Primary cSCC Research Articles

Cancer-associated fibroblast activation predicts progression, metastasis, and prognosis of cutaneous squamous cell carcinoma.

Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer and the metastatic disease is associated with poor prognosis. Cancer-associated fibroblasts (CAFs) promote progression of cancer, but their role in cSCC is largely unknown. We examined the potential of CAF markers in the assessment of metastasis risk and prognosis of primary cSCC. We utilized multiplexed fluorescence immunohistochemistry for profiling CAF landscape in metastatic and non-metastatic primary human cSCCs, in metastases, and in premalignant epidermal lesions. Quantitative high-resolution image analysis was performed with two separate panels of antibodies for CAF markers and results were correlated with clinical and histopathological parameters including disease-specific mortality. Increased stromal expression of fibroblast activation protein (FAP), α-smooth muscle actin, and secreted protein acidic and rich in cysteine (SPARC) were associated with progression to invasive cSCC. Elevation of FAP and platelet-derived growth factor receptor-β (PDGFRβ) expression was associated with metastasis risk of primary cSCCs. High expression of PDGFRβ and periostin correlated with poor prognosis. Multimarker combination defined CAF subset, PDGFRα-/PDGFRβ+/FAP+, was associated with invasion and metastasis, and independently predicted poor disease-specific survival. These results identify high PDGFRβ expression alone and multimarker combination PDGFRα-/PDGFRβ+/FAP+ by CAFs as potential biomarkers for risk of metastasis and poor prognosis.

Lymphovascular Invasion in Cutaneous Squamous Cell Carcinoma.

Although there is a large body of literature regarding risk stratification and outcomes for perineural invasion (PNI) in cutaneous squamous cell carcinoma (cSCC), there is a relative paucity of studies exploring the role of lymphovascular invasion (LVI) in cSCC and a lack of clear evidence-based guidelines for how to manage patients with these tumors. This article is intended to review the available literature regarding LVI in cSCC and formulate evidence-based recommendations for clinical management. A literature review was conducted using PubMed to find relevant articles relating to outcomes and management of primary cSCC with LVI. The available literature suggests that LVI is a major risk factor for poor outcomes and increased morbidity and mortality in cSCC. Lymphovascular invasion is a very high-risk feature that should place these tumors in the highest-risk category, and management of these tumors should be similar to that of squamous cell carcinoma with PNI.

Abstract LB207: C5ar1 is a novel biomarker and potential therapeutic target in advanced cutaneous squamous cell carcinoma

Abstract Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer and the metastatic disease is associated with poor prognosis. At present, there are no established molecular markers to predict the metastasis risk and prognosis of patients with primary cSCC. Recently, the role of specific complement components in the progression of cSCC has been shown. Here, we have examined the role of complement C5a receptor C5aR1 in the progression and metastasis of cSCC. The results show that the expression of C5aR1 is increased in cSCC cells in 3D spheroid coculture model in the presence of fibroblasts and that treatment with recombinant C5a increases invasion of cSCC cells in this model. Staining for C5aR1 was detected on the surface of tumor cells in the invasive edge of human cSCC xenograft tumors in vivo. Staining of metastatic and non-metastatic primary human cSCCs, premalignant epidermal lesions and normal skin for C5aR1 with multiplexed fluorescence and chromogenic immunohistochemistry revealed stronger expression of C5aR1 on surface of cSCC tumor cells compared to epidermal cells in cSCC in situ, actinic keratosis and normal skin. Increased expression of C5aR1 on tumor cell surface was associated with the metastasis risk of primary cSCCs and independently predicted poor disease-specific survival of patients with primary cSCC. These results reveal the role of C5aR1 in cSCC progression and identify C5aR1 as a novel biomarker for metastasis risk and poor prognosis of patients with cSCC. The results also suggest C5aR1 as a novel therapeutic target for treatment of locally advanced and metastatic cSCC. Citation Format: Lauri Heiskanen, Liisa Nissinen, Elina Siljamäki, Jaakko Knuutila, Teijo Pellinen, Markku Kallajoki, Jyrki Heino, Pilvi Riihilä, Veli-Matti Kähäri. C5ar1 is a novel biomarker and potential therapeutic target in advanced cutaneous squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB207.

Abstract CT004: A single arm phase 2 study of TVEC in patients with invasive cutaneous SCC: A novel therapeutic approach for low risk tumors

