Selective BRAF Inhibitors Research Articles

Abstract 1186: Propranolol could overcome BRAF inhibitors resistance by multiple mechanisms in melanoma

Abstract BRAF(V600E) is the most common oncogenic mutation in melanoma and leads to constitutive activation of the MAPK pathway, which results in uncontrolled cell growth. Selective BRAF inhibitors such as vemurafenib have been observed to neutralize oncogenic the signaling, inhibit cellular growth, and improve patient outcome. Although these mechanisms of vemurafenib resistance have been reported, few studies focused on how to overcome the resistance. Propranolol, a non-selective β-blocker, was confirmed to involve in multiple anticancer effects. Our previous study also showed propranolol inhibited melanoma by suppressing MAPK and AKT pathways in vitro an in vivo. But its efficacy and mechanism of overcoming vemurafenib still remain unknown in melanoma. Here, we explored the effect of propranolol on the A375, P-8 (patient-derived melanoma cell line) vemurafenib resistance cell line and resistance mice xenografts. Cell viability assay demonstrated that 2μM -20μM vemurafenib couldn’t decrease the proliferation but 24h-120h of incubation of 2μM-200μM propranolol inhibited viability with a concentration and time dependent manner in the two resistance cell line. TUNEL staining showed 24h incubation of 20μM propranolol alone or plus 4μM vemurafeinb obviously increased cell apoptosis. Mice received daily ig. administration of propranolol at the dose of 2 mg/kg alone or plus 10 mg/kg for 21days. The mean tumor volume at day 21 in resistance A375 xenografts was 221.13 ± 7.65mm3vs. 904.12 ± 70.57mm3 vs. 2021 ± 316.24mm3for the propranolol plus vemurafenib, propranolol alone, vemurafenib alone, respectively. Propranolol improved mice survival, 28.6% animal dead in plus group, 57.1% mice dead in the propranolol group, and 71.5% animal dead in vemurafenib at end of treatment. IHC showed propranolol also reduced Ki67 index both in propranolol and plus group when compared with vemurafenib treated mice. Furthermore, RNA sequencing was performed to explore the mechanism of propranolol overcoming the resistance, the data showed propranolol largely reduced mRNA levels of IGF family (IGFBP3, IGFLR1, IGFBP6, IGF2, IGF1R.etc.) but elevated the expressions of innate immune related genes (NKG7, TLR9, NCR3.etc.) in A375-vemr cell line. These results provide a strategy of therapeutic resistance for the clinic, importantly, this study also provide a clue targeting IGF family and regulating innate immune might be a potential strategy to suppress resistance in BRAF inhibitor therapies in melanoma. Citation Format: Chengfang Zhou, DongYa Shen, WeiLi Wang, Shangchen Xie, Ping Liao, Xiang Chen, Howard L McLeod, Yijing He. Propranolol could overcome BRAF inhibitors resistance by multiple mechanisms in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1186. doi:10.1158/1538-7445.AM2017-1186

Abstract 158: ASN003, a highly selective inhibitor of B-Raf and PI3 kinases, shows strong antitumor activity in B-Raf inhibitor resistant patient-derived xenograft models

