Dabrafenib Research Articles

Optimization and Enhancement of Oral Bioavailability of Dabrafenib as Nanobubbles Using Quality by Design Approach

Background: The use of Poly (lactic-co-glycolic acid) (PLGA) nanobubbles (NBs) aimed at functioning as a delivery system that encounter solubility issues for drugs like dabrafenib (DBF), which belonged to the Biopharmaceutical Classification System (BCS) class II category. These specially designed nanobubbles enhanced the drug’s solubility, stability, and bioavailability, thus improving the therapeutic effectiveness. Moreover, they offered controlled release characteristics and can potentially enhance drug delivery to tissues or cells, maximizing pharmacological results while reducing adverse effects. Methods: PLGA NBs were formulated using solvent evaporation and optimized using a Box Behnken design considering process and formulation parameters. The NBs characterization includes particle size, drug loading, entrapment efficiency, in-vitro studies, haemolytic studies, Fourier transform infrared (FTIR) spectroscopy, differential scanning calorimeter (DSC), stability studies, and as well as in-vivo studies in rats. Results: The optimized nanobubbles (NBs) displayed a particle size (PS) of 190.6 ± 18.4 nm, zeta potential of -21 ± 4.2 mV, and polydispersity index (PDI) of 0.397 ± 0.096. With 87.21 ± 3.8% of entrapment efficiency (EE) and 26.29 ± 4.01% drug loading, in-vitro studies revealed a superior drug release (99%) with ultrasound versus plain drugs (20%). FTIR and DSC studies confirmed no drug-polymer interaction. Scanning Electron Microscopy (SEM) images displayed uniform spherical nanosized particles. Haemolytic activity demonstrated safety, and stability studies indicated no significant changes after 30 days. The nanobubbles exhibited increased Cmax (4.74) and AUC0-t (6.82), thereby promising an enhanced solubility, absorption, and extended half-life. Conclusion: The current investigation showed that PLGA nanobubbles loaded with dabrafenib have a promising delayed release potential, which might make them a possible treatment alternative for breast cancer.

Probing Short-Range Interactions in Isostructural Hydrate and Perhydrate of Dabrafenib by Magic-Angle Spinning Solid-State NMR.

Dabrafenib (DBF), an anticancer drug, exhibits isostructural properties in its hydrate (DBF⊃H2O) and perhydrate (DBF⊃H2O2) forms, as revealed by single-crystal X-ray diffraction. Despite the H2O and H2O2 solvent molecules occupying identical locations, the two polymorphs have different thermal behaviors. In general, determination of stoichiometry of H2O in the perhydrate crystals is difficult due to the presence of both H2O and H2O2 in the same crystal voids. This study utilizes magic-angle spinning (MAS) solid-state NMR (SSNMR) combined with gauge-included projector augmented wave calculations to characterize the influence of solvent molecules on the local environment in pseudopolymorphs. SSNMR experiments were employed to assign 1H, 13C, and 15N peaks and identify spectral differences in the isostructural pseudopolymorphs. Proton spectroscopy at fast MAS was used to identify and quantify H2O2/H2O in DBF⊃H2O2 (mixed hydrate/perhydrate). 1H-1H dipolar-coupling-based experiments were recruited to confirm the 3D molecular packing of solvent molecules in DBF⊃H2O and DBF⊃H2O2. Homonuclear (1H-1H) and heteronuclear (1H-14N) distance measurements, in conjunction with diffraction structures and optimized hydrogen atom positions by density functional theory, helped decipher local interactions of H2O2 with DBF and their geometry in DBF⊃H2O2. This integrated X-ray structure, quantum chemical calculations, and NMR study of pseudopolymorphs offer a practical approach to scrutinizing crystallized solvent interactions in the crystal lattice even without high-resolution crystal structures or artificial sample enrichment.

