Small Molecule Tyrosine Kinase Inhibitors Research Articles

Reversing pathology in a preclinical model of Alzheimer's disease by hacking cerebrovascular neoangiogenesis with advanced cancer therapeutics.

BackgroundCognitive decline leading to dementia, accompanied by the accumulation of amyloid-beta (Aβ) in neuritic plaques together with the appearance of neurofibrillary tangles (NFT) composed of hyperphosphorylated tau protein (tau), are previously noted hallmarks of Alzheimer's disease (AD). We previously discovered hypervascularity in brain specimens from AD patients and consistent with this observation, we demonstrated that overexpression of Aβ drives cerebrovascular neoangiogenesis leading to hypervascularity and coincident tight-junction disruption and blood-brain barrier (BBB) leakiness in animal models of AD. We subsequently demonstrated that amyloid plaque burden and cerebrovascular pathogenesis subside when pro-angiogenic Aβ levels are reduced. Based on these data, we propose a paradigm of AD etiology where, as a compensatory response to impaired cerebral blood flow (CBF), Aβ triggers pathogenic cerebrovascular neoangiogenesis that underlies the conventional hallmarks of AD. Consequently, here we present evidence that repurposing anti-cancer drugs to modulate cerebrovascular neoangiogenesis, rather than directly targeting the amyloid cascade, may provide an effective treatment for AD and related vascular diseases of the brain.MethodsWe explored whether the anti-cancer drug, Axitinib, a small molecule tyrosine kinase inhibitor that targets vascular endothelial growth factor receptors (VEGFR) can inhibit aberrant cerebrovascular neoangiogenic changes, reduce Aβ deposits and reverse cognitive decline in an animal model of AD. One month post-treatment with Axitinib, we employed a battery of tests to assess cognition and memory in aged Tg2576 AD mice and used molecular analysis to demonstrate reduction of amyloid plaques, BBB leakage, hypervascularity and associated disease pathology.FindingsTargeting the pro-angiogenic pathway in AD using the cancer drug, Axitinib, dramatically reduced cerebrovascular neoangiogenesis, restored BBB integrity, resolved tight-junction pathogenesis, diminishes Aβ depositions in plaques and effectively restores memory and cognitive performance in a preclinical mouse model of AD.InterpretationModulation of neoangiogenesis, in an analogous approach to those used to treat aberrant vascularization in cancer and also in the wet form of age-related macular degeneration (AMD), provides an alternative therapeutic strategy for intervention in AD that warrants clinical investigation.

331TiP HER2CLIMB-04: Phase II trial of tucatinib + trastuzumab deruxtecan in patients with HER2+ locally advanced or metastatic breast cancer with and without brain metastases

Tucatinib (TUC) is an oral, reversible, small-molecule tyrosine kinase inhibitor highly selective for human epidermal growth factor receptor 2 (HER2). TUC + trastuzumab (T) + capecitabine (C) is approved for patients (pts) with locally advanced or metastatic breast cancer (MBC) after ≥2 prior anti-HER2 treatment regimens based on statistically significant, clinically meaningful improvements in progression-free survival (PFS) and overall survival (OS) in pts with and without brain metastases (BM) in the randomized HER2CLIMB trial (NCT02614794).

Incidence and associated risk factors for invasive fungal infections and other serious infections in patients on ibrutinib

BackgroundIbrutinib is a small molecule tyrosine kinase inhibitor that blocks the activity of B cells and other immune effectors and is used in a variety of hematologic malignancies. There have been numerous reports of increased frequency of serious infections including invasive fungal infections (IFI) in patients on ibrutinib. MethodsDemographic and clinical features of all patients receiving ibrutinib at a single tertiary care center were collected from electronic medical records. Univariate and multivariate statistical analyses were performed to find out the factors associated with infection. ResultsA total of 244 patients received ibrutinib for hematologic malignancies, of which 44 (18.0%) experienced ≥ 1 serious infection including 5 (2.0%) with IFI (1 pulmonary cryptococcosis, 4 pulmonary aspergillosis), 39 (16.0%) with bacterial infections and 8 (3.3%) with viral infections. Ten patients (4.1%) experienced multiple infections or co-infections while on ibrutinib and 10 (4.1%) expired or were transferred to hospice as a result of infection. In multivariate analysis risk factors that were less common in uninfected versus infected patients included advanced age (73 years vs. 77 years), Eastern Cooperative Oncologic Grade (ECOG) performance score ≥ 2 (6.5% vs. 31.8%) and concurrent use of steroids (4.5% vs. 20.5%) or other cytotoxic agents (0% vs. 4.6%). ConclusionsThere was a high rate of serious infection but relatively few IFI in patients receiving ibrutinib. Most patients who developed serious infections while on ibrutinib had additional predisposing risk factors including concurrent use of steroids or other cytotoxic agents, advanced age and frailty.

