Relevance of detection of RAF fusion transcripts in pan-negative melanoma in routine practice.

Pan-negative melanomas account for 30% of melanomas. In case of immunotherapy failure, therapeutic options are limited. Oncogene fusions represent a target of interest in many solid cancers. In melanoma, the frequency of oncogene fusion is not well documented and not routinely investigated. We conducted a single-center retrospective study. The objective was to determine the frequency of oncogene fusion detected by RNA sequencing, in patients with advanced or metastatic pan-negative melanoma. In parallel, an extended molecular alteration search was performed using extended targeted next-generation sequencing. We identified 59 patients with advanced pan-negative melanoma between January 2021 and January 2023. It was a cutaneous melanoma in 71.1% of the cases, a mucous melanoma in 15.2% of the cases. We identified nine patients with a RAF fusion, including seven BRAF gene fusion and two RAF1 fusion. Of the other molecular alterations, NF1 mutation was the most frequent molecular alteration identified. Among the nine patients with RAF fusions, all the patients initially received treatment with anti-PD1 ± anti-CTLA4 immunotherapy. After immunotherapy failure, five patients benefited from second-line targeted therapy (two with BRAF and MEK inhibitors combination, three MEK inhibitors alone). The response rate was 20%. In a population of pan-negative melanoma, we detected 15.2% of RAF fusion. Fusion detection allowed the introduction of a second line of targeted therapy, in the absence of a validated therapeutic option in 55.5% of cases. This study suggests the relevance of detecting RAF fusion in a selected population.

Real-World Treatment Outcomes of MET Exon14 Skipping in Non-small Cell Lung Cancer: GFPC 03-18 Study.

BACKGROUND: MET-targeted tyrosine kinase inhibitors (TKIs) demonstrated efficacy in advanced non-small cell lung cancer (aNSCLC) with MET exon14 skipping mutations (METexon14); yet, data on the management of these patients in clinical practice is sparse. OBJECTIVE: The aim of this study was to describe the management of METexon14 aNSCLC patients. PATIENTS AND METHODS: This real-life, retrospective study analyzed the management of METexon14 aNSCLC. The primary endpoint was the median overall survival (mOS). Secondary endpoints were to assess investigator-progression-free survival (PFS) and mOS in different subgroups: patients treated with (a) crizotinib, regardless of treatment line; (b) anti-MET TKIs (crizotinib, tepotinib, capmatinib); and (c) immunotherapy. RESULTS: A total of 118 patients were included between December 2015 and January 1, 2020 in 13 centers. Median age was 73 years, 62.7% were female, 83.9% had adenocarcinoma, 92.4% at stage IV, and 27% had more than three metastatic sites. The majority of the patients (106, 89.8%) received at least one systemic treatment; 73% received at least one anti-MET TKI: crizotinib (68.6%), tepotinib (16%), capmatinib (10%). Only 10% received two anti-MET TKIs in their treatment sequences. With a median follow-up of 16 months (95% CI 13.6-29.7), mOS was 27.1 months (95% CI 18-31.4). There was no significant difference between mOS of patients treated and never treated with crizotinib, 19.7 (95% CI 13.6-29.7) and 28 (95% CI 16.4-NR) months, respectively (p = 0.16); mOS of the TKI cohort and of the TKI-naïve patient cohort were 27.1 (95% CI 18-29.7) and 35.6 (95% CI 8.6-NR) months respectively, with no significant difference (p = 0.7). CONCLUSIONS: In this real-life study, there was no evidence of benefit in mOS with anti-MET TKIs.

Clinical response under MEK inhibitor alone in metastatic melanoma with a novel fusion involving the RAF1 gene.

Currently, in the absence of BRAFV600 mutation, the management of advanced melanomas is based on immunotherapies, but only half of the patients are responders. RAF1 (also named CRAF) fusions occur in 1-2.1% of wild-type melanomas. Preclinical data suggest that the presence of RAF fusion may be sensitive to MEK inhibitors. We report the case of a patient with an advanced melanoma harboring an EFCC1-RAF1 fusion who showed a clinical benefit from and a partial response to a MEK inhibitor.

MET exon 14 skipping mutation is a hepatocyte growth factor (HGF)-dependent oncogenic driver in vitro and in humanised HGF knock-in mice.

