Protein Tyrosine Kinase Inhibitor Research Articles

Role of SPRY4 in health and disease.

SPRY4 is a protein encoding gene that belongs to the Spry family. It inhibits the mitogen-activated protein kinase (MAPK) signaling pathway and plays a role in various biological functions under normal and pathological conditions. The SPRY4 protein has a specific structure and interacts with other molecules to regulate cellular behavior. It serves as a negative feedback inhibitor of the receptor protein tyrosine kinases (RTK) signaling pathway and interferes with cell proliferation and migration. SPRY4 also influences inflammation, oxidative stress, and cell apoptosis. In different types of tumors, SPRY4 can act as a tumor suppressor or an oncogene. Its dysregulation is associated with the development and progression of various cancers, including colorectal cancer, glioblastoma, hepatocellular carcinoma, perihilar cholangiocarcinoma, gastric cancer, breast cancer, and lung cancer. SPRY4 is also involved in organ development and is associated with ischemic diseases. Further research is ongoing to understand the expression and function of SPRY4 in specific tumor microenvironments and its potential as a therapeutic target.

5-Hydroxytryptamine Enhances the Pacemaker Activity of Interstitial Cells of Cajal in Mouse Colon.

We examined the localization of the 5-hydroxytryptamine (5-HT) receptor and its effects on mouse colonic interstitial cells of Cajal (ICCs) using electrophysiological techniques. Treatment with 5-HT increased the pacemaker activity in colonic ICCs with depolarization of membrane potentials in a dose-dependent manner. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channel blockers blocked pacemaker activity and 5-HT-induced effects. Moreover, an adenylate cyclase inhibitor inhibited 5-HT-induced effects, and cell-permeable 8-bromo-cAMP increased the pacemaker activity. Various agonists of the 5-HT receptor subtype were working in colonic ICCs, including the 5-HT4 receptor. In small intestinal ICCs, 5-HT depolarized the membrane potentials transiently. Adenylate cyclase inhibitors or HCN blockers did not show any influence on 5-HT-induced effects. Anoctamin-1 (ANO1) or T-type Ca2+ channel blockers inhibited the pacemaker activity of colonic ICCs and blocked 5-HT-induced effects. A tyrosine protein kinase inhibitor inhibited pacemaker activity in colonic ICCs under controlled conditions but did not show any influence on 5-HT-induced effects. Among mitogen-activated protein kinase (MAPK) inhibitors, a p38 MAPK inhibitor inhibited 5-HT-induced effects on colonic ICCs. Thus, 5-HT's effect on pacemaker activity in small intestinal and colonic ICCs has excitatory but variable patterns. ANO1, T-type Ca2+, and HCN channels are involved in 5-HT-induced effects, and MAPKs are involved in 5-HT effects in colonic ICCs.

Abstract B024: Maintenance therapy inhibition of ptk2 yields decreased disease in preclinical HRP/HRD models of recurrent HGSOC

Abstract B024: Maintenance therapy inhibition of ptk2 yields decreased disease in preclinical HRP/HRD models of recurrent HGSOC

Adaptive Darwinian off-target resistance mechanisms to selective RET inhibition in RET driven cancer

Patients treated with RET protein tyrosine kinase inhibitors (TKIs) selpercatinib or pralsetinib develop RET TKI resistance by secondary RET mutations or alterative oncogenes, of which alterative oncogenes pose a greater challenge for disease management because of multiple potential mechanisms and the unclear tolerability of drug combinations. A patient with metastatic medullary thyroid carcinoma (MTC) harboring a RET activation loop D898_E901del mutation was treated with selpercatinib. Molecular alterations were monitored with tissue biopsies and cfDNA during the treatment. The selpercatinib-responsive MTC progressed with an acquired ETV6::NTRK3 fusion, which was controlled by selpercatinib plus the NTRK inhibitor larotrectinib. Subsequently, tumor progressed with an acquired EML4::ALK fusion. Combination of selpercatinib with the dual NTRK/ALK inhibitor entrectinib reduced the tumor burden, which was followed by appearance of NTRK3 solvent-front G623R mutation. Preclinical experiments validated selpercatinib plus larotrectinib or entrectinib inhibited RET/NTRK3 dependent cells, whereas selpercatinib plus entrectinib was necessary to inhibit cells with RET/NTRK3/ALK triple alterations or a mixture of cell population carrying these genetic alterations. Thus, RET-altered MTC adapted to selpercatinib and larotrectinib with acquisition of ETV6::NTRK3 and EML4::ALK oncogenes can be managed by combination of selpercatinib and entrectinib providing proof-of-concept of urgency of incorporating molecular profiling in real-time and personalized N-of-1 care transcending one-size-fits-all approach.