Tyrosine kinase inhibitors-associated pyoderma gangrenosum, a systematic review of published case reports.

Pyoderma gangrenosum is a rare ulcerative dermatosis. It may be caused by some drugs, including small molecule tyrosine kinase inhibitors (TKIs). The aim of this study was to evaluate the reported evidence of pyoderma gangrenosum associated with the use of these drugs. A systematic electronic literature search of PubMed and Embase was conducted. In these databases, search terms describing pyoderma gangrenosum were combined with TKIs. Fifteen case reports (eight cases associated with sunitinib, two with imatinib, two with ibrutinib, one with gefitinib, one with pazopanib, and one with dabrafenib and trametinib) were identified over the 14 years. The average Naranjo score of these cases is 6.6, which indicates a probable adverse drug reaction. Pyoderma gangrenosum is a probable and reversible drug reaction associated with some TKIs. Detailed medical history can help to prompt diagnosis of drug-induced pyoderma gangrenosum. Clinicians should be aware of TKI-associated pyoderma gangrenosum when caring for the skin of oncologic patients undergoing therapy with kinase inhibitors.

Inhibition of human UDP-glucuronosyltransferase enzyme by Dabrafenib: Implications for drug-drug interactions.

Dabrafenib is a novel small molecule tyrosine kinase inhibitor (TKI) which is used to treat metastatic melanoma. The aim of this research was to survey the effects of dabrafenib on human UDP-glucuronosyltransferases (UGTs) and to evaluate the risk of drug-drug interactions (DDIs). The formation rates for 4-methylumbelliferone (4-MU) glucuronide and trifluoperazine-glucuronide in 12 recombinant human UGT isoforms with or without dabrafenib were measured and HPLC was used to investigate the inhibitory effects of dabrafenib on UGTs. Inhibition kinetic studies were also conducted. In vitro-in vivo extrapolation approaches were further used to predict the risk of DDI potentials of dabrafenib via inhibition of UGTs. Our data indicated that dabrafenib had a broad inhibitory effect on 4-MU glucuronidation by inhibiting the activities of UGTs, especially on UGT1A1, UGT1A7, UGT1A8, and UGT1A9, and dabrafenib could increase the area under the curve of co-administered drugs. Dabrafenib is a strong inhibitor of several UGTs and the co-administration of dabrafenib with drugs primarily metabolized by UGT1A1, 1A7, 1A8 or 1A9 may induce potential DDIs.

Abstract 1052: 6-phosphofructo-2-kinase enhances cytotoxicity of the EGFR inhibitor erlotinib via regulation of cell cycle in non-small lung cancer cell lines

Abstract Lung tumor development and aggressiveness is largely driven by a multitude of oncogenes including the epidermal growth factor receptor (EGFR). Targeted EGFR inhibition with small-molecule tyrosine kinase inhibitors (EGFR-TKIs) such as erlotinib have initially improved progression-free survival in patients with EGFR-driven NSCLCs but failed to provide an overall survival benefit due to the development of resistance. Clinically, responses to EGFR-TKIs are evaluated by positron emission tomography using 2[18F]- fluoro-2-deoxy-glucose (18FDG-PET) as tumors consume significantly more glucose compared to adjacent normal tissue in vivo. 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB3), a known driver of glycolytic flux, plays a key role in the metabolic shift that occurs during tumor progression. The product of PFKFB3, F2,6BP, regulates glucose metabolism by activating PFK1, which in turn facilitates glycolysis. Increased glycolysis fulfills the elevated tumor demands for energetic and biosynthetic precursors needed for survival and proliferation - the functions that confer resistance to targeted therapies. Here, we interrogated the hypothesis that PFKFB3 inhibition increases the efficacy of EGFR-TKIs in NSCLCs with different EGFR mutation status in vitro. We treated multiple NSCLCs models (wild type EGFR: H522, H1437; and mutated EGFR: PC9, HCC827) with erlotinib and observed a significant inhibition of cell proliferation in vitro. Erlotinib treatment resulted in G1 arrest, decreased expression and phosphorylation of cyclin dependent kinases (CDKs), Rb and accumulation of p27 in all NSCLCs. Notably, PFKFB3 silencing in TKI-treated cells dramatically decreased cell viability, indicating that PFKFB3 expression is required to maintain cell viability of G1 arrested cells. Further, co-treatment with the PFKFB3 inhibitor PFK-158 increased cell death in all NSCLCs. We found that inhibition of PFKFB3 reduced TKI-induced G1 arrest and altered cell cycle progression, switching the effect of erlotinib from cytostatic to cytotoxic. Importantly, dual treatment caused only minor changes in glycolytic flux when compared to the effects of PFK-158 or erlotinib alone suggesting that glucose metabolism did not contribute to the observed effect. Taken together, our findings suggest that a non-metabolic function of PFKFB3 limits the usefulness of EGFR-TKI in vitro and that targeting of PFKFB3 can be a promising approach to improve clinical efficacy of FDA-approved EGFR-TKIs in lung cancer patients. Citation Format: Nadiia Lypova, Lilibeth Lanceta, Jason Chesney, Yoannis Imbert-Fernandez. 6-phosphofructo-2-kinase enhances cytotoxicity of the EGFR inhibitor erlotinib via regulation of cell cycle in non-small lung cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1052.