Exon skipping mutations of the MET receptor tyrosine kinase (METex14), increasingly reported in cancers, occur in 3-4% of non-small-cell lung cancer (NSCLC). Only 50% of patients have a beneficial response to treatment with MET-tyrosine kinase inhibitors (TKIs), underlying the need to understand the mechanism of METex14 oncogenicity and sensitivity to TKIs. Whether METex14 is a driver mutation and whether it requires hepatocyte growth factor (HGF) for its oncogenicity in a range of in vitro functions and in vivo has not been fully elucidated from previous preclinical models. Using CRISPR/Cas9, we developed a METex14/WT isogenic model in nontransformed human lung cells and report that the METex14 single alteration was sufficient to drive MET-dependent in vitro anchorage-independent survival and motility and in vivo tumorigenesis, sensitising tumours to MET-TKIs. However, we also show that human HGF (hHGF) is required, as demonstrated in vivo using a humanised HGF knock-in strain of mice and further detected in tumour cells of METex14 NSCLC patient samples. Our results also suggest that METex14 oncogenicity is not a consequence of an escape from degradation in our cell model. Thus, we developed a valuable model for preclinical studies and present results that have potential clinical implication.

Response to Capmatinib in a MET Fusion-positive Cholangiocarcinoma.

Cholangiocarcinoma is the second most common liver cancer after hepatocellular carcinoma. In case of metastatic or unresectable disease, the recommended first-line treatment is gemcitabine-based doublet, most commonly gemcitabine and cisplatin. There is no standard treatment for further lines. MET fusions are rare alterations described in many cancers. The efficacy of specific MET inhibitors is poorly studied. We present the case of a patient with chemotherapy-refractory metastatic cholangiocarcinoma harboring a CAPZA-2-MET fusion along with MET amplification who dramatically responded to capmatinib, a specific MET tyrosine kinase inhibitor.

Safety and efficacy of nivolumab, an anti-PD1 immunotherapy, in patients with advanced basal cell carcinoma, after failure or intolerance to sonic Hedgehog inhibitors: UNICANCER AcSé NIVOLUMAB trial.

BACKGROUND: Basal cell carcinoma (BCC) is the most common human malignancy. In most cases, BCC has slow progression and can be definitively cured by surgery or radiotherapy. However, in rare cases, it can become locally advanced or, even more rarely, metastatic. The alternative recommended treatments are Sonic Hedgehog pathway inhibitors; however, the response is often short-lived. METHODS: This was a phase 2 basket study (NCT03012581) evaluating the efficacy and safety of nivolumab in a cohort of 32 advanced BCC patients, enrolled after failure of Sonic Hedgehog inhibitors, including 29 laBCC (91%) and 3 mBCC (9%). RESULTS: Compared to previously published studies, our population consisted of severe patients with a poor prognosis because they had already received multiple lines of treatment: all patients received previous Sonic Hedgehog inhibitors, 53% of patients already had chemotherapy and 75% radiotherapy. At 12 weeks, we reported 3.1% of complete responses, 18.8% of partial responses, and 43.8% of stable diseases. The best response rate to nivolumab reached 12.5% of complete responses (four patients), 18.8% of partial responses (three patients), and 43.8% of stable diseases (14 patients). Adverse events (AE) were mostly grade 2 or 3, slightly different to the adverse events observed in the treatment of metastatic melanoma (higher rate of diabetes, no thyroid dysfunction). CONCLUSION: Nivolumab is a relevant therapeutic option for patients with advanced relapsing/refractory BCC.

Unexpected cure of a toxic nodule in a multinodular goiter induced by immune checkpoint inhibitors: a case report.