Transforming Growth Factor α Evokes Aromatase Expression in Gastric Parietal Cells during Rat Postnatal Development.

Estrogen, well known as a female hormone, is synthesized primarily by ovarian aromatase. However, extra-glandular tissues also express aromatase and produce estrogen. It is noteworthy that aromatase in gastric parietal cells begins expression around 20 days after birth and continues secreting considerable amounts of estrogen into the portal vein throughout life, supplying it to the liver. Estrogen, which is secreted from the stomach, is speculated to play a monitoring role in blood triglyceride, and its importance is expected to increase. Nevertheless, the regulatory mechanisms of the aromatase expression remain unclear. This study investigated the influence of transforming growth factor α (TGFα) on gastric aromatase expression during postnatal development. The administration of TGFα (50 μg/kg BW) to male Wistar rats in the weaning period resulted in enhanced aromatase expression and increased phosphorylated ERK1+2 in the gastric mucosa. By contrast, administration of AG1478 (5 mg/kg BW), a protein tyrosine kinase inhibitor with high selectivity for the epidermal growth factor receptor and acting as an antagonist of TGFα, led to the suppression of aromatase expression. In fact, TGFα expression in the gastric fundic gland isthmus began around 20 days after birth in normal rats as did that of aromatase, which indicates that TGFα might induce the expression of aromatase in the parietal cells concomitantly.

Genistein stimulates the viability and prevents myofibroblastic transformation in human trabecular meshwork cells stimulated by TGF-β

Primary open-angle glaucoma (POAG) is the most common type of glaucoma leading to blindness. The search for ways to prevent/treat this entity is one of the main challenges of today's ophthalmology. One of such solution seems to be biologically active substances of natural origin, such as genistein (GEN), which can affect the function of isolated trabecular meshwork by the inhibition of protein tyrosine kinase. However, the role of GEN in viability as well as myofibroblastic transformation in human trabecular meshwork cells stimulated by TGF-β is unknown. Using human trabecular meshwork cells (HTMCs) we investigated the effect of genistein on cell viability and myofibroblastic transformation stimulated by TGF-β1 and TGF-β2. Using Real-Time PCR, western blot and immunofluorescence we determined the effect on the expression changes of αSMA, TIMP1, collagen 1 and 3 at mRNA and protein level. We found that genistein increases the viability of HTMCs (1, 2, 3 μg/ml; P < 0.05 and 4, 5, 10, 15, 20 μg/ml; P < 0.01). Moreover, we found that addition of 10, 15 and 20 μg/ml is able to prevent myofibroblastic transformation of HTMCs by decreasing αSMA, TIMP1, collagen 1 and 3 mRNA and protein expression (P < 0.01). Based on the obtained results, we can conclude that genistein is a potential factor that can prevent the myofibroblastic transformation of HTMCs accompanying glaucoma. Describing GEN influence on myofibroblastic transformation processes in HTMC allows us to conclude that it can be considered a potential therapeutic agent or a substance supporting treatment in patients with glaucoma.

Dermatofibrosarcoma protuberans: Under the mask of scar

Dermatofibrosarcoma protuberans (Darier–Ferrand) is a rare mesenchymal tumor of fibrohistiocytic genesis with moderate malignancy and low incidence of metastasis, the pathogenesis and etiology of which remain unclear. The diagnosis is based on the clinical picture of a reddish-brown dense plaque or papule that gradually becomes a painful node with ulceration. Histological examination reveals the infiltrate with a moire pattern in the dermis in the form of fusiform cells with diffuse CD34+ staining during immunohistochemical analysis. Treatment of dermatofibrosarcoma protuberans (Darier–Ferrand) is a serious problem due to the high tendency for recurrence after removal. The methods described in the literature include extensive excision, Mohs surgery, chemotherapy, radiation therapy and targeted treatment with an anticancer protein-tyrosine kinase inhibitor.&#x0D; The authors present their own clinical observation of a patient with dermatofibrosarcoma protuberans (Darier–Ferrand) in whom a diagnostic error by only visual method and an incorrect treatment led to the worsening of skin process. The interest of this clinical observation lies in the difficulty of diagnosis during the initial visit, in the complexity of differential and morphological diagnosis.