Abstract Background: Immune recognition and cytotoxic responses play an important role in the pathogenesis and progression of cutaneous squamous cell carcinoma (cSCC). Talimogene laherparepvec (TVEC) is an HSV-1 oncolytic immunological agent FDA approved for the local treatment of unresectable recurrent melanoma. It is proposed that T-VEC directly destroys cancer cells, and induces production of GM-CSF to enhance systemic antitumor immune priming. This proposed mechanism of action supports the novel approach to implement TVEC in the management of cSCC, particularly, in patients with increased burden of primary tumors. Methods: Study subjects were Immunocompetent, ≥ 18 years of age, and diagnosed with at least one histologically confirmed primary low-risk cSCC according to the Brigham and Women staging system. Unresectable lesions or patients unable/unwilling to undergo standard of care treatment were eligible to participate. Study lesions included target lesions injected (TLIs) and target non-injected lesions (TNILs). TNILs were selected to evaluate for abscopal effect when feasible. The TLIs were treated according to TVEC FDA approved protocol and followed for 1yr after the 1st injection. The primary study endpoint was to evaluate the overall response rate, defined as the proportion of subjects who achieved complete response and partial response in the TLIs. Safety and adverse effect profile (AEs), duration of response, time to response, durable response rate, and time to progression, were the secondary endpoints. A Simon 2 stage design was used. Total sample size was calculated to be 20 subjects. A total of 11 subjects were recruited, due to the COVID-19 pandemic and 100% CR observed in the interim analysis. NCT 03714828. Results: The analysis was conducted after all 11 participants completed Visit 8 (230-271 days from study initiation). Conclusions: TVEC showed extremely impressive ORR for patients with cSCC with a 95.8% CR for TLIs. AE profile of TVEC was mild and was well tolerated. Further studies evaluating the role of intralesional immunotherapy should be considered in patients with increased burden of cSCC. Table 1. summarizes the study results. For the primary endpoint 100% of participants achieved an overall response. Primary outcomes for patients (n = 11) Secondary outcomes for TLIs (n = 24) Time to response - days Mean ± standard deviation Median/Range (min, max) 41.6 ± 24.9 Time to response - days Mean ± standard deviation Median/Range (min, max) 48.7 ± 29.2 35/(18 - 112) 35/(18 - 112) Final response to TVEC - % Overall response rate (ORR) Partial response (PR) Complete response (CR) 100% (11/11) 9.1% (1/11) Response to TVEC - Overall response rate (ORR) Partial response (PR) Complete response (CR) 100% (24/24) 90.9% (10/11) 4.2% (1/24) 95.8% (23/24) Duration of response - days Mean ± standard deviation Median - Range (min, max) 206.0 ± 26.0 Duration of response - days Mean ± standard deviation Median - Range (min, max) 201.4 ± 29.6 212 (140 - 236) 209 (136 - 250) Durable response rate - % 90.9% (10/11) Durable response rate - % 83.3% (20/24) Secondary outcomes for TNILs (n = 2) Final response to TVEC - % 100% (2/2) Adverse Events NCI CTCAE (v. 4.0) Event n (%) 95% CI Severity Mild (Grade 1) Moderate (Grade 2) Fatigue 5 (45.5%) (16.7%, 76.6%) 4 (80%) 1 (20%) Flu-like symptoms 4 (36.4%) (10.9%, 69.2%) 3 (75%) 1 (25%) Headache 2 (18.2%) (2.3%, 51.7% 2 (100%) Citation Format: Clara Curiel-Lewandrowski, Delaney B. Stratton, Anngela C. Adams, Haiyan Cui, Denise Roe, Srinath Sundararajan. A single arm phase 2 study of TVEC in patients with invasive cutaneous SCC: A novel therapeutic approach for low risk tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT004.

Diagnosis of invasive cutaneous squamous cell carcinoma, imaging and staging.

The assessment of patients with a lesion raising the suspicion of an invasive cutaneous squamous cell carcinoma (cSCC) is a frequent clinical scenario. The management of patients with cSCC is a multistep approach, starting with the correct diagnosis. The two main diagnostic goals are to differentiate from other possible diagnoses and correctly recognize the lesion as cSCC, and then to determine the tumor spread (perform staging), that is if the patient has a common primary cSCC or a locally advanced cSCC, or a metastatic cSCC (with in-transit, regional lymph nodal, or rarely distant metastasis). The multistep diagnostic approach begins with the clinical characteristics of the primary cSCC, it is complemented with features with dermoscopy and, if available, reflectance confocal microscopy and is confirmed with histopathology. The tumor spread is assessed by physical examination and, in some cases, ultrasound and/or computed tomography or magnetic resonance imaging, mainly to investigate for regional lymph node metastasis or for local infiltration into deeper structures. In the last step, the clinical, histologic and radiologic findings are incorporated into staging systems.

Abstract 279: Iatrogenic immunosuppression emplaces tacrolimus to drive squamous skin cancer progression and influence the microenvironment

