RAS Mutations in Cutaneous Squamous-Cell Carcinomas in Patients Treated with BRAF Inhibitors

Su F, Viros A, Milagre C, Trunzer K, Bollag G, Spleiss O, et al. RAS mutations in cutaneous squamous-cell carcinomas in patients treated with BRAF inhibitors. N Engl J Med 2012;366:207–15.The clinical development of BRAF inhibitors that block the active confirmation of the BRAF kinase has led to high rates of objective responses in the 50% of melanoma patients whose cancers harbor the BRAF V600E mutation. However, 15% to 30% of these patients develop nonmelanoma skin cancers during the treatment. Su and colleagues have found a molecular mechanism for this unexpected side effect—a paradoxical activation of mitogen-activated protein kinase (MAPK) signaling. This phenomenon had already been noticed in cell lines that had oncogenic RAS or upregulated receptor tyrosine kinases. Here, the authors found that 60% of nonmelanoma skin cancers that arose during BRAF inhibitor treatment had RAS mutations, with the most common being HRAS Q61L. BRAF inhibitors accelerated soft agar growth of cells harboring this mutation, with evidence of increased extracellular signal-regulated kinase (ERK) phosphorylation and increased expression of ERK-regulated genes. The HRAS Q61L mutation also drives skin tumor development in 2-stage murine skin carcinogenesis. In this model, topical administration of DMBA [7,12-dimethylbenz(a) anthracene] causes the mutation, and tumors appear after chronic inflammation is stimulated by repeated application of the tumor promoter TPA (12-0-tetradeanoyl-phorbol-13-acetate). BRAF inhibitor treatment markedly accelerated tumor development in mice treated with DMBA TPA. A MAP–ERK (MEK) inhibitor given alongside the BRAF inhibitor prevented its tumor-accelerating actions. This elegant combination of experiments in biopsies, cell lines, and mouse models suggests that preexisting subclinical skin tumors with strong upstream MAPK signaling potential are accelerated by BRAF inhibitors. However, this unwanted side effect may be avoided by addition of a MEK inhibitor to the treatment schedule. This article also highlights the importance of genotyping tumors prior to administering BRAF-selective drugs to patients. This way it would be possible to distinguish patients who are likely to respond positively to a therapy with BRAF inhibitors from those who may develop nonmelanoma skin cancers.Garofalo M, Romano G, Di Leva G, Nuovo G, Jeon YJ, Ngankeu A, et al. EGFR and MET receptor tyrosine kinase–altered microRNA expression induces tumorigenesis and gefitinib resistance in lung cancers. Nature Med 2011;18:74–82.The tyrosine kinase inhibitors (TKI), gefitinib and erlotinib, are currently used for treating individuals with non–small cell lung cancers (NSCLC) because they target EGFR, which is activated in these tumors. Acquired resistance to these TKIs is a vexing problem for the biotechnology industry and clinicians alike. The mechanism that leads to drug resistance is not well understood, although previous studies have implicated the MET oncogene in both de novo and acquired resistance to TKIs. Still, how MET overexpression biologically triggers this process was unknown. To address this concern, Garofalo and colleagues hypothesized that microRNAs that act as master regulators of a plethora of cellular processes, including proliferation, apoptosis, invasion, and epithelial-to-mesenchymal transition (EMT), play a key role in mediating TKI resistance. Using independent short hairpin RNA approaches against EGFR and MET, they identified the microRNAs miR-30b, miR-30c, miR-221, and miR-222 as being induced by both EGF and MET receptor signaling, whereas miR-103 and miR-203 were specifically downregulated by MET. Clinical analyses in a cohort of lung tumors showed a positive association between miR-30b, miR-30c, miR-221, and miR-222, and EGFR-MET expression as well as an inverse correlation between miR-103 and miR-203, and MET. The authors further identified BIM and APAF-1 as common targets of the EGFR-MET controlled microRNA complex that lead to gefitinib resistance. Silencing miR-30 or miR-221 or overexpressing miR-103 or mir-203 induced apoptosis and decreased tumor size in response to gefitinib. Complementary to this finding, MET induced EMT via inhibition of miR-103 and miR-203, and caused enhanced migration by relieving the repressive effect of these microRNAs on their targets, PKC-ϵ and SRC. In totality, these findings suggest that modulation of specific microRNAs by EGFR-MET contributes to gefitinib resistance and therefore represents a novel therapeutic approach for treatment of NSCLCs. This multidimensional approach could have implications for lung cancer therapy, for example by stratifying patients based on MET-microRNA regulon to select those who might benefit from treatment with gefitinib.Kinross KM, Montgomery KG, Kleinschmidt M, Waring P, Ivetac I, Tikoo A, et al. An activating Pik3ca mutation coupled with Pten loss is sufficient to initiate ovarian tumorigenesis in mice. J Clin Invest 2012;122:553–7.The phosphoinositide 3-kinase (PI3K) pathway is one of the most commonly deregulated pathways in cancer. Somatic mutations in a gene encoding the p110 alpha subunit (PIK3CA) enhance PI3K activity and are often found in human cancer. However, the role of increased PIK3CA activity in tumor development and progression remains unknown. Kinross and colleagues have generated the first mouse model harboring an inducible knock-in of the human cancer-associated PIK3CA H1047R mutation (PIK3CAH1047R) to study the contribution of this mutation towards ovarian tumorigenesis. The authors demonstrate that PIK3CAH1047R expression alone is insufficient to