Simulating BRAFV600E-MEK-ERK signalling dynamics in response to vertical inhibition treatment strategies

In vertical inhibition treatment strategies, multiple components of an intracellular pathway are simultaneously inhibited. Vertical inhibition of the BRAFV600E–MEK-ERK signalling pathway is a standard of care for treating BRAFV600E-mutated melanoma where two targeted cancer drugs, a BRAFV600E-inhibitor, and a MEK inhibitor, are administered in combination. Targeted therapies have been linked to early onsets of drug resistance, and thus treatment strategies of higher complexities and lower doses have been proposed as alternatives to current clinical strategies. However, finding optimal complex, low-dose treatment strategies is a challenge, as it is possible to design more treatment strategies than are feasibly testable in experimental settings. To quantitatively address this challenge, we develop a mathematical model of BRAFV600E–MEK-ERK signalling dynamics in response to combinations of the BRAFV600E-inhibitor dabrafenib (DBF), the MEK inhibitor trametinib (TMT), and the ERK-inhibitor SCH772984 (SCH). From a model of the BRAFV600E–MEK–ERK pathway, and a set of molecular-level drug–protein interactions, we extract a system of chemical reactions that is parameterised by in vitro data and converted to a system of ordinary differential equations (ODEs) using the law of mass action. The ODEs are solved numerically to produce simulations of how pathway-component concentrations change over time in response to different treatment strategies, i.e., inhibitor combinations and doses. The model can thus be used to limit the search space for effective treatment strategies that target the BRAFV600E–MEK–ERK pathway and warrant further experimental investigation. The results demonstrate that DBF and DBF–TMT–SCH therapies show marked sensitivity to BRAFV600E concentrations in silico, whilst TMT and SCH monotherapies do not.

Dabrafenib-Panobinostat Salt: Improving the Dissolution Rate and Inhibition of BRAF Melanoma Cells.

Cocrystallization of the drug-drug salt-cocrystal of the histone deacetylase inhibitor (HDACi) panobinostat (PAN) and b-rapidly accelerated fibrosarcoma (BRAF) inhibitor dabrafenib (DBF) afforded single crystals of a two-drug salt stabilized by N+-H···O and N+-H···N- hydrogen bonds between the ionized panobinostat ammonium donor and dabrafenib sulfonamide anion acceptor in a 12-member ring motif. A faster dissolution rate for both drugs was achieved through the salt combination compared to the individual drugs in an aqueous acidic medium. The dissolution rate exhibited a peak concentration (Cmax) of approximately 310 mg cm-2 min-1 for PAN and 240 mg cm-2 min-1 for DBF at a Tmax of less than 20 min under gastric pH 1.2 (0.1 N HCl) compared to the pure drug dissolution values of 10 and 80 mg cm-2 min-1, respectively. The novel and fast-dissolving salt DBF-·PAN+ was analyzed in BRAFV600E melanoma cells Sk-Mel28. DBF-·PAN+ reduced the dose-response from micromolar to nanomolar concentrations and lowered IC50 (21.9 ± 7.2 nM) by half compared to PAN alone (45.3 ± 12.0 nM). The enhanced dissolution and lower survival rate of melanoma cells show the potential of novel DBF-·PAN+ salt in clinical evaluation.

Some things old, new and borrowed: Delivery of dabrafenib and vemurafenib to melanoma cells via self-assembled nanomicelles based on an amphiphilic dendrimer

Two clinically approved anticancer drugs targeting BRAF in melanoma patients – dabrafenib (DAB) and vemurafenib (VEM) – have been successfully encapsulated into nanomicelles formed upon self-assembly of an amphiphilic dendrimer AD based on two C18 aliphatic chains and a G2 PAMAM head. The process resulted in the formation of well-defined (∼10 nm) core-shell nanomicelles (NMs) with excellent encapsulation efficiency (∼70% for DAB and ∼60% for VEM) and good drug loading capacity (∼27% and ∼24% for DAB and VEM, respectively). Dynamic light scattering (DLS), transmission electron microscopy (TEM), small-angle x-ray scattering (SAXS), nuclear magnetic resonance (NMR), isothermal titration calorimetry (ITC), and molecular simulation (MS) experiments were used, respectively, to determine the size and structure of the empty and drug-loaded nanomicelles (DLNMs), along with the interactions between the NMs and their cargoes. The in vitro release data revealed profiles governed by Fickian diffusion; moreover, for both anticancer molecules, an acidic environment (pH = 5.0) facilitated drug release with respect to physiological pH conditions (pH = 7.4). Finally, both DAB- and VEM-loaded NMs elicited enhanced response with respect to free drug treatments in 4 different melanoma cell lines.