Abstract One in thirteen organ transplant recipients (OTRs) develop post-transplant cutaneous squamous cell carcinoma (cSCC), leading to significant morbidity upon disease progression. Although chronic ultraviolet (UV) exposure primarily underlies keratinocyte carcinogenesis, long-term immunosuppression is a critical etiologic risk factor due to reduced systemic surveillance of dysplasia. Calcineurin inhibitors such as tacrolimus are commonly administered to suppress the immune system by blocking T-cell proliferation and preventing donor organ rejection. Contrarily, studies have evinced the off-target effects of immunosuppressives, namely azathioprine and cyclosporine, but not tacrolimus — the most widely used in all immunosuppression. In addition, the preclinical research literature has overlooked the contribution of stromal fibroblasts in the cSCC tumor microenvironment (TME). Hence, we hypothesize that tacrolimus exerts direct carcinogenic effects on keratinocytes and fibroblasts in the skin. We further postulate that tacrolimus enables a tumor-permissive microenvironment by activating dermal fibroblasts to cause aggressive and invasive keratinocyte carcinoma in OTRs. To examine this cellular crosstalk phenomenon, we used human keratinocytes, primary dermal fibroblasts and cSCC cells in mono- and co-cultures to mimic cutaneous and tumor complexities. Next, we assessed the molecular mechanisms utilized by tacrolimus to potentiate tumorigenesis pertinent to cell survival, proliferation and invasion. Herein, we show that tacrolimus promotes cell survival dose-dependently in cSCC cells. We also found that a fibroblast-cSCC spheroid co-culture significantly enhances cell invasion. In a wound-healing migration assay, tacrolimus increased the migratory capacity of normal keratinocytes and cSCC cells. However, cell proliferation remains nebulous due to cyclical dose-response effects in vitro. We then queried whether cell proliferation is affected in vivo using immunodeficient mice, and similar results were observed. We confirmed via immunoblotting that tacrolimus induced elevated expression of proliferation markers at distinct drug concentrations. Altogether, our findings illuminate keratinocyte-stroma interactions and particularly isolate the contributory role of tacrolimus in non-immune mediated cSCC pathogenesis. This knowledge suggests newer immunosuppressives that do not inhibit calcineurin activity may help relieve cutaneous malignancies in transplant recipients. Citation Format: Edwin Kumah, Yash Chhabra, Vania Wang, Agrani Dixit, Ashani Weeraratna, Kristin Bibee. Iatrogenic immunosuppression emplaces tacrolimus to drive squamous skin cancer progression and influence the microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 279.

Abstract 4869: Efficient study design for the discovery of a gene expression signature predicting metastasis in cutaneous squamous cell carcinoma

Abstract Background: The StepIdent study aims to develop a gene signature predicting metastasis in patients with cutaneous squamous cell carcinoma (cSCC) to improve risk stratification, thus enabling personalized decisions about follow-up schedules and treatment options. Here we describe the unique characteristics, challenges, and best practices for an efficient design of a discovery cohort for a rare outcome (metastasis prevalence: 2-5%); for retrieving, curating, and linking the clinical and pathological data through nationwide databases; and for measuring gene expression through sequencing of archived Formalin-Fixed Paraffin-Embedded (FFPE) primary tumor samples. Methods: Following a predefined protocol, we identified a nested-case control cohort (NCC) of 305 cases and 305 controls from a nationwide cohort of 19,120 patients with a first cSCC in the Netherlands from 2007 to 2009, followed up until 2020. We chose an NCC design since it is an efficient study design in a rare outcome setting (weighting is needed to accommodate the under-sampling of the controls). Patients were identified from the Dutch National Cancer Registry (NCR) and the clinical information was retrieved from the NCR which is linked to the nationwide registry of histo- and cytopathology (PALGA). Tumor blocks were requested from PALGA, and pathological characteristics were assessed by dermatopathologists. We matched controls to cases, based on a risk score estimated by a clinicopathological model. Gene expression was measured using the Illumina RNA Prep with Enrichment kit combined with the whole exome panel and paired-end sequenced on the NextSeq 550. Results: Tissue slides for 541 samples were retrieved for sequencing. 151 samples were excluded after pathology review or due to low pre-library concentration. The final cohort includes 195 case-control pairs (n=390). The median sequencing depth was 43M (Q1-Q3: 35-52M); the median Q30 was 85% (Q1-Q3: 83-87%); the median GC content was 51% (Q1-Q3: 50-52%); a median of 1.8% of base pairs (Q1-Q3: 1.4-2.1%) was trimmed prior to the mapping/alignment; a median of 69% (Q1-Q3: 65-74%) of reads were aligned as protein-coding and a median of 7% (Q1-Q3: 6-10%) as rRNA; a median of 95% (Q1-Q3: 93-96%) of reads were aligned by STAR. Two samples were excluded based on quality control. Conclusion: We described an efficient design and implementation of a nationwide discovery study in cSCC, involving the retrieval of clinicopathological data, the collection of FFPE materials, and the execution of omics measurements. This study presents the largest cohort to date, incorporating omics measurements of primary cSCC samples, combined with simultaneous access to well-curated clinical and pathological information and follow-up data. Our findings can provide guidance for similar studies involving a rare clinical endpoint, where an efficient study design is a necessity. Citation Format: Barbara Rentroia-Pacheco, Lara Pozza, Yan Ting Chen, Daphne Huigh, Celeste J. Eggermont, Olivia FM Steijlen, Sheril Alex, Jvalini Dwarkasing, Domenico Bellomo, Harmen JG van de Werken, Antien L. Mooyaart, Marlies Wakkee, Loes M. Hollestein. Efficient study design for the discovery of a gene expression signature predicting metastasis in cutaneous squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4869.