induce AKT phosphorylation in vivo, and is only weakly tumorigenic. However, the PIK3CA mutant combined with biallelic PTEN deletion (PTENdel/del) leads to AKT hyperphosphorylation at serine-473, threonine-308, and phosphorylation of RPS6, as well as the formation of high-grade serous adenocarcinomas and granulosa cell tumors. These results are especially significant in light of the fact that PIK3CA mutations are commonly coexpressed alongside PTEN loss or oncogenic KRAS in human ovarian cancer. Importantly, Kinross and colleagues further show that pharmacologically inhibiting the PI3K and mTOR signaling pathways increases the survival rates of PIK3CAH1047R/PTENdel/del mutant mice. The authors' findings have strong clinical relevance for the development of new treatment regimens for human epithelial cancers harboring PIK3CA and PTEN mutations.Bronisz A, Godlewski J, Wallace JA, Merchant AS, Nowicki MO, Mathsyaraja H, et al. Reprogramming of the tumor microenvironment by stromal PTEN-regulated miR-320. Nature Cell Biol 2011 Dec 18. [Epub ahead of print].The functional role of stromal cells such as fibroblasts in tumor development and metastasis is important but poorly understood. Although reports in the literature demonstrate that tumorassociated fibroblasts can provide a favorable environment for cancer initiation and progression, the mechanisms underlying this effect remain unclear. In this article, Bronisz and colleagues uncover a new tumor-suppressive axis in stromal fibroblasts linking PTEN, the microRNA miR-320, and transcription factor v-ets erythroblastosis virus E26 oncogene homolog 2 (ETS2), which is important for reprogramming tumor cells in cancer initiation and progression. The authors used genetic strategies in stromal fibroblasts surrounding the mammary epithelial ducts to compare the expression of microRNAs in mouse mammary fibroblasts (MMF) with and without PTEN. They identified a group of microRNAs that are differentially expressed in PTEN−/− cells compared with wild-type cells. The work of Bronisz and colleagues functionally validates that miR-320 plays a cell–nonautonomous tumor-suppressive role in stromal cells. Specifically, coinjecting breast cancer tumor cell line DB7 with PTEN−/− MMFs resulted in a 4-fold increase in tumor growth compared with DB7 coinjected with PTEN+/+ MMFs. Interestingly, restoration of miR-320 in PTEN−/− cells significantly decreases tumor growth. The authors also show that downregulation of miR-320 in PTEN−/− MMFs influences gene expression of the surrounding epithelial or endothelial tumor cells. They further demonstrate that miR-320 in stromal fibroblasts influences the invasiveness of epithelial tumor cells through the action of secreted factors. Using a proteomics analysis of conditional media of PTEN−/− MMFs expressing either miR-320 or control, Bronisz and colleagues discovered a signature of miR-320 tumor-regulating secretory factors that, when PTEN is lost, act to increase the migration and invasive behavior of epithelial and endothelial tumor cells. The authors further demonstrate that these factors can be categorized into different groups: One group is comprised of proteins that are direct targets of miR-320, and a larger group transcriptionally regulates ETS2, which itself is a target of miR-320. Bronisz and colleagues show that the miR-320 secretome signature can be used to discriminate between normal and tumor stroma in human breast cancer. These findings highlight the functional role of stromal cells in directly initiating cancer or acting as a metastatic switch.Putluri N, Shojaie A, Vasu VT, Vareed SK, Nalluri S, Putluri V, et al. Metabolomic profiling reveals potential markers and bioprocesses in bladder cancer progression. Cancer Res 2011;71:7376–86.It is estimated that 69,250 new cases of bladder cancer and 14,990 deaths from bladder cancer occurred in 2011. The current approach for diagnosing and monitoring bladder cancer is cystoscopy, aided by cytology and biopsy. Urine cytology provides acceptable sensitivity and specificity in the context of high-grade bladder cancer, whereas sensitivity is poor for lowgrade tumors. Development of assays with enhanced sensitivity in permitting disease stratification is important in the management of this disease. Using high-throughput mass spectrometry to measure over 2,000 compounds in 58 clinical specimens, Putluri and colleagues have identified 35 metabolites displaying measurable changes in bladder cancer. These metabolomics signatures in urine provide potential markers for distinguishing bladder cancer from controls and also nonmuscle from muscle-invasive bladder cancer. A potential role for DNA methylation in altering phase I/II xenobiotic metabolism in bladder cancer was suggested by enrichment-based bioprocess mapping. With the use of bisulfite sequence analysis and methylation-specific PCR, in addition to in vitro experiments with T-24 bladder cancer cells treated with 5-aza-2′-deoxycytidine, hypomethylation in the cytochrome P450 1A1 (CYP1A1) and cytochrome P450 1 B1 (CYP1B1) promoters in bladder cancer was identified. Although further studies are required with a much larger cohort of patients, metabolomics signatures described in this article may provide a basis for potentially developing urinary biomarkers for early detection and staging of bladder cancer. Additionally, this study highlights the importance of epigenetic changes in metabolic profiles as potentially underlying factors in determining bladder tumor development and progression.Note: Breaking Advances are written by Cancer Research Editors. Readers are encouraged to consult the articles referred to in each item for full details on the findings described.