The synergistic regulatory effect of PTP1B and PTK inhibitors on the development of Oedaleus decorus asiaticus Bei-Bienko

Tyrosine phosphorylation is crucial for controlling normal cell growth, survival, intercellular communication, gene transcription, immune responses, and other processes. PTP (protein tyrosine phosphatase) and PTK (protein tyrosine kinases) can achieve this goal by regulating multiple signaling pathways. Oedaleus decorus asiaticus is an important pest that infests the Mongolian Plateau grassland. We aimed to evaluate the survival rate, growth rate, overall performance, and ovarian developmental morphology of the 4th instar nymphs of O. decorus asiaticus while inhibiting the activity of PTP1B (protein tyrosine phosphatase-1B) and PTK. In addition, the expression and protein phosphorylation levels of key genes in the MAPK signaling pathway and antioxidant enzyme activity were assessed. The results showed no significant differences in survival rate, growth rate, or overall performance between PTP1B inhibitor treatment and control. However, after PTK inhibitor treatment, these indexes were significantly lower than in the control. The ovarian size of female larvae after 15 days of treatment with PTK inhibitors showed significantly slower development, while female larvae treated with PTP1B exhibited faster ovarian growth than the control group. In comparison to controls and nymphs treated with PTK inhibitors, the expression and phosphorylation levels of key genes in the MAPK signaling pathway under PTP1B inhibitor treatments were significantly higher in 4th instar nymphs. However, ROS levels and the activities of NADPH oxidase and other antioxidant enzymes were considerably reduced, although they were significantly greater in the PTK inhibitor treatment. The results suggest that PTP1B and PTK feedback inhibition in the MAPK signal transfer can regulate the physiological metabolism of the insect as well as its developmental rate. These findings can facilitate future uses of PTP1B and PTK inhibitors in controlling insect development to help control pest populations.

A Phase 2 Study of Ponatinib for Prevention of Relapse after Allotransplantation in FLT3-ITD + AML Patients : The Ponallo Trial

Background : For acute myeloid leukemia (AML) patients relapse remains a major issue after allogeneic stem cell transplantation (allo-SCT). In those with a FMS-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD+), survivals have been improved with the use of small molecule Flt3 receptor protein-tyrosine kinase inhibitors in maintenance due to a significant decrease of relapse after allo-SCT. Ponatinib (pona) is a multikinase inhibitor that is approved for the treatment of Philadelphia chromosome-positive acute lymphoblastic and chronic myelogenous leukemias. It has also activity against FLT3-mutants including ITD and ITD691L but not D835 within the Flt3 activation segment. Pona may then represent an alternative drug for maintenance after allo-SCT in FLT3-ITD+ AML patients. Methods : This was a multicenter phase 2 non-randomized prospective study testing the efficacy and tolerability of pona after allo-SCT in FLT3-ITD+ AML patients (ClinicalTrial.gov ID : NCT03690115). The main objective was the incidence of relapse at 2 years post-transplant with the hypothesis that this incidence at 2 years will be reduced at 15% instead of 30%. Seventy-seven patients had to be included. Inclusion criteria were all patients, aged between 18 and 70 yo, with a FLT-3 ITD+ AML in cytologic complete remission (&amp;lt;5% of bone marrow blasts) and a documented engraftment at the time of starting pona after an allo-SCT using any type of conditioning regimen and donor. The control of a GVHD (if present) was mandatory as well as a platelets count ≥70 Giga/l and a neutrophils count ≥ 1 Giga/l at time of inclusion. The study was supported by Incyte. Initially, pona administration had to start between day +60 and +90/100 at the dose of 30 mg/day for 1-year. However, because of excesssive hematopoietic toxicity, an amendment was made after the 14th patient to start pona between day 100 and day 120 post-allo. During the study, It was allowed to restart pona at 15 mg/day after resolution of 30 mg dose related toxicities. After 1 year of pona it was not allowed to start a new FLT3 inhibitor or pursue pona. Results : Twenty-three patients were included between December 2019 and April 2022. Among them, 22 were effectively treated. Indeed, the last patient included did not receive pona due to hepatic abnormalities incompatible with the start of pona. Also, because of a very low rate of enrollment, the trial was closed prematurely in accordance with the sponsor in December 2022. Results for the 22 treated patients are reported here. Patient characteristics are given in Table 1. With a median follow-up of 740 days (range : 374-930), 1-y and 2-y overall survival were 81.3% (66.3-99.6) and 63.9% (45.4-90.0), and 1-y and 2-y leukemia-free survival were 54.5% (37.2-79.8) and 49% (31.8-75.7), respectively. Ten relapses occured at a median of 186 days (range=69-481) after allo-SCT with a CI of relapse of 45.5% at 2 years. Pona was started at a median of 69 days post-allo-SCT (range : 60-130). The median duration of treatment was 75.5 days (range : 4-369). Only 3 patients received one year of treatment and all at 30 mg/day as required by the protocol. The reasons for stopping pona were toxicity in 11 patients, relapse in 6, toxicity then relapse in 1, GVHD then relapse in 1. Among the 13 patients who stopped pona for toxicity or GVHD, 6 restarted pona (15 mg n=5, 30 mg n=1) then stoppped again for toxicity (n=4) or progression (n=2). A total of 36 adverse events related to pona were documented in 17 patients. The majority was grade 1 or 2 (n=27, 75%) while 6 grade 3 (17%) and 3 grade 4 (8%) occured. ( Table 2) Conclusion : Ponatinib administration after allo-SCT is associated with toxicity and do not reduce relapse or improve survival in this small cohort of FLT3+ AML patients. Other FLT3 inhibitors have to be considered in FLT3+ AML patients for maintenance after allo-SCT.