The 40-gene Expression Profile (40-GEP) Test Identifies Cutaneous Squamous Cell Carcinoma (cSCC) Patients at High Risk of Metastasis within Lower-Staged Tumors to Better Guide Treatment Decisions

Introduction- The development of metastases has a profound impact on cSCC patient survival. Lower-stage tumors are those that lack factors that portend higher risk for disease progression and therefore not typically considered for increased surveillance or treatment. However, up to one-third of all metastatic events have been reported for patients originally staged as T1. The 40-GEP test is validated to accurately classify risk for regional and/or distant metastasis in patients with primary cSCC with at least one high-risk factor. The current study investigated whether the 40-GEP test could independently improve identification of lower-staged tumors at increased risk of metastasis. 
 Methods- In an IRB-approved, retrospective, multi-center study, primary tumor tissue and associated clinical data from patients with cSCC and one or more clinicopathologic risk factors (n=897) were collected. Within this overall cohort, lower Brigham and Women’s Hospital (BWH) T-staged samples (n=444 T1 and n=335 T2a) were evaluated by Kaplan-Meier survival analysis to determine metastasis-free survival (MFS) according to 40-GEP risk class (Class 1-low metastatic risk; Class 2A-higher risk; Class 2B-highest risk).
 Results- There was a total of 118 metastatic events in the overall cohort, of which 25% (29/118) and 38% (45/118) occurred in BWH T1 and T2a, respectively. BWH T1 tumors had an overall event rate of 6.5% (29/444), with T1/Class 2B tumors having a 5.1-fold increased event rate of 33.3%. BWH T2a tumors had an overall event rate of 13.4% (45/335), with T2a/Class 2B tumors having a 2.7-fold increased event rate of 36.4%. Stratification by the 40-GEP test resulted in significantly different metastasis rates across all three 40-GEP classes within both the T1 and T2a subsets (log-rank tests, p<0.0001 each subset). 
 Conclusion- Within a cohort of cSCC patients considered lower risk by current staging alone, the 40-GEP identified patients with a substantial increase in metastatic risk. The observed rates of metastasis over 10% and 20% are clinically actionable for nodal staging or postoperative adjuvant radiation, respectively. Combining clinicopathologic risk assessment with individual biologic risk, as provided by the 40-GEP test, improves the accuracy of risk assessment used clinically as the basis of treatment decisions.

Management of Cutaneous Squamous Cell Carcinoma of the Scalp: The Role of Imaging and Therapeutic Approaches

Cutaneous squamous cell carcinoma (cSCC) is the second most common subtype of skin cancer. The scalp is one of the most frequently affected locations and is associated with a higher rate of complications, compared to other locations. In addition, it has a characteristic thickness and anatomical structure that may influence both growth pattern and treatment of primary cSCC; while clinical peripheral margins may be easily achieved during the surgery, vertical excision of the tumor is limited by the skull. Despite having a unique anatomy, current guidelines do not contemplate specific recommendations for scalp cSCC, which leads to inconsistent decision-making in multidisciplinary committees when discussing tumors with high risk factors or with close margins. This article provides specific recommendations for the management of patients with scalp cSCC, based on current evidence, as well as those aspects in which evidence is lacking, pointing out possible future lines of research. Topics addressed include epidemiology, clinical presentation and diagnosis, imaging techniques, surgical and radiation treatments, systemic therapy for advanced cases, and follow-up. The primary focus of this review is on management of primary cSCC of the scalp with localized disease, although where relevant, some points about recurrent cSCCs or advanced disease cases are also discussed.

Real-World Treatment Patterns and Outcomes of Cemiplimab in Patients with Advanced Cutaneous Squamous Cell Carcinoma Treated in US Oncology Practices.

Prior to the Food and Drug Administration approval of cemiplimab in 2018, the median overall survival (OS) for adult patients with advanced CSCC receiving systemic therapy was approximately 8 to 15 months. Limited real-world data are available on cemiplimab for this indication in the US. This retrospective cohort study included US patients with advanced CSCC initiating cemiplimab monotherapy in a real-world database (2018-2021). A clinical trial-like sub-cohort was identified using select criteria. Time to treatment discontinuation (TTD), time to next treatment (TTNT), and OS were estimated using Kaplan-Meier methods. Cox proportional hazard models were used to examine prognostic factors associated with OS in the main cohort. The main cohort included 622 patients (n = 240 in the trial-like cohort). In the main cohort, the median age was 78 years, 77.8% were male, 21.4% were immunocompromised/immunosuppressed, and 63.8% had metastatic CSCC. Median (95% CI) TTD and TTNT were 8.0 (6.6-9.0) months and 16.4 (13.3-21.0) months, respectively, in the main cohort. Median (95% CI) OS was 24.8 (21.8-29.1) months in the main cohort (not reached in the trial-like cohort). In multivariable analyses, age <60 years (hazard ratio [HR], 0.37), Eastern Cooperative Oncology Group performance status <3-4 (HR range, 0.13-0.57), and primary CSCC location in the head and neck only versus extremities only (HR, 0.59) were associated with better OS. Similar OS was observed between patients who had immunosuppressing/immunocompromising conditions and those without. These findings confirm the effectiveness of cemiplimab among a heterogenous, real-world advanced CSCC patient population and substantiate the efficacy of cemiplimab observed in clinical trials.

Prognostic Outcomes of Cutaneous Squamous Cell Carcinoma in Solid Organ Transplant Recipients: A Retrospective Comparative Cohort Study.