Tumor suppressor miR-193a-3p enhances efficacy of BRAF/MEK inhibitors in BRAF-mutated colorectal cancer.

Patients with BRAF‐mutated colorectal cancer (CRC) have a poor prognosis despite recent therapeutic advances such as combination therapy with BRAF, MEK, and epidermal growth factor receptor (EGFR) inhibitors. To identify microRNAs (miRNAs) that can improve the efficacy of BRAF inhibitor dabrafenib (DAB) and MEK inhibitor trametinib (TRA), we screened 240 miRNAs in BRAF‐mutated CRC cells and identified five candidate miRNAs. Overexpression of miR‐193a‐3p, one of the five screened miRNAs, in CRC cells inhibited cell proliferation by inducing apoptosis. Reverse‐phase protein array analysis revealed that proteins with altered phosphorylation induced by miR‐193a‐3p were involved in several oncogenic pathways including MAPK‐related pathways. Furthermore, overexpression of miR‐193a‐3p in BRAF‐mutated cells enhanced the efficacy of DAB and TRA through inhibiting reactivation of MAPK signaling and inducing inhibition of Mcl1. Inhibition of Mcl1 by siRNA or by Mcl1 inhibitor increased the antiproliferative effect of combination therapy with DAB, TRA, and anti‐EGFR antibody cetuximab. Collectively, our study demonstrated the possibility that miR‐193a‐3p acts as a tumor suppressor through regulating multiple proteins involved in oncogenesis and affects cellular sensitivity to MAPK‐related pathway inhibitors such as BRAF inhibitors, MEK inhibitors, and/or anti‐EGFR antibodies. Addition of miR‐193a‐3p and/or modulation of proteins involved in the miR‐193a‐3p–mediated pathway, such as Mcl1, to EGFR/BRAF/MEK inhibition may be a potential therapeutic strategy against BRAF‐mutated CRC.

Abstract 2361: The functional roles of miRNA-193a-3p in colorectal cancer

Abstract Background: Patients with BRAF-mutated colorectal cancer (CRC) have a poor prognosis despite recent therapeutic advances such as combination therapy with BRAF, MEK, and EGFR inhibitors. Aim: Our aim was to clarify the functions of the microRNAs (miRNAs) involved in oncogenesis and sensitivity to BRAF inhibitors and MEK inhibitors in BRAF-mutated CRC. Methods and Results: To identify miRNAs that sensitize a BRAF inhibitor dabrafenib (DAB) and a MEK inhibitor trametinib (TRA), we screened 240 miRNAs in BRAF-mutated CRC cell line RKO. Among the screened miRNAs, we tried to elucidate the novel functional roles of miR-193a-3p in CRC cell lines. We found that miR-193a-3p overexpression inhibited cell proliferation in CRC cell lines. The protein array analysis revealed that protein phosphorylation levels were altered by miR-193a-3p overexpression in various pathways such as cell-cycle-related pathways and PI3K-AKT pathways. The enhanced cytotoxicity by miR-193a-3p in addition to DAB and TRA was validated in multiple BRAF-mutated cell lines. Furthermore, the enhanced cytotoxicity by miR-193a-3p in addition to DAB and TRA was shown to be through inhibiting reactivation of MAPK signaling. This enhanced cytotoxicity by miR-193a-3p was even observed under treatment of DAB, TRA, and an anti-EGFR antibody. Conclusions: Our study shows that miR-193a-3p regulates multiple proteins involved in oncogenesis, and proposes that the combination therapy of miR-193a-3p and BRAF/MEK inhibitors could be a promising treatment strategy in BRAF-mutated CRC. Citation Format: Sakura Hiraide, Masanobu Takahashi, Yuya Yoshida, Keigo Komine, Chikashi Ishioka. The functional roles of miRNA-193a-3p in colorectal cancer [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 2361.