Reprogramming the pancreatic cancer stroma and immune landscape by a silicasome nanocarrier delivering nintedanib, a protein tyrosine kinase inhibitor

The prevailing desmoplastic stroma and immunosuppressive microenvironment within pancreatic ductal adenocarcinoma (PDAC) pose substantial challenges to therapeutic intervention. Despite the potential of protein tyrosine kinase (PTK) inhibitors in mitigating the desmoplastic stromal response and enhancing the immune milieu, their efficacy is curtailed by suboptimal pharmacokinetics (PK) and insufficient tumor penetration. To surmount these hurdles, we have pioneered a novel strategy, employing lipid bilayer-coated mesoporous silica nanoparticles (termed "silicasomes") as a carrier for the delivery of Nintedanib. Nintedanib, a triple PTK inhibitor that targets vascular endothelial growth factor, platelet-derived growth factor and fibroblast growth factor receptors, was encapsulated in the pores of silicasomes via a remote loading mechanism for weak bases. This innovative approach not only enhanced pharmacokinetics and intratumor drug concentrations but also orchestrated a transformative shift in the desmoplastic and immune landscape in a robust orthotopic KRAS-mediated pancreatic carcinoma (KPC) model. Our results demonstrate attenuation of vascular density and collagen content through encapsulated Nintedanib treatment, concomitant with significant augmentation of the CD8+/FoxP3+ T-cell ratio. This remodeling was notably correlated with tumor regression in the KPC model. Strikingly, the synergy between encapsulated Nintedanib and anti-PD-1 immunotherapy further potentiated the antitumor effect. Both free and encapsulated Nintedanib induced a transcriptional upregulation of PD-L1 via the extracellular signal-regulated kinase (ERK) pathway. In summary, our pioneering approach involving the silicasome carrier not only improved antitumor angiogenesis but also profoundly reshaped the desmoplastic stromal and immune landscape within PDAC. These insights hold excellent promise for the development of innovative combinatorial strategies in PDAC therapy.

Beta-Hydroxyisovaleryl-Shikonin Eradicates Epithelial Cell Adhesion Molecule-Positive Liver Cancer Stem Cells by Suppressing dUTP Pyrophosphatase Expression.