Cutaneous squamous cell carcinoma (cSCC) is the second most common cutaneous neoplasm, and its incidence is on the rise. While most cSCCs have an excellent prognosis, certain risk factors, especially immunosuppression, have been associated with higher rates of local recurrence (LR), metastasis, and poor prognosis. This study aims to assess the risk factors for LR and metastasis development in cSCC among solid organ transplant recipients (SOTRs) and compare these rates with those in immunocompetent patients. A retrospective observational study included cSCC cases from the University Hospital Reina Sofía in Córdoba, Spain, between 2002 and 2019. Demographic, clinical, and histopathological data were collected. Local recurrence and metastasis rates were analyzed, along with progression-free survival. Univariate analyses were performed to identify prognostic factors in SOTRs. Among 849 cSCC cases, we found higher rates of local recurrence and metastasis in tumors developed by SOTRs compared to those in immunocompetent individuals. However, no significant differences in local recurrence, metastasis, or progression-free survival were observed between the two groups. Risk factors for adverse outcomes in SOTRs included tumor size > 2 cm, depth > 4 mm, and a higher Clark level. A total of 34.4% of SOTRs developed a second primary cSCC during the follow-up. In our study, cSCCs in SOTRs did not exhibit statistically significant differences in the rates of adverse outcomes compared to immunocompetent patients. The prognosis of cSCCs in SOTRs may be more related to other tumor-dependent risk factors than to the immunosuppression status itself. Future studies are needed to refine risk stratification and follow-up protocols to ensure the optimal management of high-risk cSCC cases, particularly among immunosuppressed patients.

Risk-Stratification using the 40-Gene Expression Profile (40-GEP) Test Identifies Patients with Node Negative Cutaneous Squamous Cell Carcinoma (cSCC) at Higher Risk of Metastasis Who May Benefit from Adjuvant Radiation Therapy (ART)

ART is a standard treatment used to reduce the risk of metastasis and recurrence in moderate‒to‒high-risk cSCC patients. Indications for ART have been largely based on pathologic risk factors and informed by staging systems, and while radiation oncologists generally designate a >10% risk threshold for usage of ART, there is no consensus on which groups of tumors may benefit from ART. The 40-GEP test has been independently validated to predict a cSCC patient's risk for regional/distant metastasis in patients with one or more high-risk clinicopathologic factors and reports three biologic risk groups: Class 1 (low, ∼7%), Class 2A (moderate, 20-25%), and Class 2B (high risk, >50%) for metastasis. This study aims to evaluate whether a biomarker informed risk stratification approach using a 40-GEP result could refine the ability to select patients with node negative cSCC at higher risk of metastasis who are most likely to benefit from ART. In this retrospective study, all patients had primary cSCC tissue with verified clinicopathologic information of tumors with one or more high-risk factors, met clinical testing criteria, were comprehensively staged, and had outcomes data (n = 954). Patients with node positive disease, or those with nodal failure within 3 months of diagnosis were excluded (n = 19). From the n = 935, an intermediate risk population wherein ART is often considered was defined as Brigham and Women's Hospital (BWH) ≥T2a (n = 489). Kaplan-Meier survival analysis and log-rank test were used to assess metastasis free survival (MFS). Univariate Cox regression compared metastasis rates between 40-GEP results. The 3-year MFS rate for this eligible for ART cohort was 82.4% The 40-GEP demonstrated statistically significant risk stratification with MFS rates of 92.4%, 76.1% and 59.4% for Class 1, Class 2A and Class 2B, respectively (p<0.0001). Cox regression was significant for Class 2A and 2B compared to Class 1, with a 3.2-fold and 6.4-fold increase in metastasis, respectively (p<0.0001). 64% (59/92) of all metastases received a Class 2A result, and 44% (14/32) of Class 2B patients metastasized. 46% (223/489) of the cohort received a Class 1 result. Of patients staged BWH T1 (n = 446), those with a Class 2A and 2B had an 88.7% and 66.7% MFS rate, respectively. Within this eligible for ART population, patients with Class 2A or 2B 40-GEP results have inferior rates of MFS, while Class 1 patients have <10% risk of metastasis. Nearly half of this population received a 40-GEP Class 1 result and could be considered for treatment de-intensification trials. Conversely, patients with low-risk BWH T1 stage, who are traditionally not considered for ART, that received a Class 2A or 2B (>10% risk of metastasis) could be considered for adjuvant therapy.

Driver gene combinations dictate cutaneous squamous cell carcinoma disease continuum progression

The molecular basis of disease progression from UV-induced precancerous actinic keratosis (AK) to malignant invasive cutaneous squamous cell carcinoma (cSCC) and potentially lethal metastatic disease remains unclear. DNA sequencing studies have revealed a massive mutational burden but have yet to illuminate mechanisms of disease progression. Here we perform RNAseq transcriptomic profiling of 110 patient samples representing normal sun-exposed skin, AK, primary and metastatic cSCC and reveal a disease continuum from a differentiated to a progenitor-like state. This is accompanied by the orchestrated suppression of master regulators of epidermal differentiation, dynamic modulation of the epidermal differentiation complex, remodelling of the immune landscape and an increase in the preponderance of tumour specific keratinocytes. Comparative systems analysis of human cSCC coupled with the generation of genetically engineered murine models reveal that combinatorial sequential inactivation of the tumour suppressor genes Tgfbr2, Trp53, and Notch1 coupled with activation of Ras signalling progressively drives cSCC progression along a differentiated to progenitor axis. Taken together we provide a comprehensive map of the cSCC disease continuum and reveal potentially actionable events that promote and accompany disease progression.