PCN84 COST OF ILLNESS OF CUTANEOUS SQUAMOUS CELL CARCINOMA IN ITALY

RAS Mutations in Cutaneous Squamous-Cell Carcinomas in Patients Treated with BRAF Inhibitors

Faculty Opinions recommendation of RAS mutations in cutaneous squamous-cell carcinomas in patients treated with BRAF inhibitors.

Faculty Opinions recommendation of RAS mutations in cutaneous squamous-cell carcinomas in patients treated with BRAF inhibitors.

Metastatic squamous cell carcinoma in chronic lymphocytic leukaemia in a haematology-dermatology multidisciplinary clinic.

Multiple refined histopathological variables have been proposed to improve risk stratification of cutaneous squamous cell carcinoma (CSCC) patients, but evidence of their added value over conventional clinicopathological variables such as depth of invasion or differentiation grade is limited. To evaluate the association between refined histopathological variables and metastasis in CSCC and their value relative to routinely available variables. We conducted two nested case-control (NCC) studies using data from the Netherlands Cancer Registry and nationwide pathology registry with over 10 years of follow-up (n=19,120 patients). The first NCC study included a population-based set of 195 primary CSCCs of metastatic cases matched to 195 primary CSCCs of non-metastatic controls only based on follow-up time. The second NCC study included a metastatic risk-matched set, matching 250 cases with 250 controls based on calculated metastatic risk. Pathologists reviewed all primary CSCCs, including the refined variables: morphological subtype, solar elastosis, peritumoral infiltrate, tumour budding, and mitosis. Analyses were conducted using conditional logistic regression. In the population-based set, multivariable analyses showed no significant associations between refined histopathological variables and metastasis. In the risk-matched set, severe (OR 0.28; 95%CI 0.09-0.87) and moderate (OR 0.20; 95%CI 0.07-0.56) solar elastosis were negatively associated with metastasis. Most refined histopathological variables may not provide additional prognostic value beyond conventional risk factors for metastasis in CSCC patients. The observed inverse association between solar elastosis and metastasis warrants further research.

Ultra-short immunotherapy may spare cutaneous squamous cell carcinoma (CSCC) patients from mutilating surgery, but early identification of (non-)response is needed to safely guide treatment adaptation. This study evaluated the feasibility of sequential [18F]FDG-PET/CT (FDG-PET) as a response biomarker in resectable CSCC patients. In the MATISSE, a randomized phase-II trial, 50 CSCC patients received two courses of neoadjuvant nivolumab (weeks 0 and 2) with or without low-dose ipilimumab (week 0) before surgery (week 4). FDG-PET scans were obtained pre-treatment and shortly prior to surgery to assess the change (Δ) in maximum standardized uptake value (SUVmax), metabolic tumor volume (MTV), and total lesion glycolysis (TLG) at the primary tumour and largest (= index) lymph node metastasis (ILN). ΔMTV50%/4.0 and ΔTLG50%/4.0 were calculated using thresholds of 50% SUVmax and SUV ≥ 4.0. In 42 evaluable patients, 31 (74%) patients showed major or partial responses to immunotherapy. EORTC-criteria underestimated response but accurately identified non-responders (70% sensitivity, 100% specificity). In 28 primary tumours and 22 ILNs, a significant reduction in median SUVmax, MTV50%, MTV4.0, TLG50%, and TLG4.0 was observed in responders versus non-responders (overall, p ≤ 0.004, and p ≤ 0.03, respectively). ΔTLG50% and ΔTLG4.0 correlated strongly with response (primary: 92% and 96% accuracy; ILN: 91% and 89% accuracy). Quantitative FDG-PET-response assessment allows early identification of (non-)responders upon neoadjuvant immunotherapy prior to surgery in locoregionally advanced CSCC patients. Early changes in FDG-PET's TLG can support future trials aiming at safe de-escalation of current standard of care surgery with or without adjuvant radiotherapy. EudraCT 2020-001074-30. Registered 9 March 2020, https://www.clinicaltrialsregister.eu/ctr-search/search?query=2020-001074-30.