Cancer stem cells (CSCs) play an essential role in tumorigenesis, chemoresistance, and metastasis. Previously, we demonstrated that the development of hepatocellular carcinoma (HCC) is dictated by a subset of epithelial cell adhesion molecule-positive (EpCAM+) liver CSCs with the activation of Wnt signaling. In this study, we evaluated the expression of dUTP pyrophosphatase (dUTPase), which plays a central role in the development of chemoresistance to 5-fluorouracil, in EpCAM+ HCC cells. We further evaluated the effect of beta-hydroxyisovaleryl-shikonin (β-HIVS), an ATP-noncompetitive inhibitor of protein tyrosine kinases, on HCC CSCs. EpCAM and dUTPase were expressed in hepatoblasts in human fetal liver, hepatic progenitors in adult cirrhotic liver, and a subset of HCC cells. Sorted EpCAM+ CSCs from HCC cell lines showed abundant nuclear accumulation of dUTPase compared with EpCAM-negative cells. Furthermore, treatment with the Wnt signaling activator BIO increased EpCAM and dUTPase expression. In contrast, β-HIVS treatment decreased dUTPase expression. β-HIVS treatment decreased the population of EpCAM+ liver CSCs in a dose-dependent manner in vitro and suppressed tumor growth in vivo compared with the control vehicle. Taken together, our data suggest that dUTPase could be a good target to eradicate liver CSCs resistant to 5-fluorouracil. β-HIVS is a small molecule that could decrease dUTPase expression and target EpCAM+ liver CSCs.

RETooling the RET Inhibitor Pralsetinib for ESR1 Fusion-Positive Breast Cancer and Beyond.

Transcriptionally active fusions of ESR1 (ESR1-TAF) and somatic mutations in the estrogen receptor alpha (ERα) ligand-binding domain (LBD) cause endocrine therapy resistance in breast cancer. In searching for therapeutic target kinase(s) in these breast cancers, Gou and colleagues identified FLT4, RET, JAK1, and IGF1R as the top upregulated kinases induced by ESR1-TAFs and ERα LBD mutants in breast cancer cells. Among them, inhibition of RET by pralsetinib suppressed ESR1-TAF-driven and ERα LBD mutant-driven cell proliferation and patient-derived xenograft growth. Pralsetinib is an inhibitor of the RET protein tyrosine kinase that is approved for treating oncogenic RET mutation-positive and RET fusion-positive thyroid cancers and non-small cell lung cancer. The work by Gou and colleagues reinforces the knowledge of RET as an ESR1 target gene and highlights that RET interacts with ERα to promote breast cancer tumorigenesis and antiestrogen resistance. It also raises the prospect of repositioning pralsetinib to target wildtype RET in ER-positive breast cancer. See related article by Gou et al., p. 3237.

Lupenone attenuates thapsigargin-induced endoplasmic reticulum stress and apoptosis in pancreatic beta cells possibly through inhibition of protein tyrosine kinase 2 activity

AimsProlonged high levels of cytokines, glucose, or free fatty acids are associated with diabetes, elevation of cytosolic Ca2+ concentration ([Ca2+]C), and depletion of Ca2+ concentration in the endoplasmic reticulum (ER) of pancreatic beta cells. This Ca2+ imbalance induces ER stress and apoptosis. Lupenone, a lupan-type triterpenoid, is beneficial in diabetes; however, its mechanism of action is yet to be clarified. This study evaluated the protective mechanism of lupenone against thapsigargin-induced ER stress and apoptosis in pancreatic beta cells. Materials and methodsMIN6, INS-1, and native mouse islet cells were used. Western blot for protein expressions, measurement of [Ca2+]C, and in vivo glucose tolerance test were mainly performed. Key findingsThapsigargin increased the protein levels of cleaved caspase 3, cleaved PARP, and the phosphorylated form of JNK, ATF4, and CHOP. Thapsigargin increased the interaction between stromal interaction molecule1 (Stim1) and Orai1, enhancing store-operated calcium entry (SOCE). SOCE is further activated by protein tyrosine kinase 2 (Pyk2), which is Ca2+-dependent and phosphorylates the tyrosine residue at Y361 in Stim1. Lupenone inhibited thapsigargin-mediated Pyk2 activation, suppressed [Ca2+]C, ER stress, and apoptosis. Lupenone restored impaired glucose-stimulated insulin secretion effectuated by thapsigargin and glucose intolerance in a low-dose streptozotocin-induced diabetic mouse model. SignificanceThese results suggested that lupenone attenuated thapsigargin-induced ER stress and apoptosis by inhibiting SOCE; this may be due to the hindrance of Pyk2-mediated Stim1 tyrosine phosphorylation. In beta cells that are inevitably exposed to frequent [Ca2+]C elevation, the attenuation of abnormally high SOCE would be beneficial for their survival.