Personalised decision making to predict absolute metastatic risk in cutaneous squamous cell carcinoma: development and validation of a clinico-pathological model

Cutaneous squamous cell carcinoma (cSCC) is a common skin cancer, affecting more than 2 million people worldwide yearly and metastasising in 2-5% of patients. However, current clinical staging systems do not provide estimates of absolute metastatic risk, hence missing the opportunity for more personalised treatment advice. We aimed to develop a clinico-pathological model that predicts the probability of metastasis in patients with cSCC. Nationwide cohorts from (1) all patients with a first primary cSCC in The Netherlands in 2007-2008 and (2) all patients with a cSCC in 2013-2015 in England were used to derive nested case-control cohorts. Pathology records of primary cSCCs that originated a loco-regional or distant metastasis were identified, and these cSCCs were matched to primary cSCCs of controls without metastasis (1:1 ratio). The model was developed on the Dutch cohort (n=390) using a weighted Cox regression model with backward selection and validated on the English cohort (n=696). Model performance was assessed using weighted versions of the C-index, calibration metrics, and decision curve analysis; and compared to the Brigham and Women's Hospital (BWH) and the American Joint Committee on Cancer (AJCC) staging systems. Members of the multidisciplinary Skin Cancer Outcomes (SCOUT) consortium were surveyed to interpret metastatic risk cutoffs in a clinical context. Eight out of eleven clinico-pathological variables were selected. The model showed good discriminative ability, with an optimism-corrected C-index of 0.80 (95% Confidence interval (CI) 0.75-0.85) in the development cohort and a C-index of 0.84 (95% CI 0.81-0.87) in the validation cohort. Model predictions were well-calibrated: the calibration slope was 0.96 (95% CI 0.76-1.16) in the validation cohort. Decision curve analysis showed improved net benefit compared to current staging systems, particularly for thresholds relevant for decisions on follow-up and adjuvant treatment. The model is available as an online web-based calculator (https://emc-dermatology.shinyapps.io/cscc-abs-met-risk/). This validated model assigns personalised metastatic risk predictions to patients with cSCC, using routinely reported histological and patient-specific risk factors. The model can empower clinicians and healthcare systems in identifying patients with high-risk cSCC and offering personalised care/treatment and follow-up. Use of the model for clinical decision-making in different patient populations must be further investigated. PPP Allowance made available by Health-Holland, Top Sector Life Sciences & Health, to stimulate public-private partnerships.

Transcriptomic analysis of cutaneous squamous cell carcinoma reveals a multigene prognostic signature associated with metastasis

Metastasis of cutaneous squamous cell carcinoma (cSCC) is uncommon. Current staging methods are reported to have sub-optimal performances in metastasis prediction. Accurate identification of patients with tumors at high risk of metastasis would have a significant impact on management. To develop a robust and validated gene expression profile signature for predicting primary cSCC metastatic risk using an unbiased whole transcriptome discovery-driven approach. Archival formalin-fixed paraffin-embedded primary cSCC with perilesional normal tissue from 237 immunocompetent patients (151 nonmetastasizing and 86 metastasizing) were collected retrospectively from four centers. TempO-seq was used to probe the whole transcriptome and machine learning algorithms were applied to derive predictive signatures, with a 3:1 split for training and testing datasets. A 20-gene prognostic model was developed and validated, with an accuracy of 86.0%, sensitivity of 85.7%, specificity of 86.1%, and positive predictive value of 78.3% in the testing set, providing more stable, accurate prediction than pathological staging systems. A linear predictor was also developed, significantly correlating with metastatic risk. This was a retrospective 4-center study and larger prospective multicenter studies are now required. The 20-gene signature prediction is accurate, with the potential to be incorporated into clinical workflows for cSCC.

European consensus-based interdisciplinary guideline for invasive cutaneous squamous cell carcinoma: Part 2. Treatment–Update 2023

In order to update recommendations on treatment, supportive care, education, and follow-up of patients with invasive cutaneous squamous cell carcinoma (cSCC), a multidisciplinary panel of experts from the European Association of Dermato-Oncology (EADO), the European Dermatology Forum (EDF), the European Society for Radiotherapy and Oncology (ESTRO), the European Union of Medical Specialists (UEMS), the European Academy of Dermatology and Venereology (EADV), and the European Organisation of Research and Treatment of Cancer (EORTC) was formed. Recommendations were based on an evidence-based literature review, guidelines, and expert consensus. Treatment recommendations are presented for common primary cSCC (low risk, high risk), locally advanced cSCC, regional metastatic cSCC (operable or inoperable), and distant metastatic cSCC. For common primary cSCC, the first-line treatment is surgical excision with postoperative margin assessment or micrographically controlled surgery. Achieving clear surgical margins is the most important treatment consideration for patients with cSCCs amenable to surgery. Regarding adjuvant radiotherapy for patients with high-risk localised cSCC with clear surgical margins, current evidence has not shown significant benefit for those with at least one high-risk factor. Radiotherapy should be considered as the primary treatment for non-surgical candidates/tumours. For cSCC with cytologically or histologically confirmed regional nodal metastasis, lymph node dissection is recommended. For patients with metastatic or locally advanced cSCC who are not candidates for curative surgery or radiotherapy, anti-PD-1 agents are the first-line systemic treatment, with cemiplimab being the first approved systemic agent for advanced cSCC by the Food and Drugs Administration/European Medicines Agency. Second-line systemic treatments for advanced cSCC, include epidermal growth factor receptor inhibitors (cetuximab) combined with chemotherapy or radiotherapy. Multidisciplinary board decisions are mandatory for all patients with advanced cSCC, considering the risks of toxicity, the age and frailty of patients, and co-morbidities, including immunosuppression. Patients should be engaged in informed, shared decision-making on management and be provided with the best supportive care to improve symptom management and quality of life. The frequency of follow-up visits and investigations for subsequent new cSCC depends on underlying risk characteristics.