Cutaneous adverse events (CAEs) after treatment with BRAF and MEK inhibitors in patients with melanoma remain incompletely characterized. To determine the association of BRAF and MEK inhibitor treatment with CAEs in patients with melanoma compared with BRAF inhibitor alone. PubMed, Cochrane, Embase and Web of Science were systematically searched for BRAF and MEK inhibitors from database inception through 10 May 2024. Randomized clinical trials reporting on CAEs in patients with melanoma being treated with BRAF and MEK inhibitors compared with patients with melanoma being treated with BRAF inhibitor monotherapy were selected. Pooled Risk ratios (RRs) and 95% CIs were determined using random-effects analyses. The selected end points were alopecia, cutaneous squamous-cell carcinoma, hyperkeratosis, keratoacanthoma, palmoplantar erythrodysaesthesia syndrome, palmoplantar keratoderma, rash, photosensitivity reaction, and skin papilloma. All-grade and high-grade (≥3) CAEs were recorded. Comparing with BRAF and MEK inhibitors, treatment with BRAF inhibitors alone was associated with an increased risk of rash (RR, 0.73; 95% CI, 0.54-0.99; p = 0.039; I2 = 88%), alopecia (RR, 0.28; 95% CI, 0.20-0.41; P < 0.001; I2 = 76%), hyperkeratosis (RR, 0.30; 95% CI, 0.22-0.41; P < 0.001; I2 = 56%), palmoplantar erythrodysaesthesia syndrome (RR, 0.21; 95% CI, 0.10-0.47; P < 0.001; I2 = 81%), palmoplantar keratoderma (RR, 0.39; 95% CI, 0.26-0.57; P < 0.001; I2 = 29%), Skin papilloma (RR, 0.25; 95% CI, 0.12-0.52; P < 0.001; I2 = 77%), cutaneous squamous-cell carcinoma (RR, 0.21; 95% CI, 0.11-0.42; P < 0.001; I2 = 50%), and keratoacanthoma (RR, 0.22; 95% CI, 0.12-0.40; P < 0.001; I2 = 0%). Therapy with BRAF and MEK inhibitors was associated with a lower risk of CAEs, especially rash, alopecia, hyperkeratosis, palmoplantar erythrodysaesthesia syndrome, palmoplantar keratoderma, skin papilloma, cutaneous squamous-cell carcinoma, and keratoacanthoma, compared with BRAF inhibitor alone. The risks of photosensitivity reaction was similar between the assessed groups. The findings may help to balance between beneficial melanoma treatment and cutaneous morbidity and mortality.

Rapid response to cemiplimab for advanced cutaneous squamous cell carcinoma

A 5.7-year prospective study of 1380 patients treated for psoriasis with oral methoxsalen (8-methoxypsoralen) and ultraviolet A photochemotherapy (PUVA) revealed that after adjustment for exposures to ionizing radiation and topical tar preparations, the risk that cutaneous squamous-cell carcinoma would develop at least 22 months after the first exposure to PUVA was 12.8 times higher in patients exposed to a high dose than in those exposed to a low dose (95 per cent confidence interval, 5.8 to 28.5). No substantial dose-related increase was noted for basal-cell carcinoma. The dose-dependent risk of cutaneous squamous-cell carcinoma suggests that PUVA can act as an independent carcinogen. In our study, morbidity associated with these tumors has been limited, but further follow-up is needed. Meanwhile, patients treated with PUVA should be followed closely for the possible development of cutaneous squamous-cell carcinoma.