Introduction: Immune checkpoint inhibitors (ICI) are used to treat cancers including metastatic melanomas and can induce endocrine side effects. The thyroid is frequently affected with classically transient thyrotoxicosis followed by hypothyroidism. The evolution of thyroid nodules and goiters under ICI therapy is poorly described. Case presentation: A 72-year-old male presenting with hyperthyroidism due to a toxic nodule in a multinodular goiter (MNG) started ICI therapy combining ipilimumab and nivolumab to treat metastatic melanoma. After an initial worsening of thyrotoxicosis, treated with carbimazole, he developed profound hypothyroidism, persisting after carbimazole discontinuation, needing a long-term levothyroxine supplementation. Ultrasound control performed 6 months after ICIs treatment initiation revealed diffuse thyroid atrophy with involution of all nodules. 123I-scintigraphy confirmed a destructive mechanism. Discussion: The evolution of MNG and toxic nodules is poorly described in patients treated with ICI since systematic US evaluations are lacking. We describe for the first time a toxic nodule cured by ICI therapy inducing destructive thyroiditis. Conclusion: Pre-existing nodules and MNG, even if toxic, are not a contraindication for ICI treatment provided the patients are carefully monitored.

Dramatic response of refractory metastatic squamous cell carcinoma of the skin with cetuximab/pembrolizumab.

Cutaneous squamous cell carcinoma (cSCC) accounts for 20% of skin cancers. At an advanced stage the prognosis is poor, making cSCC the second leading cause of death from skin cancer. In cases of metastatic or unresectable disease, anti-programmed cell death 1 (anti-PD1) treatment has shown promising results in a recent phase II study. Although anti-PD1 treatment now offers higher response rates, the responses remain inconsistent and may lead to therapeutic impasses. Preclinical data have suggested synergy between anti-epidermal growth factor receptor (anti-EGFR) and immunotherapy. We report the case of a patient with metastatic cSCC that proved refractory first to anti-EGFR/carboplatin and then to immunotherapy, but who showed a complete and durable response with cetuximab/pembrolizumab combination. This response could reflect synergy of the two treatments.

When the MET receptor kicks in to resist targeted therapies.

Although targeted therapies have increased the life expectancy of patients with druggable molecular alterations directly involved in tumor development, the efficacy of these therapies is limited by acquired resistances leading to treatment failure. Most targeted therapies, including ones exploiting therapeutic antibodies and kinase inhibitors, are directed against receptor tyrosine kinases (RTKs) or major signaling hubs. Resistances to these therapies arise when inhibition of these targets is bypassed through activation of alternative signaling pathways. In recent years, activation of the receptor tyrosine kinase MET has been shown to promote resistance to various targeted therapies. This casts MET as important actor in resistance. In this review, we describe how the MET receptor triggers resistance to targeted therapies against RTKs such as EGFR, VEGFR, and HER2 and against signaling hubs such as BRAF. We also describe how MET can be its own resistance factor, as illustrated by on-target resistance of lung tumors harboring activating mutations causing MET exon 14 skipping. Interestingly, investigation of all these situations reveals functional physiological relationships between MET and the target of the therapy to which the cancer becomes resistant, suggesting that resistance stems from preexisting mechanisms. Identification of MET as a resistance factor opens the way to co-treatment strategies that are being tested in current clinical trials.

Complete and prolonged response to anti-PD1 therapy in an ALK rearranged lung adenocarcinoma.

OBJECTIVE: Immune checkpoint inhibitors (ICI) have become a major treatment in advanced non small cell lung cancer (NSCLC). However, some patients do not benefit from ICI, especially those harboring an ALK rearrangement. Here, we report a case of prolonged complete tumor response to immunotherapy in an ALK-rearranged NSCLC patient. MATERIALS AND METHODS: We verify ALK expression and rearrangement on formalin-fixed paraffin-embedded tumor samples of the patient by Immunohistochemistry and Fluorescence In Situ Hybridization analysis. The patient provided written informed consent authorizing publication of clinical case. RESULTS: We report the case of 48 years old man with a ALK-rearranged NSCLC. This patient displayed a complete response for 16 months under nivolumab therapy in third line setting after ceritinib and platin based chemotherapy. CONCLUSION: This is the first case of complete and prolonged response to ICI in ALK rearranged NSCLC. This case supports the idea that some ALK rearranged NSCLC could durably benefit from immunotherapy.

Alterations in the PI3K Pathway Drive Resistance to MET Inhibitors in NSCLC Harboring MET Exon 14 Skipping Mutations.