Abstract The RAS-RAF-MEK and PI3K-AKT-mTOR pathways are two major signaling pathways involved in human cancer. Components of these two pathways are frequently mutated in a wide variety of solid tumors. Concurrent double mutations in the two pathways are also observed quite often in a broad range of tumor types. Additionally, inhibition of one of these pathways often leads to the upregulation of the other pathway and development of resistance. In preclinical models, combined inhibition of both pathways has been shown to impart greater efficacy as compared to inhibition of either pathway alone. ASN003 is a novel, highly selective, small-molecule inhibitor of both RAS-RAF and PI3K pathways, discovered using a rational design approach. ASN003 shows potent inhibitory activity against B-RAF and PI3K kinases (low nM IC50). Within the PI3K family, ASN003 has high selectivity for inhibition of PI3Kα and PI3Kδ over PI3Kβ. In a panel of 292 kinases, ASN003 showed high selectivity for inhibiting B-RAF and PI3 kinases, and associated mutant kinases. In cell-based mechanistic studies, ASN003 inhibited phosphorylation of ERK, AKT and S6, and showed strong antiproliferative activity (IC50 = 60-300 nM) in cell lines with B-RAF and PI3K pathway mutations as well as in vemurafenib-resistant cell lines. In pharmacodynamic studies in multiple tumor models, ASN003 showed strong inhibition of the phosphorylation of downstream targets of B-RAF and PI3K, confirming appropriate target engagement. In in vivo efficacy studies, ASN003 showed strong tumor growth inhibition or regression in multiple tumor xenograft models, including A375 (B-Raf V600E mutation), RKO (B-Raf V600E and PIK3CA mutations), and A2058 (B-Raf V600E mutation and PTEN loss). We now report that ASN003 also showed strong tumor growth inhibition (>80%) in a patient-derived xenograft (PDX) model established from a relapsed patient with progressive B-Raf mutant melanoma who showed initial response to vemurafenib. Sequencing analysis showed that the vemurafenib resistant tumor acquired a concurrent PIK3CA mutation. Dual targeting of the B-RAF and PI3K pathways with ASN003 has the potential to treat and/or prevent the acquired resistance to selective B-RAF inhibitors, and may also treat a broader patient population and provide greater efficacy and survival benefit than selective B-RAF inhibitors or selective PI3K pathway inhibitors alone. ASN003 is currently in Phase I clinical development in patients with advanced solid tumors, including tumors with B-Raf V600 mutation, PI3 kinase pathway alterations or PTEN loss. Citation Format: Scott K. Thompson, Roger A. Smith, Niranjan Rao, Michael J. Wick, Sanjeeva P. Reddy. ASN003, a highly selective inhibitor of B-Raf and PI3 kinases, shows strong antitumor activity in B-Raf inhibitor resistant patient-derived xenograft models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 158. doi:10.1158/1538-7445.AM2017-158

A combinatorial strategy using YAP and pan-RAF inhibitors for treating KRAS-mutant pancreatic cancer

KRAS mutation is the most common genetic event in pancreatic cancer. Whereas KRAS itself has proven difficult to inhibit, agents that target key downstream signals of KRAS, such as RAF, are possibly effective for pancreatic cancer treatment. Because selective BRAF inhibitors paradoxically induce downstream signaling activation, a pan-RAF inhibitor, LY3009120 is a better alternate for KRAS-mutant tumor treatment. Here we explored a new combinational strategy using a YAP inhibitor and LY3009120 in pancreatic cancer treatment. We found that reduced YAP expression closely correlates with longer relapse-free and overall survival of patients. Stable knockdown of YAP significantly inhibited pancreatic cancer cell proliferation and tumor growth. In addition, LY3009120 exhibited a dramatically enhanced antitumor effect in combination with YAP knockdown. YAP depletion blocks the activation of a parallel AKT signal pathway after LY3009120 treatment. Finally, combination with a YAP inhibitor, verteporfin, significantly enhanced the antitumor efficacy of LY3009120. Collectively, our results demonstrate that genetic or pharmacological inhibition of YAP can increase sensitivity to LY3009120 in pancreatic cancer through blocking compensatory activation of a parallel AKT signal pathway, thereby validating a combinatorial approach for treating KRAS-mutant pancreatic cancer.

ASN003, a highly selective BRAF and PI3K inhibitor: Preclinical and phase 1 clinical data in patients with advanced solid tumors.