Primary Malignant Melanoma of the Esophagus: Prognosis and Therapeutic Challenges

Primary malignant melanoma of the esophagus (PMME) is a very rare and highly aggressive tumor, representing <1% of esophageal malignancies. We are reporting a case of PMME in a 54-year-old White, Non-Hispanic male, who presented to the hospital with dysphagia and weight loss. Computed tomography (CT) of the chest revealed a large mid-to-distal esophageal mass with mass effect on adjacent structures, with no evidence of invasion. The patient subsequently underwent esophagogastroduodenoscopy (EGD) with guided biopsy of the esophageal mass. Histopathology and immunohistochemistry of the biopsied specimen were performed and consistent with malignant melanoma. Tumor staging by positron emission tomography and computed tomography (PET/CT) scans revealed extensive, locally advanced disease with lymph node involvement. Subsequently, esophageal ultrasound (EUS) with biopsy of the involved lymph node was performed and was positive for malignant melanoma. The mass was deemed unresectable due to the extensive degree of locally advanced disease. To assist in our therapeutic decision making, the patient was screened for the presence of a BRAF V600 mutation. He was then started on a combination of the BRAF inhibitor (dabrafenib) in combination with the mitogen-activated extracellular kinase (MEK) Inhibitor (trametinib) given the presence of a BRAF V600 mutation.

LIMK1 promotes peritoneal metastasis of gastric cancer and is a therapeutic target

Peritoneal metastasis is a common form of metastasis among advanced gastric cancer patients. In this study, we reported the identification of LIM domain kinase 1 (LIMK1) as a promoter of gastric cancer peritoneal metastasis, and its potential to be a therapeutic target of dabrafenib (DAB). Using transcriptomic sequencing of paired gastric cancer peritoneal metastasis, primary tumors, and normal gastric tissues, we first unveiled that LIMK1 is selectively up-regulated in metastatic tumors. Increased LIMK1 in gastric cancer peritoneal metastasis was validated by immunohistochemistry analysis of an independent patient cohort. In vitro functional studies demonstrated that LIMK1 knockout or knockdown significantly inhibited cell migration and invasion of gastric cancer cells. LIMK1 knockout also abrogated peritoneal and liver metastases of gastric cancer cells in nude mice in vivo. Dabrafenib, a small molecule targeting LIMK1, was found to decrease cell migration and invasion of gastric cancer cells in vitro and abolish peritoneal and liver metastasis formation in vivo. Mechanistically, either LIMK1 knockout or Dabrafenib inhibited LIMK1 expression and phosphorylation of its downstream target cofilin. Taken together, our results demonstrated that LIMK1 functions as a metastasis promoter in gastric cancer by inhibiting LIMK1-p-cofilin and that Dabrafenib has the potential to serve as a novel treatment for gastric cancer peritoneal metastasis.

Determination of dabrafenib and trametinib in serum by dispersive solid phase extraction with multi-walled carbon nanotubes and capillary electrophoresis coupled to ultraviolet/visible detection