Application of Simplex Lattice Mixture design and desirability function in the development and Optimization of SEDDS for protein kinase inhibitor-Pazopanib Hydrochloride

Pazopanib Hydrochloride is a tyrosine protein kinase inhibitor molecule approved by USFDA and European agencies for the treatment of renal cell carcinoma (RCC) patients and other renal malignancies, but it has very poor aqueous solubility. Therefore, it is essential need to improve the solubility and in vitro dispersion or release characteristics. The purpose of this study was to investigate the Pazopanib hydrochloride drug solubility in various vehicles and screening of suitable solubilizers for the preparation of self-emulsifying lipid-based drug delivery systems (SE-LBDDS) of a poorly water-soluble drug (BCS class II), Pazopanib HCl by using simplex lattice mixture design. Ternaryplots wereconstructed by using oil (Labrafac WL 1349l), surfactant (Labrasol), and co-surfactant (Transcutol-P), and the concentration ranges were determined by using a simplex lattice design. The composition of pazopanib HCl SEEDS was optimized through various dependent variables (responses)such as solubility (Y1), precipitation after 15 min (Y2), and, particle size (Y3).Solubility study of pazopanib HCl in different oils, surfactants, and co-surfactants was carried out by shake flask method at 37°C. Three formulation components were chosen based on the maximum solubility results inthe oil, surfactant, and co-surfactant category and included in the experimental design. The results were analyzed by model fitting using the standard least-squares method. Pazopanib HCl were shown maximum solubility i.e. 25.64±0.24mg/g, 57.84 ±2.91mg/g and, 44.61±1.51mg/g in Labrafac WL 1349 (oil), Labrasol (surfactant) and Transcutol-P (co-surfactant) respectively. Hence these chosen formulation component's concentrations were further optimized by using a simplex lattice design (SLD). Derived mathematical polynomial equations and models were exercised to evaluate the impact of formulation input variables on the output variables (responses) using JMP software. The model p-value for both the responses i.e. solubility and particle size were found less than 0.05 hence models were significant. The results of the mathematical analysis demonstrated that the formulation components have a significant impact on the studied responses. Hence simplex lattice mixture design can be used as a powerful quality design to predict the optimized SEDDS formulation.The applicability of simplex lattice design with desirability function helped optimize a SEEDS formulation of pazopanib HCl and the selected model has made it possible to identify the impact of the critical factors to optimize the required responses.

Involvement of the AKT Pathway in Resistance to Erlotinib and Cabozantinib in Triple-Negative Breast Cancer Cell Lines.

Resistance to protein tyrosine kinase inhibitors (TKIs) presents a significant challenge in therapeutic target development for cancers such as triple-negative breast cancer (TNBC), where conventional therapies are ineffective at combatting systemic disease. Due to increased expression, the receptor tyrosine kinases EGFR (epidermal growth factor receptor) and c-Met are potential targets for treatment. However, targeted anti-EGFR and anti-c-Met therapies have faced mixed results in clinical trials due to acquired resistance. We hypothesize that adaptive responses in regulatory kinase networks within the EGFR and c-Met signaling axes contribute to the development of acquired erlotinib and cabozantinib resistance. To test this, we developed two separate models for cabozantinib and erlotinib resistance using the MDA-MB-231 and MDA-MB-468 cell lines, respectively. We observed that erlotinib- or cabozantinib-resistant cell lines demonstrate enhanced cell proliferation, migration, invasion, and activation of EGFR or c-Met downstream signaling (respectively). Using a SILAC (Stable Isotope Labeling of Amino acids in Cell Culture)-labeled quantitative mass spectrometry proteomics approach, we assessed the effects of erlotinib or cabozantinib resistance on the phosphoproteome, proteome, and kinome. Using this integrated proteomics approach, we identified several potential kinase mediators of cabozantinib resistance and confirmed the contribution of AKT1 to erlotinib resistance in TNBC-resistant cell lines.

A meta-analysis: the clinical value of PD-1 inhibitor or protein tyrosine kinase inhibitors in the treatment of advanced osteosarcoma.