e14102 Background: RAS-RAF-MEK and PI3K-AKT-mTOR are two major signaling pathways involved in tumorigenesis. Components of these two pathways are frequently mutated in a broad range of tumors. ASN003 is a novel and highly selective small-molecule inhibitor of the RAS-RAF-MEK and PI3K pathways. Methods: The activity of ASN003 was determined using PI3K and BRAF enzymes, and efficacy was studied in human tumor xenograft models in mice. ASN003 is currently being investigated in patients with solid tumors in a Phase 1 trial using an accelerated dose titration design. In Part A, safety and tolerability of ASN003 is being studied in patients with advanced solid tumors. In Part B, safety, tolerability and preliminary efficacy of ASN003 will be evaluated in melanoma, CRC and NSCLC patients with a BRAF, PIK3CA or PTEN mutation. Pharmacokinetic (PK) profile and the pharmacodynamic (PD) effects of ASN003 on biomarkers such as pERK and pS6 are investigated in both parts of the study. Results: ASN003 showed potent and highly selective inhibition of BRAF and PI3K-α and -δ, and low affinity for PI3K-ß. ASN003 showed strong antiproliferative activity in cell lines and caused significant tumor growth inhibition in xenograft models harboring BRAF and PIK3CA or PTEN mutations. ASN003 showed antiproliferative activity in B-RAF and MEK inhibitor resistant cell lines. ASN003 had a strong antitumor activity in a BRAFV600mutant melanoma PDX model resistant to BRAF inhibitors, vemurafenib and dabrafenib. In humans, to date, ASN003 was well tolerated at 10 and 20 mg QD. Adverse events were mild and peak plasma level of 120 nM at 10 mg QD was achieved with a half-life of > 12 h. Dose escalation is ongoing. Conclusions: ASN003 is a unique small molecule, with highly selective and potent inhibition of BRAF, PI3-α and -δ kinases. ASN003 has strong antitumor activity in various xenograft tumor models harboring both BRAF and PIK3CA/PTEN mutations, and in a BRAF inhibitor resistant melanoma PDX model. To date, ASN003 was well tolerated and achieved good systemic exposure. Updated and detailed clinical, PK and PD results will be presented. Clinical trial information: NCT02961283.

BRAF-V600 mutational status affects recurrence patterns of melanoma brain metastasis.

Brain metastasis is common and carries a poor prognosis in melanoma. A single institution, retrospective cohort of 225 melanoma patients was analyzed to determine if BRAF-V600 mutational status was associated with brain metastasis. Eighty-three of the 225 patients (37%) had BRAF-V600 mutations. At initial diagnosis, BRAF-V600 mutations were associated with younger age (p ≤ 0.001), higher proportion of females (p = 0.0037), higher AJCC stage (p = 0.030), regional lymph node involvement (p = 0.047), and family history of cancer (p = 0.044). Compared to BRAF-WT, BRAF-V600 patients had an increased risk of brain metastasis in multivariate analysis (OR = 2.24; 95% CL = 1.10-4.58; p = 0.027). However, BRAF-V600 patients treated with a selective BRAF inhibitor (BRAFi) had a similar risk of brain metastasis compared to BRAF-WT patients (OR = 1.00; 95% CL = 0.37-2.65; p = 0.98). Moreover, treatment with BRAFi significantly prolonged the time from initial diagnosis to brain metastasis diagnosis (HR = 0.30; 95% CL = 0.11-0.79; p = 0.015). Compared to other tissues, the brain was the most frequent site of metastasis in BRAF-V600 patients without BRAFi (42% ± 7%). The frequency of brain metastasis was lower in BRAF-WT and BRAF-V600 patients with BRAFi (25% ± 4% and 25% ± 8%, respectively). The proportion of patients with brain metastasis as the only site was 40%, 60%, and 0% in the BRAF-WT, BRAF-V600 without BRAFi, and BRAF-V600 with BRAFi groups, respectively. This study provides evidence on the clinical importance of BRAF-V600 mutations and BRAF inhibition in the progression to melanoma brain metastasis.

Clinical trials of targeted therapy in the treatment of RAI failure or advanced DTC

Regarding radioactive iodine-refractory failure or advanced differentiated thyroid cancer, multiple multikinase inhibitors including sorafenib and lenvatinib, which target platelet derived growth factor receptor, vascular endothelial growth factor pathway, and rearranged during transfection (RET) pathway were proved to have obvious antitumor activity. Moreover, selective BRAF inhibitor, promoting drug uptake of radioactive iodine also showed a certain therapeutic effect. These molecular targets which are relevant with differentiated thyroid cancer occurrence, development, invasion and metastasis have become of its moment, and, selective inhibitors and re-differentiation agents were shown to be promising. In the future, individual genetic testing would provide more specific information in directing individualized molecular-targeted therapy. Key words: Thyroid neoplasms/TH; Review

Erratum to: Ongoing Response in BRAF V600E-Mutant Melanoma After Cessation of Intermittent Vemurafenib Therapy: A Case Report.