Hepatocyte growth factor receptor (MET) tyrosine kinase inhibitors (MET TKIs) have been found to have efficacy against advanced NSCLC with mutations causing MET exon 14 skipping (METex14 mutations), but primary resistance seems frequent, as response rates are lower than those for targeted TKIs of other oncogene-addicted NSCLCs. Given the known interplay between MET and phosphoinositide 3-kinases (PI3K), we hypothesized that in METex14 NSCLC, PI3K pathway alterations might contribute to primary resistance to MET TKIs. We reviewed clinical data from 65 patients with METex14 NSCLC, assessing PI3K pathway alterations by targeted next-generation sequencing (mutations) and immunohistochemistry (loss of phosphatase and tensin homolog [PTEN]). Using a cell line derived from a patient with primary resistance to a MET TKI and cell lines harboring both a METex14 mutation and a PI3K pathway alteration, we assessed sensitivity to MET TKIs used alone or with a PI3K inhibitor and investigated relevant signaling pathways. We found a phosphatidylinositol 3-kinase catalytic subunit alpha (PIK3CA) mutation in two of 65 samples (3%) and loss of PTEN in six of 26 samples (23%). All three of the MET TKI-treated patients with a PI3K pathway alteration had been found to have progressive disease at first assessment. Likewise, MET TKIs had no effect on the proliferation of METex14-mutated cell lines with a PI3K pathway alteration, including the PTEN-lacking patient-derived cell line. Treatment combining a MET TKI with a PI3K inhibitor caused inhibition of both PI3K and MAPK signaling and restored sensitivity to MET TKIs. PI3K pathway alterations are common in METex14 NSCLC and may confer primary resistance to MET TKIs. In preclinical models, PI3K inhibition restores sensitivity to MET TKIs.

High MET Overexpression Does Not Predict the presence of MET exon 14 Splice Mutations in NSCLC: Results From the IFCT PREDICT.amm study.

INTRODUCTION: MET proto-oncogene (MET) exon 14 splice site (METex14) mutations were recently described in NSCLC and has been reported to correlate with efficacy of MET tyrosine kinase inhibitors. High diversity of these alterations makes them hard to detect by DNA sequencing in clinical practice. Because METex14 mutations induce increased stabilization of the MET receptor, it is anticipated that these mutations are associated with MET overexpression. We aim to determine whether NSCLC with high MET overexpression could define a subset of patients with a high rate of METex14 mutations. METHODS: From The French Cooperative Thoracic Intergroup PREDICT.amm cohort of 843 consecutive patients with a treatment-naive advanced NSCLC who were eligible for a first-line therapy, 108 NSCLC samples with high MET overexpression defined by an immunochemistry score 3+ were tested for METex14 mutations using fragment length analysis combined with optimized targeted next-generation sequencing. MET copy number analysis was also derived from the sequencing data. RESULTS: METex14 mutations were detected in two patients (2.2%) who also displayed a TP53 mutation and a PIK3CA mutation, respectively. An MET gene copy number increase was observed in seven additional patients (7.7%). Next-generation sequencing analysis revealed inactivating mutations in TP53 (52.7%) and PTEN (1.1%), and oncogenic mutations in KRAS (28.6%), EGFR (7.7%), PIK3CA (4.4%), BRAF (4.4%), NRAS (2.2%), GNAS (1.1%), and IDH1 (1.1%). CONCLUSIONS: The rate of METex14 mutations in NSCLC with high MET overexpression was similar to that found in unselected NSCLC. Moreover, we observed a high frequency of driver alterations in other oncogenes. Consequently these findings do not support the use of MET immunohistochemistry as a surrogate marker for METex14 mutations.

Relevance of Detection of Mechanisms of Resistance to ALK Inhibitors in ALK-Rearranged NSCLC in Routine Practice.