MO32-4 A real-world multicentre retrospective study of low-dose apatinib for HER2-negative metastatic breast cancer

There are fewer treatments in for patients with HER2-negative breast cancer than for HER2-positive patients, especially for metastatic breast cancer. Apatinib is a small-molecule tyrosine kinase inhibitor that selectively targets VEGFR-2. Apatinib was prescribed orally at a dose of 250mg once daily. If conditions permitted, patients also received an NGS test before treatment. The median PFS of the 115 patients was 4.0 months. The objective response rate was 25.5%, and the disease control rate was 80.9%. Univariate analysis showed that patients without liver metastases before treatment were given longer PFS (4.4 months vs. 3.0 months, p=0.022), and that patients with hypertension or hand-foot syndrome during treatment had significantly longer PFS (hypertension: 4.4 vs 3.0 months, p=0.010; Hand-foot syndrome: 4.7 vs 3.3 months, p=0.016). NGS found that patients with BRCA mutations had significantly longer PFS (10.5 months versus 2.6 months, p=0.023). In addition, there were 15 patients having continued to treat with apatinib combined with another chemotherapy agent after PD for first time, the PFS slightly extended (5.2 vs. 4.0 months), but no statistical difference (p =6.22). Most of the adverse effects were grade 1 to 2. Low-dose apatinib is a new choice after failure of multi-line treatment for HER2-negative breast cancer, and the adverse reactions are well tolerated. Through NGS detection to find the predictive factors of apatinib, to the first time, we found that patients with BRCA mutations had longer PFS. Moreover, patients without liver metastasis received significantly longer PFS, suggesting the organ selectivity of apatinib treatment. It provides effective evidence for the clinical use of low-dose apatinib in HER2-negative advanced breast cancer. We believe that patients with HER2-negative advanced breast cancer should choose low-dose apatinib to increase the efficacy, especially those with BRCA mutations.

P-142 Safety and efficacy of apatinib for non-surgical advanced esophageal cancer: A real-world study

Esophageal cancer (EC) remains an aggressive cancer with short-term survival due to limited therapeutic options, and there is currently no effective treatment for patients after first-line chemotherapy failure. Apatinib is a small-molecule tyrosine kinase inhibitor that selectively inhibits vascular endothelial growth factor receptor-2. Our goal was to evaluate the effiicacy and safety of apatinib in patients with non-surgical esophageal cancer.

A Case of Chronic Lymphocytic Leukemia with Unmutated Variable Region of the Immunoglobulin Heavy Chain Gene with Markedly High Lactate Dehydrogenase Responding to Ibrutinib

Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults. Several prognostic factors exist that help stratify patients into different risk categories. Unmutated status of the immunoglobulin heavy chain (IGHV) gene has been shown to be among the most important high-risk prognostic factors in CLL. Elevated serum level of lactate dehydrogenase has also been shown to be associated with worst outcomes. Recently, the management of CLL has progressed favorably and improved outcomes were seen with the use of the small-molecule Bruton's tyrosine kinase inhibitor, ibrutinib, when compared with traditional chemotherapy. Here, we present a CLL case with an unmutated IGHV gene status presenting with markedly elevated LDH and responding dramatically to ibrutinib.

Progress in Treatment of Non-Small Cell Lung Cancer Harboring HER2 Aberrations.