The selective BRAF inhibitors vemurafenib and dabrafenib yield high response rates and improved overall survival in patients with BRAF V600E-mutant metastatic melanoma. Treatment traditionally continues until disease progression or the development of unacceptable toxicity. Acquired drug resistance and toxicity are key challenges with the use of these drugs. Resistance to vemurafenib usually develops within 6-8 months. Management of drug toxicity typically involves stopping vemurafenib until resolution, before restarting at a lower dose, or permanently ceasing vemurafenib therapy. We have recently considered whether intermittent dosing could be used as an alternative to dose reduction/termination in the management of vemurafenib toxicity. One patient treated with intermittent vemurafenib was an 89-year-old woman with metastatic melanoma, who initially showed a good response to continuous dosing. Recurrent toxicity meant that the continuous vemurafenib dosage was repeatedly ceased before restarting at a lower dose. Ten months after vemurafenib was first begun, an intermittent dosing regimen was introduced in an attempt to control toxicity. This continued for 2 months, before cessation due to continued unacceptable toxicity. A further 24 months later, the patient remains fit and well in complete clinical remission, with no recurrence of her previous melanoma and no new primary malignancies. To the best of our knowledge, a continued response after the cessation of selective BRAF inhibitors has never before been described in melanoma. Induction of an immune response and/or epigenetic changes could explain continued disease response after cessation of vemurafenib therapy. Care should be taken when extrapolating the findings from the continued response after vemurafenib cessation to other tumour types. We recommend the collection and analysis of data to investigate the clinical responses seen after cessation of vemurafenib due to intolerable toxicities, which could help further explain vemurafenib's mechanism of action.

CpG-based immunotherapy impairs antitumor activity of BRAF inhibitors in a B-cell-dependent manner

Combining immunotherapy with targeted therapy has increasingly become an appealing therapeutic paradigm for cancer treatment due to its great potential for generating durable and synergistic antitumor response. In this study, however, we unexpectedly found that two types of CpG-based tumor peptide vaccine treatments consistently negated the antitumor activity of a selective BRAF inhibitor in tumors with BRAF mutation rather than showing a synergistic antitumor effect. Our further studies demonstrated that CpG alone was sufficient to dampen BRAF inhibitor-induced antitumor responses, suggesting that the impaired antitumor activity of the BRAF inhibitor observed in mice receiving CpG-based peptide vaccine is mainly dependent upon the use of CpG. Mechanistically, CpG increased the number of circulating B cells, which produced elevated amounts of tumor necrosis factor-α (TNFα) that contributed to the increased tumor resistance to BRAF inhibitors. More importantly, B-cell depletion or TNFα neutralization can restore the antitumor effect of BRAF inhibition in mice receiving CpG treatment, indicating that TNFα-secreting B cells play an indispensable role in BRAF inhibitor resistance induced by CpG. Taken together, our results strongly suggest that precautions must be implemented when designing combinatorial approaches for cancer treatment, because distinct regimens, despite their respective therapeutic benefit as monotherapy, may together provide antagonistic clinical outcomes.

Critical role of reactive oxygen species (ROS) for synergistic enhancement of apoptosis by vemurafenib and the potassium channel inhibitor TRAM-34 in melanoma cells

Inhibition of MAP kinase pathways by selective BRAF inhibitors, such as vemurafenib and dabrafenib, have evolved as key therapies of BRAF-mutated melanoma. However, tumor relapse and therapy resistance have remained as major problems, which may be addressed by combination with other pathway inhibitors. Here we identified the potassium channel inhibitor TRAM-34 as highly effective in combination with vemurafenib. Thus apoptosis was significantly enhanced and cell viability was decreased. The combination vemurafenib/TRAM-34 was also effective in vemurafenib-resistant cells, suggesting that acquired resistance may be overcome. Vemurafenib decreased ERK phosphorylation, suppressed antiapoptotic Mcl-1 and enhanced proapoptotic Puma and Bim. The combination resulted in enhancement of proapoptotic pathways as caspase-3 and loss of mitochondrial membrane potential. Indicating a special mechanism of vemurafenib-induced apoptosis, we found strong enhancement of intracellular ROS levels already at 1 h of treatment. The critical role of ROS was demonstrated by the antioxidant vitamin E (α-tocopherol), which decreased intracellular ROS as well as apoptosis. Also caspase activation and loss of mitochondrial membrane potential were suppressed, proving ROS as an upstream effect. Thus ROS represents an initial and independent apoptosis pathway in melanoma cells that is of particular importance for vemurafenib and its combination with TRAM-34.