BACKGROUND: Anaplastic lymphoma kinase (ALK) tyrosine kinase inhibitors (TKIs) have shown efficacy in the treatment of ALK-rearranged non-small-cell lung cancer (NSCLC), but the disease eventually progresses in all patients. In many cases, resistance to ALK TKIs arises through ALK mutations. Although clinical and biological data suggest variations in TKI efficacy according to the mechanism of resistance, ALK mutations are still rarely investigated in routine practice. MATERIALS AND METHODS: We performed a retrospective multicentric study with an aim to determine the frequency and clinical relevance of ALK alterations detected using targeted next-generation sequencing in patients with advanced ALK-rearranged NSCLC after progression during an ALK TKI treatment. Data on clinical, pathological, and molecular characteristics and patient outcomes were collected. RESULTS: We identified 23 patients with advanced ALK-rearranged NSCLC who, between January 2012 and May 2017, had undergone at least 1 repeat biopsy at progression during an ALK TKI treatment. A resistance mechanism was identified in 9 of the 23 patients (39%). The anomalies involved included 9 ALK mutations in 8 patients and one ALK amplification. The ALK mutation rate was 15% after failure of a first ALK TKI and 33% after failure of 2 ALK TKI treatments. Five of 7 patients who received a different ALK TKI after detection of an ALK mutation achieved an objective response. All of the patients who received a TKI presumed to act on the detected ALK mutant achieved disease control. CONCLUSION: Targeted next-generation sequencing is suitable for detecting ALK resistance mutations in ALK-rearranged NSCLC patients in routine practice. It might help select the best treatment at the time of disease progression during treatment with an ALK TKI.

Optimization of Routine Testing for MET Exon 14 Splice Site Mutations in NSCLC Patients.

INTRODUCTION: Genomic alterations affecting splice sites of MNNG HOS transforming gene (MET) exon 14 were recently identified in NSCLC patients. Objective responses to MET tyrosine kinase inhibitors have been reported in these patients. Thus, detection of MET exon 14 splice site mutations represents a major challenge. So far, most of these alterations were found by full-exome sequencing or large capture-based next-generation sequencing (NGS) panels, which are not suitable for routine diagnosis. METHODS: Aiming to provide a molecular testing method applicable in routine practice, we first developed a fragment-length analysis for detecting deletions in introns flanking MET exon 14. Second, we designed an optimized targeted NGS panel called CLAPv1, covering the MET exon 14 and flanking regions in addition to the main molecular targets usually covered in genomic testing. In patients with MET exon 14 mutations, MET gene amplification, gene copy number and MET receptor expression were also determined. RESULTS: Among 1514 formalin-fixed paraffin-embedded NSCLC samples, nonoptimized NGS allowed detection of MET exon 14 mutations in only 0.3% of the patients, and fragment length analysis detected deletions in 1.1% of the patients. Combined, the optimized CLAPv1 panel and fragment-length analysis implemented for routine molecular testing revealed MET exon 14 alterations in 2.2% of 365 additional NSCLC patients. MET gene amplification or high gene copy number was observed in 6 of 30 patients (20%) harboring MET exon 14 mutations. CONCLUSIONS: These results show that optimized targeted NGS and fragment-length analysis improve detection of MET alterations in routine practice.

Personalized therapy based on sequential molecular analysis leads to 30 months of survival in a patient with diffuse unresectable gastric linitis plastica.

INTRODUCTION: Diffuse gastric cancer is associated with poor prognosis. We report a patient with metastatic gastric linitis plastica harboring human epidermal growth factor receptor 2 (HER2) activating mutation and HER2 amplification. CASE DESCRIPTION: The patient received 5-fluorouracil/folinic acid and oxaliplatin combined with trastuzumab/pertuzumab, resulting in disease control for 8 months. Second-line therapy with nivolumab and trastuzumab/pertuzumab was well-tolerated, with macroscopic peritoneal response. Following ovarian progression and surgical resection of ovarian metastases, immunohistochemistry of PD-L1 was negative; proteomics demonstrated normal expression of HER2 and absence of PD-L1, while genomics showed HER2 amplification, suggesting mechanisms of escape to dual HER2 blockade by downregulation of HER2 and to nivolumab by the absence of PD-L1. Based upon this and nonexpression of biomarkers of taxane resistance, therapy was changed to paclitaxel. Two and a half years after diagnosis, the patient is undergoing treatment, with excellent performance status. CONCLUSIONS: Molecular analysis and personalized therapy can help optimize treatment in difficult-to-treat cancers.

The multiple paths towards MET receptor addiction in cancer.