P26 Risk factors for complications in cutaneous squamous cell carcinoma and proposal of a simplified follow-up strategy, making use of available healthcare resources

Abstract Cutaneous squamous cell carcinoma (cSCC) is a common malignancy, generally cured by surgical excision. A minority of such tumours recur, metastasize or even cause death. The 2021 British Association of Dermatologists (BAD) guidelines outline high-risk features that are used to guide the follow-up of patients after a primary cSCC. The aims of this study were to identify histological features of cSCC that are significantly associated with the development of complications, and to compare the sensitivity, specificity and number of appointments required to follow-up patients using different follow-up criteria. In total, 556 cases of primary invasive cSCCs excised between January 2014 and March 2018 were included and followed-up for a minimum of 4 years. Five per cent (n = 26) of tumours were associated with at least one cSCC-related complication, namely, local recurrence 3.8% (n = 21), nodal metastasis 1.6% (n = 9), distant metastasis 0.4% (n = 2) and disease-specific death 0.7% (n = 4). The initial diagnosed complication was local recurrence in 20 of 26 cases. Only two patients had the first complication beyond 2 years of the original cSCC. Factors that were statistically significantly associated with the development of a complication (P &amp;lt; 0.001) are larger diameter, larger depth of invasion, tumour invasion beyond the dermis, perineural invasion (irrespective of size), lymphovascular invasion, gross bone erosion and deep margin &amp;lt; 1 mm. Differentiation and peripheral margin were not significantly associated with complications. Therefore, it may be proposed that patients with tumours with at least one of the following risk factors be followed-up: diameter &amp;gt; 40 mm; depth of invasion &amp;gt; 6 mm; invasion beyond the dermis; presence of perineural or lymphovascular invasion or deep margin &amp;lt; 1 mm. In adopting these criteria, 739 appointments in 156 patients would be required over 2 years (4 monthly during the first year and 6 monthly during the second year). Twenty of the patients who developed a complication would be followed-up (sensitivity 76.9%, specificity 69.4%). Using the BAD guidelines, up to four more patients with complications would have been followed-up (sensitivity 92.3%, specificity 44.7%), but this would have required a total of 1556 follow-up visits in 242 patients. Given the limited resources, patient burden and the fact that local recurrence is most often the first complication, it may be reasonable to offer routine follow-up only to patients who have at least one of the proposed criteria. Coupled with patient education, this strategy could greatly reduce the number of follow-up visits required. Such a strategy may allow resources to be utilized to reduce waiting times, particularly for 2-week wait referrals, decreasing the delay in new skin cancer diagnoses.

DS08 (P26) Risk factors for complications in cutaneous squamous cell carcinoma and proposal of a simplified follow-up strategy: making use of available healthcare resources

Abstract Cutaneous squamous cell carcinoma (cSCC) is a common malignancy, generally cured by surgical excision. A minority of such tumours recur, metastasize or even cause death. The 2021 British Association of Dermatologists (BAD) guidelines outline high-risk features that are used to guide the follow-up of patients after a primary cSCC. The aims of this study were to identify histological features of cSCC that are significantly associated with the development of complications and to compare the sensitivity, specificity and number of appointments required to follow-up patients using different follow-up criteria. In total, 556 cases of primary invasive cSCCs excised between January 2014 and March 2018 were included and followed-up for a minimum of 4 years. Five per cent (n = 26) of tumours were associated with at least one cSCC-related complication, namely, local recurrence 3.8% (n = 21), nodal metastasis 1.6% (n = 9), distant metastasis 0.4% (n = 2) and disease-specific death 0.7% (n = 4). The initial diagnosed complication was local recurrence in 20 of 26 cases. Only two patients had the first complication beyond 2 years of the original cSCC. Factors that were statistically significantly associated with the development of a complication (P &amp;lt; 0.001) were larger diameter, larger depth of invasion, tumour invasion beyond the dermis, perineural invasion (irrespective of size), lymphovascular invasion, gross bone erosion and deep margin &amp;lt; 1 mm. Differentiation and peripheral margin were not significantly associated with complications. Therefore, it may be proposed that patients with tumours with at least one of the following risk factors be followed-up: diameter &amp;gt; 40 mm, depth of invasion &amp;gt; 6 mm, invasion beyond the dermis, presence of perineural or lymphovascular invasion or deep margin &amp;lt; 1 mm. In adopting these criteria, 739 appointments in 156 patients would be required over 2 years (4-monthly during the first year and 6-monthly during the second year). Twenty of the patients who developed a complication would be followed-up (sensitivity 76.9%, specificity 69.4%). Using the BAD guidelines, up to four more patients with complications would have been followed-up (sensitivity 92.3%, specificity 44.7%), but this would have required a total of 1556 follow-up visits in 242 patients. Given the limited resources, patient burden and the fact that local recurrence is most often the first complication, it may be reasonable to offer routine follow-up only to patients who have at least one of the proposed criteria. Coupled with patient education, this strategy could greatly reduce the number of follow-up visits required. Such a strategy may allow resources to be utilized to reduce waiting times, particularly for 2-week wait referrals, decreasing the delay in new skin cancer diagnoses.