PD-1 inhibitors and TKIs have been used to treat advanced osteosarcoma, but there is still a lack of intuitive data for the comparison of their efficacy. We conducted a meta-analysis to evaluate their therapeutic benefits. A systematic methodological search of five primary electronic databases was performed. Studies with a randomized design of any type about PD-1 inhibitors or TKIs for the treatment of advanced osteosarcoma were included. The primary outcomes mainly included CBR, PFS, OS and ORR, The CR, PR, SD and AEs were the secondary outcomes. The survival period (months) of patients was taken as the main analysis data. Random-effects models were used for meta-analysis. Eight immunocheckpoint inhibitors in 327 patients from 10 clinical trials were finally evaluated. For OS, TKIs [11.67 months (95% CI, 9.32-14.01)] show more obvious advantages than PD-1 inhibitors [6.37 months (95% CI, 3.96-8.78)]. For PFS, TKIs [4.79 months (95% CI, 3.33-6.24)] are longer than PD-1 inhibitors [1.46 months (95% CI, 1.23-1.69)]. Although there was no fatal event, attention should still be paid, especially during the combined application of PD-1 inhibitors with TKIs since their obvious AEs. The findings of this study suggest that patients with advanced osteosarcoma, TKIs may be more beneficial than PD-1 inhibitors. TKIs combined with PD-1 inhibitors has a bright future in the treatment of advanced osteosarcoma, but we should always pay attention to the strong side effects.

Response to Pralsetinib in Multi-Drug-Resistant Breast Cancer With CCDC6-RET Mutation

Triple-negative breast cancers (TNBC) represent a pathological subtype of breast cancer, which are characterized by strong invasiveness, high metastasis rate, low survival rate, and poor prognosis, especially in patients who have developed resistance to multiline treatments. Here, we present a female patient with advanced TNBC who progressed despite multiple lines of treatments; next-generation sequencing (NGS) was used to find drug mutation targets, which revealed a coiled-coil domain-containing protein 6 (CCDC6)-rearranged during transfection (RET) gene fusion mutation. The patient was then given pralsetinib, and after one treatment cycle, a CT scan revealed partial remission and adequate tolerance to therapy. Pralsetinib (BLU-667) is a RET-selective protein tyrosine kinase inhibitor that can inhibit the phosphorylation of RET and downstream molecules as well as the proliferation of cells expressing RET gene mutations. This is the first case in the literature of metastatic TNBC with CCDC6-RET fusion treated with pralsetinib, an RET-specific antagonist. This case demonstrates the potential efficacy of pralsetinib in cases of TNBC with RET fusion mutations and suggests that NGS may reveal new opportunities and bring new therapeutic interventions to patients with refractory TNBC.

Dasatinib in combination with BMS-754807 induce synergistic cytotoxicity in lung cancer cells through inhibiting lung cancer cell growth, and inducing autophagy as well as cell cycle arrest at the G1 phase.

Lung cancer is the leading cause of cancer-related deaths worldwide. Combination of drugs targeting independent signaling pathways would effectively block the proliferation of cancer cells with lower concentrations and stronger synergy effects. Dasatinib, a multi-targeted protein tyrosine kinase inhibitor targeting BCR-ABL and kinases of SRC family, has been successfully applied in the treatment of chronic myeloid leukemia (CML). BMS-754807, an inhibitor targeting the insulin-like growth factor 1 receptor (IGF-IR) and insulin receptor (IR) family kinases, has been in phase I development for the treatment of a variety of human cancers. Herein, we demonstrated that dasatinib in combination with BMS-754807 inhibited lung cancer cell growth, while induced autophagy as well as cell cycle arrest at the G1 phase. Dasatinib in combination with BMS-754807 suppressed the expression of cell cycle marker proteins, Rb, p-Rb, CDK4, CDK6 and Cyclin D1, and the PI3K/Akt/mTOR signaling pathway. Dasatinib in combination with BMS-754807 induced autophagy in lung cancer cells, evidenced by the upregulation of LC3B II and beclin-1, the downregulation of LC3B I and SQSTM1/p62, and the autophagic flux observed with a confocal fluorescence microscopy. Furthermore, dasatinib (18mg/kg) in combination with BMS-754807 (18mg/kg) inhibited the growth of tumors in NCI-H3255 xenografts without changing the bodyweight. Overall, our results suggest that dasatinib in combination with BMS-754807 inhibits the lung cancer cell proliferation in vitro and tumor growth in vitro, which indicates promising evidence for the application of the drug combination in lung cancer therapy.