Dispersive solid phase extraction (DSPE) prior to capillary electrophoresis with ultraviolet and visible detection was used for the first time for the determination of dabrafenib (DB) and trametinib (TT), the main drugs used for the treatment of metastatic melanoma, in serum. Six different magnetic and non-magnetic nanomaterials were evaluated as DSPE sorbents for the extraction and preconcentration of the analytes. The best results were obtained with 4 mg of pristine multi-walled carbon nanotubes in phosphate buffer (pH 12.5; 10 mM). The electrophoretic separation was carried out in a fused silica capillary (30 cm × 75 µm i.d.) at 22 °C in a voltage ramp (8–12 kV) with a background electrolyte consisting of borate buffer (pH 9.2; 25 mM), 37 mM sodium dodecylsulfate, 22% (v/v) acetonitrile and 3% (v/v) methanol.The preconcentration factor achieved was 4, and the limits of detection were 0.012 and 0.008 mg L−1 for DB and TT in serum, respectively. Linearity was observed up to 5 mg/L and precision (n = 7) was below 3% in migration time and 7.8% in peak areas. The present method was applied to six patients under treatment with DB and TT for metastatic melanoma. It requires a less sophisticated instrumentation and lower volume of harmful organic solvents than the methods reported up to present.

Metastatic Ovarian Tumor of Vulvar Malignant Melanoma in a 43-Year-Old Woman: A Case Report and Literature Review

An ovarian malignant melanoma sometimes occurs from ovarian teratoma. Ovarian metastatic malignant melanoma is extremely rare. We describe a patient in whom vulvar melanoma (previously resected) metastasized to the ovary, making ovarian metastatic malignant melanoma. A 43-year-old Japanese woman was referred to us because of left ovarian tumor. She had undergone resection for malignant melanoma on the right labia minora with inguinal lymph node metastasis (pT1bN1aM0, stage IIIA, FIGO 2008). Eighteen months after this surgery, CT scans revealed left ovarian tumor and swelled pelvic lymph nodes, with a pelvic examination disclosing a left adnexal solid mass, with normal serum CA125 level (21.7 U/mL). Laparotomy revealed a left solid ovarian tumor measuring 4 cm, which was covered with a smooth grayish capsule. The right ovary, uterus, and pelvic cavity appeared normal. Upon sectioning during the surgery, the cross-sectional surface of the left ovary revealed a dark brown solid tumor. Following an intra-operative frozen-section diagnosis as metastatic melanoma, total hysterectomy with bilateral salpingo-oophorectomy and pelvic lymph dissection was performed. Histological examination confirmed the diagnosis as malignant melanoma metastasis to the left ovary and the obturator lymph node: the same laterality (left) as the primary site. The tumor was entirely composed of malignant melanoma cells with no evidence of teratoma. Combined chemotherapy with dabrafenib mesylate and trametinib was planned based on the positive BRAF mutations. This case highlights the importance that physicians should have high index of suspicion for the occurrence of ovarian melanoma metastasis after melanoma surgery. We also made extensive literature review on this issue, of which description may contribute to better understanding of this condition.

Malignant gastrointestinal neuroectodermal tumor with BRAF mutation and a history of malignant melanoma: A case report.

Malignant gastrointestinal neuroectodermal tumors (GNETs), also called clear-cell sarcoma-like tumors of the gastrointestinal tract, are rare and highly aggressive tumors originating in the gastrointestinal tract. These tumors are generally immunohistochemically positive for S-100 protein (S-100) and SRY-related HMG-box 10 (SOX10), and often contain EWSR1-ATF1 or EWSR1-CREB1. The histological features of GNETs overlap with those of clear-cell sarcoma of the tendons and aponeuroses. However, GNETs immunohistochemically lack melanocyte-specific markers and often demonstrate positivity for CD56, synaptophysin and neuron-specific enolase. The present case reports a woman with a history of desmoplastic malignant melanoma exhibiting a BRAF mutation, which later transformed into a GNET of the small intestine with both a BRAF mutation and two subtypes of EWSR1-ATF1 fusion genes. Tumor cells were revealed to be weakly immunoreactive or negative for S-100 and SOX10, lacked markers of melanocytic differentiation and were focally positive for CD56. Combination therapy with dabrafenib mesylate and trametinib dimethyl sulfoxide proved to be temporarily effective against this tumor. The present case is relatively unique as, to the best of our knowledge, there is no case of GNET with a history of melanoma. Furthermore, there is no report of GNET exhibiting both a BRAF mutation and an EWSR1-ATF1 fusion gene. Further accumulation of similar cases is necessary to elucidate the pathological significance of this GNET having a BRAF mutation.