OA08.07 BRAF-V600E Advanced Lung Adenocarcinoma with Leptomeningeal (LM) Disease Treated with Vemurafenib

BRAF mutations occur in around 3% of non-small cell lung cancers (NSCLC) and V600E accounts for 50%. BRAF V600E is an attractive molecular target for cancer tyrosine kinase treatment, but ideal treatment is still not defined. A case of a patient with BRAF-mutated non–small cell lung cancer (NSCLC) detected by NGS Ion torrent technology who presented with LM disease and was treated with the selective BRAF inhibitor vemurafenib is described. 58 years old, female, non-smoker, who presented in the emergency department with pericardial effusion. Pericardial fluid cytology confirmed adenocarcinoma TTF1 positive. Multi-organ metastatic disease was diagnosed (bone, lung and thyroid) without EGFR mutation or ALK-EML4 translocation. Four cycles of chemotherapy with pemetrexed and carboplatin were done. She started with refractory headache and vomiting, brain CT and MRI showed no evidence of metastasis, a lumbar puncture confirmed malignant cells in the cerebrospinal fluid. A BRAF V600E was detected by NGS, Ion Torrence PGM technology in the initial tumor sample and in plasma circulating free DNA. An off-label treatment with vemurafenib 960 mg q12hr was offered to the patient, with clinical improvement and radiologic lung stability. At month 2 of treatment, the patient developed respiratory insufficiency with lung infiltrates and Influenza A virus was identified in a nasal swab. Vemurafenib was temporary suspended and re-introduced until 720 mg q12h and maintained until disease progression (large volume pleural effusion with positive cytology), at month 6 of vemurafenib tretament. Third line treatment is being planned. The authors highlight the importance of using a multiplex screening strategy to detect targetable mutations in advanced lung cancer patients. The application of next generation sequencing to the tumor and plasma cfDNA allowed the detection of a BRAF-V600E mutation. The improvement of neurologic symptoms and disease control achieved with vemurafenib supports vemurafenib's efficacy. Care should be taken to the possibility of occurring lung toxicity.

ACY-1215 accelerates vemurafenib induced cell death of BRAF-mutant melanoma cells via induction of ER stress and inhibition of ERK activation.

BRAFV600E mutation is found in ~50% of melanoma patients and BRAFV600E kinase activity inhibitor, vemurafenib, has achieved a remarkable clinical response rate. However, most patients treated with vemurafenib eventually develop resistance. Overcoming primary and secondary resistance to selective BRAF inhibitors remains one of the most critically compelling challenges for these patients. HDAC6 has been shown to confer resistance to chemotherapy in several types of cancer. Few studies focused on the role of HDAC6 in vemurafenib resistance. Here we showed that overexpression of HDAC6 confers resistance to vemurafenib in BRAF-mutant A375 cells. ACY-1215, a selective HDAC6 inhibitor, inhibits the proliferation and induces the apoptosis of A375 cells. Moreover, ACY-1215 sensitizes A375 cells to vemurafenib induced cell proliferation inhibition and apoptosis induction, which occur partly through induction of endoplasmic reticulum (ER) stress and inactivation of extracellular signal-regulated kinase (ERK). Taken together, our results suggest that the inhibition of HDAC6 may be a promising strategy for the treatment of melanoma and overcoming resistance to vemurafenib.

LY3009120, a panRAF inhibitor, has significant anti-tumor activity in BRAF and KRAS mutant preclinical models of colorectal cancer.

Activating mutations in the KRAS and BRAF genes, leading to hyperactivation of the RAS/RAF/MAPK oncogenic signaling cascade, are common in patients with colorectal cancer (CRC). While selective BRAF inhibitors are efficacious in BRAFmut melanoma, they have limited efficacy in BRAFmut CRC patients. In a RASmut background, selective BRAF inhibitors are contraindicated due to paradoxical activation of the MAPK pathway through potentiation of CRAF kinase activity. A way to overcome such paradoxical activation is through concurrent inhibition of the kinase activity of both RAF isoforms. Here, we further examined the effects of LY3009120, a panRAF and RAF dimer inhibitor, in human models of CRC with various mutational backgrounds. We demonstrate that LY3009120 induced anti-proliferative effects in BRAFmut and KRASmut CRC cell lines through G1-cell cycle arrest. The anti-proliferative effects of LY3009120 in KRASmut CRC cell lines phenocopied molecular inhibition of RAF isoforms by simultaneous siRNA-mediated knockdown of ARAF, BRAF and CRAF. Additionally, LY3009120 displayed significant activity in in vivo BRAFmut and KRASmut CRC xenograft models. Examination of potential resistance to LY3009120 demonstrated RAF-independent ERK and AKT activation in the KRASmut CRC cell line HCT 116. These findings describe the preclinical activity of a panRAF inhibitor in a BRAFmut and KRASmut CRC setting.