Epidermal growth factor receptor 2 (HER2/ErbB2/neu), a member of ErbB receptor tyrosine kinase family, forms homo- or heterodimers with ErbB1 (HER1/EGFR), ErbB3 (HER3), or ErbB4 (HER4), to activate signal transduction pathways and promote proliferation, differentiation and tumorigenesis. Preliminary clinical trials of monoclonal antibodies, antibody conjugates and small-molecule tyrosine kinase inhibitors targeting HER2 have indicated that HER2 is a potential therapeutic target in non-small cell lung cancer (NSCLC). HER2 aberrations in NSCLC patients mainly include mutation, amplification, and overexpression. While there are significant differences in the outcome of NSCLC with these HER2 changes, no consensus has been reached for the incidence, detection method and targeted treatments for the three types of HER2 aberration. HER2 mutation is generally considered to have more clinical relevance and response to HER2-targeted therapies. In this review, we discuss HER2 alterations in NSCLC, including diagnostic challenges and treatment strategies particular to the HER2 mutation.

Surufatinib for the treatment of advanced extrapancreatic neuroendocrine tumors

ABSTRACT Introduction: Surufatinib (also known as HMPL-012, sulfatinib) is a novel oral tyrosine kinase inhibitor (TKI), which has the dual activity of anti-angiogenesis and immune regulation. In December 2020, surufatinib was approved as a monotherapy for unresectable locally advanced or metastatic, progressive nonfunctioning, well differentiated (grade 1 or 2) extrapancreatic neuroendocrine tumors (epNETs) in China. Areas covered: In this paper, the chemical properties, mechanism of action, pharmacokinetics, clinical efficacy, safety, and tolerability of surufatinib for treatment of advanced extrapancreatic NETs are introduced in detail. We performed a systematic review of the literature in PubMed and the following keywords were used: ‘surufatinib,’ ‘sulfatinib’ and ‘HMPL-012.’ Expert opinion: Surufatinib is a potent, selective, and small-molecule TKI that targets vascular endothelial growth factor receptor (VEGFR), fibroblast growth factor receptor 1 (FGFR1) and colony stimulating factor 1 receptor (CSF1R). Surufatinib showed an acceptable safety profile and encouraging antitumor activity in patients with advanced epNETs. The most frequently observed adverse events (AEs) were hypertension and proteinuria. Surufatinib provides a new treatment option for patients with advanced epNETs. More clinical trials of surufatinib are ongoing to develop a combination of therapy strategies and new indications for malignancies.

Bone Marrow Stromal Cell Mediated Resistance to Mertk Inhibition in Acute Myeloid Leukemia

MERTK is a receptor tyrosine kinase of the TAM family (TYRO-3, AXL, MERTK) that is aberrantly expressed in 80-100% of pediatric and adult primary acute myeloid leukemia (AML) samples. MERTK inhibition mediated by small molecule tyrosine kinase inhibitors (TKIs), UNC2025 or MRX-2843, altered cell cycle progression and induced apoptosis in leukemia cell cultures. Moreover, treatment with these MERTK TKIs prolonged survival in mouse xenograft models of acute leukemia, but was not curative. In these models, treatment with the TKI effectively reduced peripheral disease burden but was less effective in the bone marrow, suggesting a role for the bone marrow microenvironment in therapeutic resistance.To evaluate the role of the bone marrow stromal niche in mediating resistance to MERTK inhibition by MRX-2843, AML cell lines were cultured in the presence of a fibroblast-like cell line (HS27) and induction of cell death and apoptosis was determined by flow cytometry after treatment with MRX-2843 or vehicle. Co-culture with the HS27 cell line significantly reduced cell death in Kasumi-1 AML cell cultures in response to treatment with 200nM MRX-2843 compared to leukemia cells alone (31.9% versus 61.2%, p<0.05). Similar results were observed in NOMO-1 and OCI-AML5 AML cell cultures. Additionally, cell cycle profiles were determined by flow cytometry in the presence or absence of stromal cells and MRX-2843. Treatment of Kasumi-1 cell cultures with 200nM MRX-2843 in the absence of stromal cells resulted in a significant accumulation of cells in G2 phase relative to vehicle-treated cultures (8.9% versus 14.8%, p<0.01) and was accompanied by an increase in γH2AX expression detected by immunoblot. In contrast, there was no change in cell cycle progression or γH2AX expression in cultures treated with MRX-2843 when co-cultured with the stromal cell line. NOMO-1 cells also accumulated in G2 phase in response to treatment with MRX-2843 and were protected from changes in cell cycle progression by co-culture with HS27 cells. Additionally, MERTK inhibition resulted in induction of polyploidy in NOMO-1 cell cultures, but this effect was abrogated in the presence of stromal cells. Collectively these results indicate abrogation of cell cycle progression in leukemia cells in response to treatment with MRX-2843 and demonstrate protection from this effect mediated by stromal cell interactions.The MERTK ligand, GAS6, mediates resistance to cytotoxic chemotherapy in leukemia cells and is a poor prognostic marker in AML. To evaluate the role of the MERTK ligands GAS6 and PROS1 in resistance to MERTK inhibition, their expression in stromal cells was determined by immunoblot. GAS6, but not PROS1, was expressed in HS27 cells; however, MRX-2843 retained its ability to inhibit MERTK phosphorylation in stromal cell co-cultures. We hypothesized that upregulation of the other TAM kinases, AXL and/or TYRO3, may mediate resistance as they share common ligands. To test this hypothesis, Kasumi-1 cells were treated with UNC4241, a small molecule pan-TAM inhibitor, in the presence of stromal cells. While co-culture protected Kasumi-1 cells from MRX-2843 induced apoptosis, treatment with UNC4241 induced apoptosis in the presence of stromal cells (32.7% versus 20.4% in vehicle control, p<0.01).To further evaluate the interactions between stromal cells and leukemia cells, Kasumi-1 cells were cultured in HS27-conditioned medium in the presence or absence of MRX-2843. Conditioned medium was not sufficient to provide protection from MRX-2843 induced apoptosis (74.0% vs. 69.8% in unconditioned medium, p>0.05). Similarly, addition of 500nM exogenous GAS6 or PROS1 did not protect leukemia cells from apoptosis in the presence of MRX-2843. Together, these results suggest that direct cell-cell contact between leukemia and stromal cells is necessary to provide protection.Together these data support a model whereby direct cell-cell contact with stromal cells in the bone marrow niche protects leukemia cells from MRX-2843 induced apoptosis and cell cycle alterations and suggest that bone marrow mobilizing agents may be effective to overcome resistance to MERTK inhibitors. DisclosuresWang:Meryx: Equity Ownership; University of North Carolina: Patents & Royalties. Frye:University of North Carolina: Patents & Royalties; Meryx: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Earp:Meryx: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. DeRyckere:Meryx: Equity Ownership. Graham:Meryx: Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