Simultaneous quantification of dabrafenib, hydroxy-dabrafenib and trametinib in human plasma by liquid chromatography-tandem mass spectrometry

A new liquid chromatography-tandem mass spectrometry (LC–MS/MS) method for the simultaneous quantification of dabrafenib (DAB), its main metabolite hydroxy-dabrafenib (OHD) and trametinib (TRA) in human plasma has been developed and validated. After addition of internal standard (dabrafenib-d9), extraction was achieved after protein precipitation with acetonitrile containing 1 % (v/v) formic acid. Chromatographic separation was performed on an Accucore® C18 (2.1 × 50 mm; 2.6 μm) column using a gradient elution of water acidified with 0.1 % (v/v) formic acid (A) and acetonitrile containing 0.1 % (v/v) formic acid (B) at a flow rate of 500 μL/min. The calibration ranged from 10 to 2000 ng/mL for DAB and OHD and from 5 to 50 ng/mL for TRA. This method was validated with satisfactory results including good precision (intra- and inter-assay coefficient of variation from 2.0 %–14.9 %) and good accuracy (inter- and intra-day bias between -1.2 % and 10.9 %), as well as long term stability in unprocessed plasma at −20 °C. This newly proposed method is useful for clinical research purposes as well as therapeutic drug monitoring for patients with a Rapidly Accelerated Fibrosarcoma kinase B (BRAF)-mutated cancer.

Asian Case of Metastatic Melanoma in Which a Complete Response Was Maintained after Discontinuation of Dabrafenib and Trametinib

A 54-year-old man diagnosed with metastatic melanoma of the right inguinal node with occult primary developed liver and bone metastases. The combination of dabrafenib plus trametinib was initiated, and a complete response (CR) was achieved 24 months after starting treatment. One month later, the target therapy was discontinued at the patient’s decision, and he has remained free from progression for 21 months since discontinuation. To the extent of our knowledge, real-world data in Asian melanoma concerning the discontinuation of dabrafenib plus trametinib after achieving CR have not been published; therefore, our case is a meaningful one for considering to cease target drugs and to rescue their financial toxicity.

Functionalized Graphene Oxide Thin Films for Anti-tumor Drug Delivery to Melanoma Cells.

Since Graphene discovery, their associated derivate nanomaterials, Graphene Oxide (GO) and reduced-GO were in the forefront of continuous developments in bio-nano-technology due to unique physical-chemical properties. Although GO nano-colloids (GON) were proposed as drug release matrix for targeting cancer cells, there is still a concern regarding its cytotoxicity issues. In this study, we report on the fabrication of functional GON bio-coatings by Matrix-Assisted Pulsed Laser Evaporation (MAPLE) to be used as drug carriers for targeting melanoma cells. We first performed a thorough in vitro cytotoxicity assay for comparison between GON and protein functionalized GON coatings. As functionalization protein, Bovine Serum Albumin (BSA) was non-covalently conjugated to GO surface. Safe concentration windows were identified in cytotoxicity tests by live/dead staining and MTS assays for five different human melanoma cell lines as well as for non-transformed melanocytes and human dermal fibroblasts. Hybrid GON-BSA nano-scaled thin coatings incorporating Dabrafenib (DAB) and Trichostatin A (TSA) inhibitors for cells bearing BRAFV600E pathway activating mutation were assembled on solid substrates by MAPLE technique. We further demonstrated the successful immobilization for each drug-containing GON-BSA assembling systems by evaluating cellular BRAF activity inhibition and histone deacetylases activity blocking, respectively. DAB activity was proven by the decreased ERK phosphorylation in primary melanoma cells (SKmel28 BRAFV600E cell line), while TSA effect was evidenced by acetylated histones accumulation in cell's nuclei (SKmel23 BRAF WT cell line). In addition, melanoma cells exposed to GON-BSA coatings with compositional gradient of inhibitors evidenced a dose-dependent effect on target activity. Such functional bio-platforms could present high potential for cell-biomaterial interface engineering to be applied in personalized cancer therapy studies.