Multimodal tumor suppression by miR-302 cluster in melanoma and colon cancer

The miR-302 family is one of the main groups of microRNAs, which are highly expressed in embryonic stem cells (ESCs). Previous reports have indicated that miR-302 can reduce the proliferation rate of some cancer cells while compromising on their oncogenic potential at the same time without having the same effect on normal somatic cells. In this study we aimed to further investigate the role of the miR-302 cluster in multiple cancer signaling pathways using A-375 melanoma and HT-29 colorectal cancer cells. Our results indicate that the miR-302 cluster has the potential to modulate oncogenic properties of cancer cells through inhibition of proliferation, angiogenesis and invasion, and through reversal of the epithelial-to-mesenchymal transition (EMT) in these cells. We showed for the first time that overexpression of miR-302 cluster sensitized A-375 and HT-29 cells to hypoxia and also to the selective BRAF inhibitor vemurafenib. MiR-302 is a pleiotropically acting miRNA family which may have significant implications in controlling cancer progression and invasion. It acts through a reprogramming process, which has a global effect on a multitude of cellular pathways and events. We propose that reprogramming of cancer cells by epigenetic factors, especially miRNAs might provide an efficient tool for controlling cancer and especially for those with more invasive nature.

Preclinical efficacy of a RAF inhibitor that evades paradoxical MAPK pathway activation in protein kinase BRAF-mutant lung cancer

Oncogenic activation of protein kinase BRAF drives tumor growth by promoting mitogen-activated protein kinase (MAPK) pathway signaling. Because oncogenic mutations in BRAF occur in ∼2-7% of lung adenocarcinoma (LA), BRAF-mutant LA is the most frequent cause of BRAF-mutant cancer mortality worldwide. Whereas most tumor types harbor predominantly the BRAFV600E-mutant allele, the spectrum of BRAF mutations in LA includes BRAFV600E (∼60% of cases) and non-V600E mutant alleles (∼40% of cases) such as BRAFG469A and BRAFG466V The presence of BRAFV600E in LA has prompted clinical trials testing selective BRAF inhibitors such as vemurafenib in BRAFV600E-mutant patients. Despite promising clinical efficacy, both innate and acquired resistance often result from reactivation of MAPK pathway signaling, thus limiting durable responses to the current BRAF inhibitors. Further, the optimal therapeutic strategy to block non-V600E BRAF-mutant LA remains unclear. Here, we report the efficacy of the Raf proto-oncogene serine/threonine protein kinase (RAF) inhibitor, PLX8394, that evades MAPK pathway reactivation in BRAF-mutant LA models. We show that PLX8394 treatment is effective in both BRAFV600E and certain non-V600 LA models, in vitro and in vivo. PLX8394 was effective against treatment-naive BRAF-mutant LAs and those with acquired vemurafenib resistance caused by an alternatively spliced, truncated BRAFV600E that promotes vemurafenib-insensitive MAPK pathway signaling. We further show that acquired PLX8394 resistance occurs via EGFR-mediated RAS-mTOR signaling and is prevented by upfront combination therapy with PLX8394 and either an EGFR or mTOR inhibitor. Our study provides a biological rationale and potential polytherapy strategy to aid the deployment of PLX8394 in lung cancer patients.

