membranes. The kinase inhibitors act on the growth factor receptor signaling pathways (survival factor pathways) and initiate the caspase cascade. The key enzyme for the action of both pro-apoptotic and anti-apoptotic compounds is caspase 3. [ABSTRACT FROM AUTHOR]

A novel somatostatin analogue, TT-232 (which inhibits the proliferation of various cell cultures and transplantable mouse tumours), was examined regarding its effect on human melanoma and lymphoma xenografts as a single treatment or in combination with DTIC (dacarbazine) and etoposide. TT-232 inhibited the growth of HT-18 melanoma xenografts, a dose of 5 mg kg(-1) being the most effective. Combination of 1 mg kg(-1) TT-232 with 30 or 60 mg kg(-1) DTIC (administered daily) resulted in a stronger inhibitory effect compared to TT-232 or DTIC as a single modality. Antimetastatic effect of TT-232 treatment combined with DTIC was studied using the B16 mouse melanoma muscle - lung metastasis model. The number of lung metastases of B16 melanoma could be decreased by the daily administration of 1 mg kg(-1) TT-232 or 60 mg kg(-1), but not of 30 mg kg(-1) DTIC. TT-232, combined with 30 or 60 mg kg(-1) DTIC decreased the lung metastasis number significantly lower than the control. Nearly 50% growth inhibition of HT-58 lymphoma was achieved by daily treatment with 1 mg kg(-1) TT-232. 5 mg kg(-1) etoposide, administered daily, resulted in a similar effect. The combination of 1 mg kg(-1) TT-232 and 5 mg kg(-1) etoposide was significantly more effective than TT-232 or etoposide as a single treatment. The very strong tumour growth inhibitory effect of 10 mg kg(-1) etoposide could even be increased by combination with TT-232. These experimental data suggest that TT-232 may be an effective new tool in the combination chemotherapy of malignant tumours like melanoma and lymphoma. [ABSTRACT FROM AUTHOR]

Abstract: Imatinib (IMT) is a selective tyrosine kinase inhibitor, used in the treatment of chronic myeloid leukemia and gastrointestinal stromal tumors. Its strong plasma protein binding was found to belong to the F1*S genetic variant of α1-acid glycoprotein (AGP). In this work, comparative AGP binding studies were performed with IMT fragment molecules to reveal which parts of the molecule are important in the high-affinity interaction provoking specific spectral changes. Molecular modeling calculations indicated that IMT docked into the X-ray structure of AGP/F1 adopts a bent, compact conformation. This binding mode is similar to those found in its complexes with some low-affinity kinases and a quinone reductase, being strikingly different from the extended conformation of IMT in its high-affinity kinase targets. [Copyright &y& Elsevier]

We provided strong proof of concept evidence that somatostatin mediates potent analgesic and anti-inflammatory actions via its receptor subtype 4 (sst 4 ) located both at the periphery and the central nervous system. Therefore, sst 4 agonists are promising novel drug candidates for neuropathic pain and neurogenic inflammation, but rational drug design was not possible due to the lack of knowledge about its 3-dimensional structure. We modeled the sst 4 receptor structure, described its agonist binding properties, and characterized the binding of our novel small molecule sst 4 agonists (4-phenetylamino-7H-pyrrolo[2,3-d]pyrimidine derivatives) using an in silico platform. In addition to the in silico binding data, somatostatin displacement by Compound 1 was demonstrated in the competitive binding assay on sst 4 -expressing cells. In vivo effects were investigated in rat models of neurogenic inflammation and chronic traumatic neuropathic pain. We defined high- and low-affinity binding pockets of sst 4 for our ligands, binding of the highest affinity compounds were similar to that of the reference ligand J-2156. We showed potent G-protein activation with the highest potency of 10 nM EC 50 value and highest efficacy of 342%. Oral administration of 100 μg/kg of 5 compounds significantly inhibited acute neurogenic plasma protein extravasation in the paw skin by 40-60%, one candidate abolished and 3 others diminished sciatic nerve-ligation induced neuropathic hyperalgesia by 28-62%. The in silico predictions on sst 4 -ligands were tested in biological systems. Low oral dose of our novel agonists inhibit neurogenic inflammation and neuropathic pain, which opens promising drug developmental perspectives for these unmet medical need conditions., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Targeted therapies against cancer types with more than one driver gene hold bright but elusive promise, since approved drugs are not available for all driver mutations and monotherapies often result in resistance. Targeting multiple driver genes in different pathways at the same time may provide an impact extensive enough to fight resistance. Our goal was to find synergistic drug combinations based on the availability of targeted drugs and their biological activity profiles and created an associated compound library based on driver gene-related protein targets. In this study, we would like to show that driver gene pattern based customized combination therapies are more effective than monotherapies on six cell lines and patient-derived primary cell cultures. We tested 55-102 drug combinations targeting driver genes and driver pathways for each cell line and found 25-85% of these combinations highly synergistic. Blocking 2-5 cancer pathways using only 2-3 targeted drugs was sufficient to reach high rates of tumor cell eradication at remarkably low concentrations. Our results demonstrate that the efficiency of cancer treatment may be significantly improved by combining drugs against multiple tumor specific drivers., Competing Interests: CONFLICTS OF INTEREST The authors declare no conflicts of interest.

Pancreatic cancer is an increasing cause of cancer related death worldwide. KRAS is the dominant oncogene in this cancer type and molecular rationale would indicate, that inhibitors of the downstream target MEK could be appropriate targeted agents, but clinical trials have failed so far to achieve statistically significant benefit in unselected patients. We aimed to identify predictive molecular biomarkers that can help to define subgroups where MEK inhibitors might be beneficial alone or in combination. Next-generation sequencing data of 50 genes in three pancreatic cancer cell lines (MiaPaCa2, BxPC3 and Panc1) were analyzed and compared to the molecular profile of 138 clinical pancreatic cancer samples to identify the molecular subtypes of pancreatic cancer these cell lines represent. Luminescent cell viability assay was used to determine the sensitivity of cell lines to kinase inhibitors. Western blot was used to analyze the pathway activity of the examined cell lines. According to our cell viability and pathway activity data on these model cell lines only cells harboring the rare G12C KRAS mutation and low EGFR expression are sensitive to single MEK inhibitor (trametinib) treatment. The common G12D KRAS mutation leads to elevated baseline Akt activity, thus treatment with single MEK inhibitors fails. However, combination of MEK and Akt inhibitors are synergistic in this case. In case of wild-type KRAS and high EGFR expression MEK inhibitor induced Akt phosphorylation leads to trametinib resistance which necessitates for MEK and EGFR or Akt inhibitor combination treatment. In all we provide strong preclinical rational and possible molecular mechanism to revisit MEK inhibitor therapy in pancreatic cancer in both monotherapy and combination, based on molecular profile analysis of pancreatic cancer samples and cell lines. According to our most remarkable finding, a small subgroup of patients with G12C KRAS mutation may still benefit from MEK inhibitor monotherapy.