Improving the Dissolution Rate of the Anticancer Drug Dabrafenib

Dabrafenib (DBF) is an anticancer drug with selective B-Raf (BRAF) inhibition activity. In 2013, DBF was approved by the U.S. Food and Drug Administration (US-FDA) as a dabrafenib mesylate salt (DB...

Protective effect of dabrafenib on renal ischemia-reperfusion injury in vivo and in vitro

AimsIschemia-reperfusion injury (IRI) is a major cause of acute kidney injury (AKI), which can lead to poor outcome and increased risk of mortality. Dabrafenib (DAB) is an approved cancer treatment. Little is known about the effect of DAB in prevention or treatment of renal IRI. MethodsFor in vivo experiments, C57BL/6 mice were divided into four groups: sham (no IRI, no DAB), IRI, DAB, and DAB + IRI. IRI was induced by clamping of bilateral renal pedicles for 30 min. For in vitro experiments, HK-2 cells were used to establish the hypoxia/reoxygenation (H/R) injury model, with four groups: control (no H/R, no DAB), H/R, DAB, and DAB + H/R. Renal function and renal histological changes were recorded. Expression of NGAL and KIM-1 proteins and mRNAs were determined by western blotting and qRT-PCR; secretion of inflammatory cytokines (IL-6 and TNF- α) was determined by qRT-PCR; Cell death was determined using the TUNEL assay, measurement of cleaved caspase-3, and flow cytometry. Necroptosis-related proteins were determined by western blotting. ResultsIn mice, DAB pretreatment improved renal function and also reduced histological injury, inflammation, cell death, and expression of necroptosis-associated proteins. In HK-2 cells, DAB significantly decreased the levels of NGAL and KIM-1, inflammatory cytokines, cell death, and necroptosis-related proteins. ConclusionOur in vitro and in vivo experiments indicated that DAB appears to alleviate renal IRI by suppressing cell death and inhibiting inflammatory responses. DAB has potential use for the clinical prevention and treatment of AKI-induced IRI.

Dabrafenib and trametinib in patients with tumors with BRAF V600E/K mutations: Results from the molecular analysis for therapy choice (MATCH) Arm H.

3002 Background: The NCI-MATCH precision medicine trial assigns patients (pts) with solid tumors, lymphomas, or multiple myeloma with progression on prior treatment to a targeted therapy based on genetic alterations identified in pre-treatment biopsies. Arm H (EAY131-H) evaluated the combination of the BRAF inhibitor (inh) dabrafenib (DAB), and the MEK inh, trametinib (TRM), in pts with BRAF V600E/K mutations. Methods: Pts with melanoma, thyroid, or colorectal cancer were excluded. Pts with NSCLC were excluded after the U.S. Food and Drug Administration (FDA) approved DAB/TRM for this indication. Pts received DAB 150 mg po BID and TRM 2 mg PO daily on 28 day cycles until disease progression or intolerable toxicity; restaging was performed every 2 cycles. The primary endpoint was objective response rate (ORR); secondary endpoints included progression-free survival (PFS), 6-month PFS, and overall survival (OS). Results: A total of 35 pts were enrolled from 1/2016-2/2018; 2 were ineligible (CrCl below criteria; labs out of window). Over 17 distinct tumor histologies were represented. 58% of pts were female, median age was 63 (range 21-85), 94% were Caucasian, and 48% of pts had received at least 3 prior therapies (range 1- 8). The confirmed ORR was 33.3% (90% CI 19.9%, 49.1%), with a median duration of response (DoR) of 12 months (mon). Varied histologies had a DoR of &gt; 12 mon: histiocytic sarcoma, cholangiocarcinoma and mixed adenoneuroendocrine carcinoma of unknown primary, among others. Median PFS was 9.4 mon; the 6 mon PFS rate was 70.6% (90% CI 58.2%-85.5%), and an additional 10 pts had a PFS &gt; 5.5 mon. Median OS has not been reached. At the time of data cutoff (12/2018) 11 pts continue on treatment. Adverse events (AE) were comparable to previously reported profiles of DAB/TRM; no new AEs were identified. The most frequent grade 3 AEs were fatigue, neutropenia, hyponatremia, hypophosphatemia, and urinary tract infection; there was 1 grade 4 sepsis; no grade 5 AEs. Conclusions: In this pre-treated, mixed histology cohort, DAB and TRM showed promising activity outside of currently approved FDA indications warranting further investigations. Correlative analyses are planned. Clinical trial information: NCT02465060.