Objective To measure serum and tissue levels of soluble syndecan-1 (SDC1) in patients with cutaneous squamous cell carcinoma (CSCC) , and to explore the relationship between the expression of SDC1 and clinicopathologic features of CSCC as well as between the serum and tissue levels of SDC1. Methods An immunohistochemical study was performed to measure SDC1 expression in the epidermis of lesional specimens from 93 patients with CSCC and normal skin specimens from 30 healthy human controls, and enzyme-linked immunosorbent assay (ELISA) to detect serum levels of soluble SDC1 in 81 patients with CSCC and 30 healthy human controls. Results The expression of SDC1 was significantly lower in CSCC tissues than in normal skin tissues (Z = 3.85, P 0.05) . CSCC patients with lymphatic metastasis showed significantly higher serum levels of SDC1 compared with those without (Z = 5.30, P < 0.01) . The serum levels of soluble SDC1 were significantly negatively correlated with its tissue levels in CSCC patients (rs =-0.625, P < 0.01) . Receiver operating characteristic (ROC) curve analysis showed that the best cutoff point of serum SDC1 levels was 65.5 μg/L for the diagnosis of lymphatic metastasis, with the sensitivity, specificity and area under the curve (AUC) being 73.7% , 87.1% and 0.904 (0.840-0.968) respectively. Conclusion The down-regulated tissue expression but up-regulated serum levels of SDC1 may be associated with the invasiveness of CSCC, and elevated serum SDC1 levels are somewhat valuable to the diagnosis of lymphatic metastasis. Key words: Carcinoma, squamous cell; Skin; Serum; Pathology; Syndecan-1

BRAF inhibitor-based therapies have been shown to induce cutaneous toxicities, with onset generally in the first 8-26weeks of therapy. To determine whether cutaneous toxicities persist in patients who have remained on BRAF inhibitor-based therapies for longer than 52weeks, and therefore whether ongoing dermatology assessment is required. All patients treated with the BRAF inhibitors vemurafenib or dabrafenib or combination BRAF inhibitor and mitogen-activated protein kinase kinase (MEK) inhibitor therapy at Westmead Hospital, Sydney, Australia underwent regular dermatological assessments for the duration of therapy. All patients enrolled in a clinical trial, and 18% of patients in the compassionate access scheme underwent a baseline assessment prior to commencement of therapy and every 4-8weeks thereafter. Patients' adverse events were recorded in a specific database. Patients continued to develop cutaneous adverse events after 52weeks of continuous therapy. Patients on single-agent BRAF inhibitor therapy suffered from Grover disease (45%), plantar hyperkeratosis (45%), verrucal keratosis (18%) and even cutaneous squamous cell carcinoma (16%). The most frequent adverse event seen in patients in the combination BRAF and MEK inhibitor group was an acneiform eruption (40%). Patients on BRAF inhibitor-based therapies need to continue to have regular dermatological follow-up independent of the duration of their therapy.

Radiotherapy for management of basal and squamous cell carcinoma

Since hedgehog (HH)/Gli1 that contributes to cancer proliferation and metastasis has been masked for decades, the signaling pathway was investigated about its exact role in proliferation and metastasis of cutaneous squamous cell carcinoma (SCC). Sonic hedgehog homolog (Shh), GLI family zinc finger 1 (Gli1), and vascular endothelial growth factor (VEGF) expressions in cutaneous SCC tissues were analyzed with immunohistochemistry, and their correlations with cutaneous SCC patients' prognosis were conducted with Kaplan-Meier curve. Regarding in vitro studies, effects of the HH signaling pathway, and cyclopamine on patched 1 (Ptch1), smoothened/frizzled class receptor (Smo) and VEGF expressions were assessed in A431 cells based on western blot and quantitative real-time polymerase chain reaction (qRT-PCR). Besides, Cell Counting Kit-8 (CCK-8) assay was implemented to evaluate cell proliferation, while wound-healing assay and transwell assay were performed to assess cell migration and invasion, respectively. Mice models were also established to observe effects of Gli1 on tumor diversity and incidence during a period of 20weeks. Positively expressed VEGF, Gli1, and Shh proteins in cutaneous SCC tissues were correlated with poor survival of patients (P<0.05). Besides, Gli1 messenger RNA (mRNA) and VEGF mRNA were observed to be significantly over-expressed in A431 cells (P<0.05), and they were associated with incremental cell proliferation, invasiveness, and migration, which can be reversed by the interference of VEGF siRNA. Furthermore, cyclopamine treatment could induce inhibition of cell proliferation, invasiveness, and migration and suppression of Smo, Gli1, and VEGF expressions. The mice models also confirmed that Gli1 could significantly induce rise of tumor incidence and tumor diversity, while cyclopamine statistically relieved this transformation (P<0.05). Abnormal activation of the HH signaling pathway plays critical roles in development of cutaneous SCC either in vivo or in vitro.