The Novel, Irreversible FLT3 Inhibitor FF-10101 Demonstrates Broad in Vitro Activity Against FLT3 Inhibitor Resistance Mutations

Background: Small-molecule FLT3 tyrosine kinase inhibitors (TKIs) have improved remission rates in relapsed/refractory AML driven by FLT3 internal tandem duplication (ITD) mutations, although nearly all responders eventually relapse. Acquired clinical resistance to type II inhibitors such as quizartinib and sorafenib has largely been due to kinase domain (KD) mutations that destabilize drug binding. Early inhibitors, limited by short half-life and fluctuating plasma levels, were also only able to transiently inhibit FLT3. FF-10101 is a first-in-class, type I inhibitor that covalently binds FLT3 irreversibly, suggesting its activity should be limited by the half-life of FLT3 itself. In a profiling assay of 216 human recombinant kinases FF-10101 showed selectivity for wild type FLT3 and FLT3 D835Y. It also demonstrated potent activity against FLT3-ITD AML cell lines harboring TKI-resistant D835Y and Y842C/H mutations. Here, we report in vitro testing of FF-10101 against 19 additional FLT3 TKI-resistant mutations, as well as six cysteine-to-serine mutations within the KD that could impact covalent drug binding. Profiling of this activity is relevant as FF-10101 enters single-agent phase 1/2a clinical trial in relapsed/refractory FLT3 mutant AML.Results: FF-10101 was profiled in an in vitro viability assay against murine BaF3 and human AML cell lines harboring a spectrum of mutations. FF-10101 inhibited the proliferation of FLT3-driven Molm14 and MV4;11 cells (IC50 4.1 and 1.1 nM, respectively). BaF3 FLT3-ITD cells were comparably inhibited (IC50 3.1 ± 1.6 nM), while growth of BCR-ABL-driven K562 cells were unaffected. To confirm covalent binding of FF-10101, we then analyzed its activity against all cysteine residues in the FLT3 KD. Every C->S substitution except C695S remained sensitive to FF-10101, confirming C695 as the FF-10101 binding site and suggesting C695S may impart clinical resistance to FF-10101. We next assessed 10 BaF3 FLT3-ITD cell lines harboring mutations at D835 and the gatekeeper F691, given discovery of resistance mutations at these sites in patients treated with AC220 and sorafenib. FF-10101 inhibited F691L and all substitutions/deletions at the D835 residue. N676K, a mutation found in a patient at relapse on midostaurin, was also potently inhibited. Notably, F691L previously exhibited relative resistance to crenolanib, a reversible, type I FLT3 inhibitor. Beyond clinically described mutants, we also tested FF-10101 against six mutations previously identified in mutagenesis screening assays. A D698N mutation found in separate crenolanib and PLX3397 mutagenesis assays demonstrated relative resistance to crenolanib but was sensitive to FF-10101 (IC50 8.4 ± 7.8 nM), as were four D839 amino acid substitutions identified in the PLX3397 mutagenesis assay alone. Alternatively, a Y693C mutation found in a crenolanib mutagenesis screen remained relatively insensitive to FF-10101. Finally, two resistance mutations co-occurring in the same KD are well-described in chronic myelogenous leukemia (CML) patients treated with sequential TKI therapy. While not yet found in AML patients treated similarly, we hypothesize that analogous compound mutations could emerge. As such, we tested FF-10101 against BaF3 FLT3-ITD cells containing three compound mutations. Unlike crenolanib, FF-10101 potently inhibited D698N/D835V, but F691L/D835V and Y693C/D835V each exhibited relatively moderate resistance to both drugs. Analysis of additional activation loop (AL) mutations (N831H/K, R845G, G846R, Y842S) and biochemical studies of FLT3 phosphorylation and downstream effector pathways in response to FF10101 is ongoing and will be presented.Conclusions: FF-10101 is a type I, irreversible FLT3 inhibitor that has demonstrated potent activity against all mutations associated with clinical resistance to FLT3 TKIs. Expanded profiling shows anti-proliferative activity against nearly all mutations associated with FLT3 TKI resistance except for C695S (the site of FF-10101 binding) and the crenolanib-resistant Y693C mutation, which has yet to be associated with clinical TKI resistance. These data suggest FF-10101 has potential to be clinically active against the majority of FLT3 TKI resistant KD mutations. FF-10101 is currently beginning phase I/2a clinical assessment as a single agent in patients with relapsed/refractory AML harboring a FLT3 mutation. [Display omitted] DisclosuresSmith:Fujifilm: Research Funding; Astellas: Research Funding.