Multiple Squamous Cell Carcinoma after Vemurafenib Therapy

Vemurafenib is a low molecular weight, orally available, inhibitor of oncogenic V600 BRAF serine–threonine kinase. It is effective in the treatment of v-raf murine sarcoma viral oncogene homolog B1 (BRAF) V600-located melanomas.1 The drug specifically targets tumor cells that harbor activating mutations in BRAF gene, most commonly the substitution of glutamic acid for valine at codon 600 (V600E). Selective BRAF inhibition therapy is associated with malignant and benign growths. Among these are keratoacanthomas, cutaneous squamous cell carcinomas, warty dyskeratomas, verrucous keratoses, acantholytic dyskeratoses, and widespread eruptions. Recent studies showed that squamous cell carcinoma (SCC) and keratoacanthomas developed in 20%–57% of patients who are on vemurafenib therapy.2,3 Cutaneous malignancies occur more frequently than other kinase inhibitor molecules after vemurafenib therapy

Non-Invasive Glutamine PET Reflects Pharmacological Inhibition of BRAFV600E In Vivo.

This study aimed to study whether cancer cells possess distinguishing metabolic features compared with surrounding normal cells, such as increased glutamine uptake. Given this, quantitative measures of glutamine uptake may reflect critical processes in oncology. Approximately, 10% of patients with colorectal cancer (CRC) express BRAF V600E , which may be actionable with selective BRAF inhibitors or in combination with inhibitors of complementary signaling axes. Non-invasive and quantitative predictive measures of response to these targeted therapies remain poorly developed in this setting. The primary objective of this study was to explore 4-[18F]fluoroglutamine (4-[18F]F-GLN) positron emission tomography (PET) to predict response to BRAFV600E-targeted therapy in preclinical models of colon cancer. Tumor microarrays from patients with primary human colon cancers (n=115) and CRC liver metastases (n=111) were used to evaluate the prevalence of ASCT2, the primary glutamine transporter in oncology, by immunohistochemistry. Subsequently, 4-[18F]F-GLN PET was evaluated in mouse models of human BRAF V600E -expressing and BRAF wild-type CRC. Approximately 70% of primary colon cancers and 53% of metastases exhibited positive ASCT2 immunoreactivity, suggesting that [18F]4-F-GLN PET could be applicable to a majority of patients with colon cancer. ASCT2 expression was not associated selectively with the expression of mutant BRAF. Decreased 4-[18F]F-GLN predicted pharmacological response to single-agent BRAF and combination BRAF and PI3K/mTOR inhibition in BRAF V600E -mutant Colo-205 tumors. In contrast, a similar decrease was not observed in BRAF wild-type HCT-116 tumors, a setting where BRAFV600E-targeted therapies are ineffective. 4-[18F]F-GLN PET selectively reflected pharmacodynamic response to BRAF inhibition when compared with 2-deoxy-2[18F]fluoro-D-glucose PET, which was decreased non-specifically for all treated cohorts, regardless of downstream pathway inhibition. These findings illustrate the utility of non-invasive PET imaging measures of glutamine uptake to selectively predict response to BRAF-targeted therapy in colon cancer and may suggest further opportunities to inform colon cancer clinical trials using targeted therapies against MAPK activation.

Targeting the MAPK and PI3K pathways in combination with PD1 blockade in melanoma

ABSTRACTImmunotherapy of advanced melanoma with CTLA-4 or PD-1/PD-L1 checkpoint blockade induces in a proportion of patients long durable responses. In contrast, targeting the MAPK-pathway by selective BRAF and MEK inhibitors induces high response rates, but most patients relapse. Combining targeted therapy with immunotherapy is proposed to improve the long-term outcomes of patients. Preclinical data endorsing this hypothesis are accumulating. Inhibition of the PI3K-Akt-mTOR pathway may be a promising treatment option to overcome resistance to MAPK inhibition and for additional combination with immunotherapy.We therefore evaluated to which extent dual targeting of the MAPK and PI3K-Akt-mTOR pathways affects tumor immune infiltrates and whether it synergizes with PD-1 checkpoint blockade in a BRAFV600E/PTEN−/−-driven melanoma mouse model. Short-term dual BRAF + MEK inhibition enhanced tumor immune infiltration and improved tumor control when combined with PD-1 blockade in a CD8+ T cell dependent manner. Additional PI3K inhibition did not impair tumor control or immune cell infiltration and functionality. Analysis of on-treatment samples from melanoma patients treated with BRAF or BRAF + MEK inhibitors indicates that inhibitor-mediated T cell infiltration occurred in all patients early after treatment initiation but was less frequent found in on-treatment biopsies beyond day 15.Our findings provide a rationale for clinical testing of short-term BRAF + MEK inhibition in combination with immune checkpoint blockade, currently implemented at our institutes. Additional PI3K inhibition could be an option for BRAF + MEK inhibitor resistant patients that receive targeted therapy in combination with immune checkpoint blockade.