Interaction of Anticancer Drugs with Human Organic Anion Transporter hOAT4.

Human organic anion transporter 4 (hOAT4) belongs to a family of multispecific organic anion transporters that play critical roles in the disposition of numerous drugs and therefore are the major sites for drug-drug interaction. Drug-drug interactions contribute significantly to the individual variation in drug response. hOAT4 is expressed in the kidney and placenta. In the current study, we examined the interaction of 36 anticancer drugs with hOAT4 in kidney COS-7 cells and placenta BeWo cells. Among the drugs tested, only epirubicin hydrochloride and dabrafenib mesylate exhibited > 50% cis-inhibitory effect, in COS-7 cells, on hOAT4-mediated uptake of estrone sulfate, a prototypical substrate for the transporter. The IC50 values for epirubicin hydrochloride and dabrafenib mesylate were 5.24±0.95 μM and 8.30±3.30 μM, respectively. Dixon plot analysis revealed that inhibition by epirubicin hydrochloride was noncompetitive with a Ki = 3 μM whereas inhibition by dabrafenib mesylate was competitive with a Ki = 4.26 μM. Our results established that epirubicin hydrochloride and dabrafenib mesylate are inhibitors of hOAT4. Furthermore, by comparing our data with clinically relevant exposures of these drugs, we conclude that although the tendency for dabrafenib mesylate to cause drug-drug interaction through hOAT4 is insignificant in the kidney, the propensity for epirubicin hydrochloride to cause drug-drug interaction is high.

BRAF inhibitors stimulate inflammasome activation and interleukin 1 beta production in dendritic cells.

Melanoma is the most dangerous form of skin cancer with a growing incidence over the last decades. Fourty percent of all melanomas harbor a mutation in the signaling adaptor BRAF (V600E) that results in ERK hyperactivity as an oncogenic driver. In these cases, treatment with the BRAFV600E inhibitors Vemurafenib (VEM) or Dabrafenib (DAB) coapplied with the MEK1/2 inhibitors Cobimetinib (COB) or Trametinib (TRA) can result in long-term suppression of tumor growth. Besides direct suppression of ERK activity, these inhibitors have been reported to also modulate tumor immune responses, and exert pro-inflammatory side effects such as fever and rash in some patients. Here we asked for potential effects of BRAFV600E inhibitors on dendritic cells (DC) which are essential for the induction of adaptive anti-tumor responses. Both splenic and bone marrow-derived (BM) mouse dendritic cells (DC) up-regulated costimulator expression (CD80, CD86) in response to DAB but not VEM treatment. Moreover, DAB and to lesser extent VEM enhanced IL-1β (interleukin 1 beta) release by splenic DC, and by LPS-stimulated BMDC. We demonstrate that DAB and VEM activated the NLRC4/Caspase-1 inflammasome. At high concentration, DAB also induced inflammasome activation independent of Caspase-1. TRA and COB elevated MHCII expression on BMDC, and modulated the LPS-induced cytokine pattern. Immunomodulatory activity of DAB and VEM was also observed in human monocyte-derived DC, and DAB induced IL-1β in human primary DC. Altogether, our study shows that BRAFV600E inhibitors upregulate IL-1β release by mouse and human DC which may affect the DC-mediated course of anti-tumor immune responses.