Nanotechnology of Tyrosine Kinase Inhibitors in Cancer Therapy: A Perspective.

Nanotechnology is an important application in modern cancer therapy. In comparison with conventional drug formulations, nanoparticles ensure better penetration into the tumor mass by exploiting the enhanced permeability and retention effect, longer blood circulation times by a reduced renal excretion and a decrease in side effects and drug accumulation in healthy tissues. The most significant classes of nanoparticles (i.e., liposomes, inorganic and organic nanoparticles) are here discussed with a particular focus on their use as delivery systems for small molecule tyrosine kinase inhibitors (TKIs). A number of these new compounds (e.g., Imatinib, Dasatinib, Ponatinib) have been approved as first-line therapy in different cancer types but their clinical use is limited by poor solubility and oral bioavailability. Consequently, new nanoparticle systems are necessary to ameliorate formulations and reduce toxicity. In this review, some of the most important TKIs are reported, focusing on ongoing clinical studies, and the recent drug delivery systems for these molecules are investigated.

Digital PCR detection of EGFR somatic mutations in non-small-cell lung cancer formalin fixed paraffin embedded samples

BackgroundDigital PCR (dPCR) is proposed to replace real time PCR and Sanger sequencing for detection and quantification of rare mutations, frequently unnoticed in the mass of tumoral cells. Screening of endothelial growth factor receptor (EGFR) mutations is mandatory before treatment with EGFR-targeted therapy with small-molecule tyrosine kinase inhibitors, which has been approved for the treatment of advanced non-small-cell lung cancer (NSCLC). ObjectiveIn order to establish a cost-effective method for detection of mutations, we optimized dPCR identification of EGFR mutations in exons 18–21, and determined dPCR sensitivity, limits of detection (LoD) and quantification (LoQ). MethodsFor clinical validation, we compared the performance of dPCR and castPCR in 57 NSCL formalin fixed paraffin embedded samples and 10 lung cancer-free formalin fixed paraffin embedded samples. ResultsEGFR mutations DEL19, p.L858R, p.G719X, p.L861Q and p.T790 M were detected by dPCR in 27 samples versus 11 detected by castPCR (p = 0.014). LoD was determined as 100 molecules of DNA/uL and LoQ as 1%. Most of the samples (87%) identified by competitive Allele-Specific TaqMan (castPCR) as wild-type and by dPCR as mutated, presented less than 10% mutated DNA molecules (mean 4.57%). Accuracy of dPCR was 94.44%, as measured with the assay recommended by the College of American Pathologists. ConclusionThese results indicated higher sensibility and specificity of dPCR for screening EGFR mutations in NSCLC biopsies, compared to castPCR.

Apatinib enhances the anti-tumor effect of paclitaxel via the PI3K/p65/Bcl-xl pathway in triple-negative breast cancer.