Performance analysis of four machine learning algorithms for the accurate prediction of metastatic disease in cutaneous squamous cell carcinoma.

e13579 Background: Cutaneous squamous cell carcinoma (cSCC) are the most common form of metastasising skin cancer. Whilst rates of metastatic cSCC are low, they account for a significant proportion of skin cancer related morbidity and mortality, particularly within elderly cohorts, which poses a significant burden to healthcare services. Established cSCC tumour staging systems perform poorly at predicting metastatic risk. Additionally, we lack clinically validated prognostic biomarkers – highlighting the unmet need for novel risk stratification tools to guide clinical practice and improve outcomes for patients with advanced disease. We aimed to train four recognised machine learning (ML) algorithms on a large clinic-pathological dataset of primary cSCC, with the objective of optimising an ML strategy and developing a reliable and clinically useful risk stratification tool capable of accurately predicting metastatic events following primary cSCC. Methods: A dataset of primary cSCC registrations was derived from the Northern Cancer Registry, UK. This identified 7003 histologically confirmed primary cSCC registered between 2010–2020; providing a minimum of 2 years clinical follow-up. We conducted a retrospective analysis of standardised pathology datasets, recording clinical-pathological features. Primary outcome measure was regional and/or distant metastasis. Four machine learning algorithms, were trained based on these features, including: a Logistic Regression Trainer, a Decision Tree Classifier, a Random Forest Classifier and a fully connected artificial neural network (ANN). The algorithms were optimised on training data using five-fold cross validation. Subgroup analysis was performed using mean Shapley additive explanations (SHAP). Results: Accuracy scoring identified the ANN as the optimal predictor of metastasis (0.94), followed by: Logistic Regression Trainer (0.82), Random Forest Classifier (0.80), and Decision Tree Classifier (0.71). Preliminary subgroup analysis identified immunosuppression as most sensitive risk factor for developing metastatic disease (SHAP = 0.122). Conclusions: Significant heterogeneity in current morbidity and mortality data has limited the capacity of traditional statistical models and tumour staging systems to identify very high-risk cSSC. Our findings demonstrate that ML algorithms can accurately predict metastatic events in cSSC populations. Further development of a model user-interface is necessary to support the development of a useful risk stratification tool to guide clinical practice.

Abstract 2101: Cancer-associated fibroblast activation predicts progression, metastasis, and prognosis of cutaneous squamous cell carcinoma

Abstract Purpose: Cutaneous squamous cell carcinoma (cSCC) is the most common metastatic skin cancer and the metastatic disease is associated with poor prognosis. At present, no established biomarkers are available for predicting the metastasis risk and prognosis of patients with cSCC. Cancer-associated fibroblasts (CAFs) promote the progression of cancer, but their role in cSCC is largely unknown. Here, we have examined the potential of CAF markers in the assessment of metastasis risk and prognosis of primary cSCC. Experimental Design: We utilized multiplexed fluorescence immunohistochemistry for profiling of CAF landscape in metastatic and non-metastatic primary human cSCCs, the actual metastases, and in premalignant and benign epidermal lesions. Quantitative high-resolution image analysis was performed with two separate panels of antibodies for CAF markers and the results were correlated with clinical and histopathological parameters including disease-specific mortality. Results: Increased stromal expression of fibroblast activation protein (FAP), α-smooth muscle actin, and secreted protein acidic and rich in cysteine (SPARC) were associated with progression of premalignant epidermal lesions to invasive cSCC. Furthermore, elevation of FAP and platelet-derived growth factor receptor-β (PDGFRβ) expression was associated with the risk of metastasis of primary cSCCs. High expression of PDGFRβ and periostin correlated with poor prognosis. Furthermore, a multi-marker defined CAF subset, PDGFRα-/PDGFRβ+/FAP+, was associated with invasion and metastasis and independently predicted poor disease-specific survival. Conclusions: These results show, that fibroblast activation evolves during the progression of cSCC. High PDGFRβ expression alone and multimarker combination PDGFRα-/PDGFRβ+/FAP+ are potential biomarkers for the progression of primary cSCC to metastatic stage, and for poor prognosis. Citation Format: Jaakko S. Knuutila, Pilvi Riihilä, Liisa Nissinen, Roosa Kallionpää, Teijo Pellinen, Veli-Matti Kähäri. Cancer-associated fibroblast activation predicts progression, metastasis, and prognosis of cutaneous squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2101.