Targeting RET Solvent-Front Mutants with Alkynyl Nicotinamide-Based Inhibitors.

Selpercatinib (LOXO292) and pralsetinib (BLU667) are RET protein tyrosine kinase inhibitors (TKIs) recently approved for treating RET-altered cancers. However, RET mutations that confer selpercatinib/pralsetinib resistance have been identified, necessitating development of next-generation RET TKIs. While acquired RET G810C/R/S/V mutations were reported in selpercatinib-treated patients, it was unclear whether all of these and other potential G810 mutants are resistant to selpercatinib and pralsetinib. Here, we profiled selpercatinib and pralsetinib on all six possible G810 mutants derived from single nucleotide substitution and developed novel alkynyl nicotinamide-based RET TKIs to inhibit selpercatinib/pralsetinib-resistant RET G810 mutants. Surprisingly, the G810V mutant found in a clinical study was not resistant to selpercatinib or pralsetinib. Besides G810C/R/S, G810D also conferred selpercatinib/pralsetinib resistance. Alkynyl nicotinamide compounds such as HSN608, HSL476, and HSL468 have better drug-like properties than alkynyl benzamides. Six of these compounds inhibited all six G810 solvent-front mutants and the V804M gatekeeper mutant with IC50 < 50 nmol/L in cell culture. Oral administration of HSN608 at a well-tolerated dose (30 mg/kg) gave plasma level > 30x the IC50s of inhibiting all G810 mutants in cell culture. In cell-derived xenograft tumors driven by KIF5B-RET (G810C) that contains the most frequently observed solvent-front mutant in selpercatinib-treated patients, HSN608, HSL476, and HSL468 significantly suppressed and caused regression of the selpercatinib-resistant tumors. This study clarifies the sensitivities of different RET solvent-front mutants to selpercatinib and pralsetinib and identifies novel alkylnyl nicotinamide-based RET TKIs for inhibiting selpercatinib/pralsetinib-resistant G810 mutants.

Calcineurin inhibitors stimulate Kir4.1/Kir5.1 of the distal convoluted tubule to increase NaCl cotransporter.

We examine whether calcineurin or protein phosphatase 2B (PP2B) regulates the basolateral inwardly rectifying potassium channel Kir4.1/Kir5.1 in the distal convoluted tubule (DCT). Application of tacrolimus (FK506) or cyclosporine A (CsA) increased whole-cell Kir4.1/Kir5.1-mediated K+ currents and hyperpolarized the DCT membrane. Moreover, FK506-induced stimulation of Kir4.1/Kir5.1 was absent in kidney tubule-specific 12 kDa FK506-binding protein-knockout mice (Ks-FKBP-12-KO). In contrast, CsA stimulated Kir4.1/Kir5.1 of the DCT in Ks-FKBP-12-KO mice, suggesting that FK506-induced stimulation of Kir4.1/Kir5.1 was due to inhibiting PP2B. Single-channel patch-clamp experiments demonstrated that FK506 or CsA stimulated the basolateral Kir4.1/Kir5.1 activity of the DCT, defined by NPo (a product of channel number and open probability). However, this effect was absent in the DCT treated with Src family protein tyrosine kinase (SFK) inhibitor or hydroxyl peroxide. Fluorescence imaging demonstrated that CsA treatment increased membrane staining intensity of Kir4.1 in the DCT of Kcnj10fl/fl mice. Moreover, CsA treatment had no obvious effect on phosphorylated NaCl cotransporter (pNCC) expression in Ks-Kir4.1-KO mice. Immunoblotting showed acute FK506 treatment increased pNCC expression in Kcnj10fl/fl mice, but this effect was attenuated in Ks-Kir4.1-KO mice. In vivo measurement of thiazide-induced renal Na+ excretion demonstrated that FK506 enhanced thiazide-induced natriuresis. This effect was absent in Ks-FKBP-12-KO mice and blunted in Ks-Kir4.1-KO mice. We conclude that inhibition of PP2B stimulates Kir4.1/Kir5.1 of the DCT and NCC and that PP2B inhibition-induced stimulation of NCC is partially achieved by stimulation of the basolateral Kir4.1/Kir5.1.