BackgroundApatinib is a new generation of small molecule tyrosine kinase inhibitor, which can highly selectively inhibit phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR-2). This study aimed to investigate the synergistic effects of apatinib and paclitaxel (PTX) on triple-negative breast cancer (TNBC) in vivo and in vitro, and to explore the molecular mechanism of the PI3K/p65/Bcl-xl pathway.MethodsIn vitro, 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide (MTT) method, flow cytometry (FCM), wound healing assay, and transwell matrix assay were conducted to measure the effects of apatinib and PTX on cell viability, apoptosis, migration, and invasion in TNBC cell line MDA-MB-468. Western blot (WB) was conducted to detect protein expression levels of PI3K, p65, and Bcl-xl after the application of apatinib and PTX. In vivo, MDA-MB-468 tumor-bearing nude mice were treated with apatinib and PTX, and tumor growth was observed.ResultsIn vitro, apatinib and PTX could synergistically suppress the cell viability, the combined group had the most obvious effect. Apatinib and PTX could promote apoptosis and suppress migration and invasion of TNBC cells. Apatinib could reduce the expression of p-PI3K, p65, and Bcl-xl proteins (P<0.05). In vivo, apatinib and PTX could inhibit tumor size and weight of model mice, and the combined agents had a more significant effect.ConclusionsApatinib could enhance the anti-tumor effect of PTX on TNBC cells through the PI3K/p65/Bcl-xl molecular pathway, and apatinib combined with PTX might be a promising option for TNBC treatment.

A Rapid and Selective UPLC-MS/MS Assay for Accurate Analysis of Apatinib in Rat Plasma and its Application to a Pharmacokinetic Study

Objective: Apatinib, a novel small-molecule Tyrosine Kinase Inhibitor (TKI), is under development to treat advanced gastric cancer. For the pharmacokinetic evaluation and routine drug monitoring of apatinib, a quantitative ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) method in rat plasma was developed with tinidazole used as an internal standard (IS). Methods:: Protein precipitation (PPT) was selected as a sample pre-treatment method to extract apatinib. Then, chromatography was performed on a Kinetex C8 column (2.1×100 mm, 2.6 μm) using a constant mobile phase including 0.2% formic acid and 10 mM ammonium acetate in water and methanol (30:70, v/v) with a gradient flow rate from 0.2 mL/min to 0.4 mL/min. Chromatographic analysis was performed in only 4.5 min. Mass spectrometric detection was carried on positive electrospray ionization (ESI+) mode with Multiple-Reaction Monitoring (MRM). Results: The standard calibration curve showed good linearity in 2-1000 ng/mL with the correlation coefficient (R2) &gt; 0.99. The Lower Limit of Quantitation (LLOQ) was 2 ng/mL. The precision, accuracy, extraction recovery, matrix effect, stability and carryover were all within the acceptable range. Conclusion: This method was simple, accurate, selective and successfully used for a pharmacokinetic study following seven rats orally administrated a single of 60 mg/kg apatinib.

Immunotherapy in Treating EGFR-Mutant Lung Cancer: Current Challenges and New Strategies.

Lung cancer is the leading cause of cancer-related deaths worldwide. Immune checkpoint inhibitors, including monoclonal antibodies against programmed death-1 (PD-1) and programmed death ligand-1 (PD-L1), have dramatically improved the survival and quality of life of a subset of non-small cell lung cancer (NSCLC) patients. Multiple predictive biomarkers have been proposed to select the patients who may benefit from the immune checkpoint inhibitors. EGFR-mutant NSCLC is the most prevalent molecular subtype in Asian lung cancer patients. However, patients with EGFR-mutant NSCLC show poor response to anti-PD-1/PD-L1 treatment. While small-molecule EGFR tyrosine kinase inhibitors (TKIs) are the preferred initial treatment for EGFR-mutant NSCLC, acquired drug resistance is severely limiting the long-term efficacy. However, there is currently no further effective treatment option for TKIs-refractory EGFR-mutant NSCLC patients. The reasons mediating the poor response of EGFR-mutated NSCLC patients to immunotherapy are not clear. Initial investigations revealed that EGFR-mutated NSCLC has lower PD-L1 expression and a low tumor mutational burden, thus leading to weak immunogenicity. Moreover, the use of PD-1/PD-L1 blockade prior to or concurrent with osimertinib has been reported to increase the risk of pulmonary toxicity. Furthermore, emerging evidence shows that PD-1/PD-L1 blockade in NSCLC patients can lead to hyperprogressive disease associated with dismal prognosis. However, it is difficult to predict the treatment toxicity. New biomarkers are urgently needed to predict response and toxicity associated with the use of PD-1/PD-L1 immunotherapy in EGFR-mutated NSCLC. Recently, promising data have emerged to suggest the potentiation of PD-1/PD-L1 blockade therapy by anti-angiogenic agents and a few other novel therapeutic agents. This article reviews the current investigations about the poor response of EGFR-mutated NSCLC to anti-PD-1/PD-L1 therapy, and discusses the new strategies that may be adopted in the future.