Prospective Case Series of Cutaneous Adverse Effects Associated With Dabrafenib and Trametinib.

Dabrafenib, a novel selective small-molecule inhibitor of BRAF, has been shown to increase overall survival in patients with unresectable metastatic melanoma harboring the BRAF V600E mutation. The development of resistance has led to combination therapy with selective MEK inhibitor trametinib. Compared with vemurafenib, dabrafenib is a more recent BRAF inhibitor approved by the Food and Drug Administration in May 2013 for metastatic melanoma; fewer data are available in the current literature regarding cutaneous toxicity. We sought to present additional cutaneous side effects of dabrafenib and trametinib and follow their evolution and management. We carried out a prospective study of 14 patients treated with dabrafenib alone or with trametinib. Patients were followed every 4 weeks, and we collected detailed cutaneous symptoms, photos, and biopsy specimens. All patients presented with at least 1 adverse skin reaction. The mean duration of treatment was 24 weeks. The most common adverse effect was papillomas (7/14), followed by palmoplantar hyperkeratosis (5/14), alopecia (5/14), and seborrheic dermatitis-like eruption (2/14). Three patients who received trametinib developed an acneiform eruption (3/5). One patient developed a keratoacanthoma-like squamous cell carcinoma. Side effects presented as early as 2 weeks after starting therapy, with a mean time of onset of 9 weeks. Selective BRAF inhibitor dabrafenib and MEK inhibitor trametinib are associated with multiple skin adverse effects. Given their recent approval and the potential for malignant lesions to develop on treatment, awareness of potential adverse effects and their management is necessary.

MEK inhibitor-associated retinopathy (MEKAR) in metastatic melanoma: Long-term ophthalmic effects

BackgroundMitogen-activated protein kinase kinase (MEK) inhibitors have aroused considerable interest in oncology. Activity has been demonstrated in various types of cancer, especially melanoma. MEK inhibitors induce a transient retinopathy, considered to be a class effect. At present, only sparse data are available on retinal effects with long-term MEK inhibition. Patients and methodsIn this prospective, observational study, patients with advanced melanoma participating in different phase 1/2 or phase 3 clinical trials were treated with the MEK inhibitor binimetinib, with a v-Raf murine sarcoma viral oncogene homolog B (BRAF) inhibitor, or with combination therapy. They underwent regular ophthalmological examinations including determination of visual function, biomicroscopy, dilated fundoscopy and optical coherence tomography (OCT) for a period of up to 2 years. Retinopathy was diagnosed on defined OCT criteria. ResultsSixty-two patients were investigated between 1st October 2011 and 31st July 2015: 13 were treated with the MEK inhibitor binimetinib alone, 10 with a selective BRAF inhibitor, and 39 with combination therapy. In 92% of patients on monotherapy and 100% of those on combination treatment, binimetinib caused dose-related lesions with serous neuroretinal detachments and oedema, strongly dependent on the time after medication. With continued treatment, retinal volume and thickness decreased to levels below baseline, without any apparent functional deficits or changes in structural integrity. ConclusionsBinimetinib induces a specific retinopathy with daily fluctuations depending on the time interval after medication. The retinopathy partially recovers, but can still be detected many months later. Retinal thinning, possible first signs of retinal atrophy have been observed after long-term treatment, but, so far, without functional relevance.

Agminated melanocytic nevus status post dabrafenib therapy for metastatic melanoma

Agminated melanocytic nevus is an uncommon type of mole, characterized by a local group of macular or papular pigmented lesions, well demarcated, without a common pigmented background. This pattern has also been associated with Spitz nevi, dysplastic melanocytic nevi, and non-melanocytic lesions.We describe the onset of an acquired agminated melanocytic nevus after dabrafenib treatment. Our case highlights paradoxical MAPK activation in the setting of single-agent BRAF blockade and underscores the importance of characterizing the diverse side effects of selective BRAF inhibitors. This is the first case, to our knowledge, of agminated melanocytic nevus in association with dabrafenib.