Targeted therapies against receptor tyrosine kinases (RTKs) are currently used with success on a small proportion of patients displaying clear oncogene activation. Lung cancers with a mutated EGFR provide a good illustration. The efficacy of targeted treatments relies on oncogene addiction, a situation in which the growth or survival of the cancer cells depends on a single deregulated oncogene. MET, a member of the RTK family, is a promising target because it displays many deregulations in a broad panel of cancers. Although clinical trials having evaluated MET inhibitors in large populations have yielded disappointing results, many recent case reports suggest that MET inhibition may be effective in a subset of patients with unambiguous MET activation and thus, most probably, oncogene addiction. Interestingly, preclinical studies have revealed a particularity of MET addiction: it can arise through several mechanisms, and the mechanism involved can differ according to the cancer type. The present review describes the different mechanisms of MET addiction and their consequences for diagnosis and therapeutic strategies. Although in each cancer type MET addiction affects a restricted number of patients, pooling of these patients across all cancer types yields a targetable population liable to benefit from addiction-targeting therapies.

Tumour and cellular distribution of activated forms of PR in breast cancers: a novel immunohistochemical analysis of a large clinical cohort.

BACKGROUND: The progesterone receptor (PR) is expressed by ∼70% of early breast tumours and is implicated in the progression of breast cancer. In cancerous tissues PR may be activated in the absence of a ligand, or when ligand concentrations are very low, resulting in aberrantly activated PR (APR). The presence of APR may indicate that patients with breast cancer are more likely to respond to antiprogestins. The aims of this study were to describe and classify the histological subnuclear morphology of active and inactive PR in archival breast cancer samples. METHODS: Archived tumour specimens from 801 women with invasive breast cancer were collected. Tissue samples (n=789) were analysed for PR isoforms A and B (PRA and PRB), Ki67 and estrogen receptors (ERα) status, using immunohistochemistry. Medical records were used to determine human epidermal growth factor 2 (HER2) status, tumour stage and grade. RESULTS: A total of 79% of tumours stained positive for either PRA or PRB, and of these 25% of PRA-positive and 23% of PRB-positive tumours had PR present in the activated form. APRA was associated with higher tumour grade (p=0.001). APRB was associated with a higher tumour grade (p=0.046) and a trend for a more advanced stage. Patients with PR-positive tumours treated with antiestrogens had better disease-free survival (DFS) than those with PR-negative tumours (p<0.0001). Cumulative progression rate and DFS were similar irrespective of APR status. Both APRA and APRB were independent of HER2, ERα and Ki67 expression. CONCLUSIONS: APR had a binary mode of expression in the breast cancer specimens tested, allowing separation into two tumour subsets. APR is an independent target at the cellular and tumour level and may therefore be a suitable predictive marker for antiprogestins, such as onapristone. Using the described technique, a companion diagnostic is under development to identify APR in solid tumours.

Ipilimumab in anti-PD1 refractory metastatic melanoma: a report of eight cases.

Targeted therapy and immunotherapy in metastatic melanoma have led to a marked improvement in patients' survival and their quality of life. Although there are data on anti-programmed-death-receptor-1 (anti-PD1) after ipilimumab, only few data are available on ipilimumab following anti-PD1 as the first-line treatment. The aim of our study was to evaluate tolerance and survival of patients treated with ipilimumab as the second-line immunotherapy among metastatic melanoma patients following anti-PD1 treatment. Retrospective and descriptive epidemiological studies were carried out at the Dermatology Department of the University Hospital of Lille. We describe a case series of patients treated with ipilimumab after anti-PD1 failure for metastatic melanomas. For each patient, we assessed disease extension since ipilimumab introduction using RECIST 1.1. The time between ipilimumab introduction and other systemic treatment and overall survival (between ipilimumab introduction and last patient visit) was assessed. The effect of ipilimumab after anti-PD1 treatment was evaluated in eight patients. Four patients responded to ipilimumab: three showed a complete response and one showed a partial response. For these patients, the time period between the first ipilimumab injection and another systemic treatment ranged from 209 to 391 days and the overall survival ranged from 314 to 581 days. One patient showed grade 3 chorioretinitis, an unusual toxicity with ipilimumab or anti-PD1 to our knowledge. We have described the efficacy of ipilimumab following anti-PD1 in metastatic melanoma in eight patients. Several comparative studies are still in progress, and their results will be important to develop an optimal therapeutic strategy for our patients.