Application of neratinib in the treatment of hormone receptor and HER-2 double positive breast cancer

Hormone receptor (HR), human epidermal growth factor receptor-2(HER-2) double positive breast cancer is a highly aggressive disease, and it is more susceptible to drug resistance and metastasis. Currently, although anti-HER-2 monoclonal antibody combined with endocrine therapy is commonly used in clinical practice, patients with HR-positive and HER-2-positive still have a poor prognosis. That may be caused by the activation of downstream PI3K/Akt/mTOR signaling pathway of HER-2. Therefore, simultaneously blocking the extracellular, intracellular and downstream PI3K/Akt/mTOR signaling pathway of HER-2, as well as the dual blockade of HER-2 and HR, is more conducive to the treatment of HR positive and HER-2 positive breast cancer. The small-molecule tyrosine kinase inhibitor neratinib not only inhibits the phosphorylation of HER-1, HER-2 and HER-4, but also blocks the downstream signal transduction of PI3K/Akt and Ras/Raf/MEK/ERK pathways. The intensive adjuvant chemotherapy studies has confirmed that nelatinib can improve the prognosis of patients with HER-2 positive breast cancer, especially those with HR and HER-2 double positive patients. Nelatinib has also achieved positive effect on the treatment of advanced breast cancer, but relevant data on the treatment of HR-positive and HER-2 positive advanced breast cancer are still lacking. Lapatinib, a small-molecule HER inhibitor similar to neratinib, has shown some effects on the treatment of HR-positive and HER-2 positive patients with advanced breast cancer. Given that nelatinib has a superior efficacy than lapatinib, it is expected to provide more ideas and options for the treatment strategies of breast cancer.

Pyrotinib plus capecitabine for HER2-positive metastatic breast cancer patients with brain metastases (PERMEATE): A multicenter, single-arm phase II study.

1037 Background: HER2-positive metastatic breast cancer (BC) has a high risk of brain metastases (BM), leading to poor survival. Small molecule tyrosine kinase inhibitor (TKI) with enhanced penetrability to the blood brain barrier combined with capecitabine have demonstrated promising clinical outcomes in HER2-positive metastatic BC patients with untreated (such as lapatinib) or previously treated (such as neratinib) BM. The randomized phase III PHOEBE trial has proved better efficacy of pyrotinib, an irreversible pan-HER receptor TKI, versus lapatinib when in combination with capecitabine in HER2-positive local relapsed or metastatic BC. This study was conducted to investigate the efficacy and safety of pyrotinib plus capecitabine in HER2-positive metastatic BC patients with BM. Methods: In this multicenter phase II trial (NCT03691051), eligible patients received pyrotinib 400 mg orally once daily without breaks and capecitabine 1000 mg/m2 orally twice daily for 14 days followed by 7 days off. Treatment was continued until disease progression or intolerable toxicity. Prior HER2 TKIs were not allowed. Cohort A included patients with radiotherapy-naive BM, and cohort B included those with progressive BM after whole brain radiotherapy or stereotactic conformal radiotherapy. The primary endpoint was confirmed central nervous system (CNS) objective response rate (ORR), as assessed according to the Response Evaluation Criteria In Solid Tumors version 1.1. Results: Between January 2018 and July 2020, a total of 78 female patients were included (Table). For cohort A (n = 59), the CNS ORR was 74.6% (95%CI: 61.6%-85.0%). For cohort B (n = 19), the CNS ORR was 42.1% (95%CI: 20.3%-66.5%). By the cutoff date on 25 January 2021, the median progression-free survival was 12.1 months (95%CI: 9.0-14.7) in cohort A and 5.6 months (95%CI: 3.4-10.7) in cohort B. The most common grade ≥3 adverse events were diarrhea (23.1% [18/78]), neutrophil count decreased (12.8% [10/78]), white blood cell count decreased (12.8% [10/78]), anemia (9.0% [7/78]), hand-foot syndrome (7.7% [6/78]), hypertriglyceridemia (6.4% [5/78]), and hypokalemia (5.1% [4/78]). Conclusions: Pyrotinib plus capecitabine resulted as an effective and safe treatment for HER2-positive BC patients with radiotherapy-naive BM, but the efficacy was modest in those with radiotherapy-treated BM. Clinical trial information: NCT03691051 .[Table: see text]