Novel chemokine biomarkers in melanoma†.

The melanoma tumor microenvironment is a complex milieu of cancer, inflammatory, and stromal cells. In this context, chemokines play a pivotal role in recruiting inflammatory cells and influence the tumor, exerting both pro-tumorigenic and anti-tumoral roles. Interactions between these cells is what ultimately hold together and transform the tumor into an efficient machine. A recent study found that chemokines CCL8, CCL15, and CCL20 were upregulated in melanoma cells when co-cultured with macrophages and were associated with poor survival rates. CCL8 and CCL15 also stimulated melanoma cell growth, invasion, and metastasis, and were highly expressed in tumors prone to metastasize, suggesting these chemokines are attractive and independent biomarkers. Understanding the intricated interactions within the tumor microenvironment could lead to prognostic biomarkers and to the development of new therapeutic strategies for melanoma. © 2024 The Author(s). The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Chondroitin sulfate proteoglycan 4 (CSPG4) increases invasion of recessive dystrophic epidermolysis bullosa-associated cutaneous squamous cell carcinoma by modifying TGFß signaling.

BACKGROUND: Recessive dystrophic epidermolysis bullosa (RDEB) is a rare genetic skin-blistering disorder often progressing to metastatic cutaneous squamous cell carcinoma (cSCC) at chronic wound sites. Chondroitin sulfate proteoglycan 4 (CSPG4) is a cell-surface proteoglycan that is an oncoantigen in multiple malignancies, where it modulates oncogenic signaling, drives epithelial-to-mesenchymal transition (EMT), and enables cell motility. OBJECTIVES: To evaluate CSPG4 expression and function in RDEB-cSCC. METHODS: RDEB-cSCC cell lines were used to assess CSPG4-dependent changes in invasive potential, TGFß1-stimulated signal activation, and clinically relevant cytopathology metrics in an in vitro full-thickness tumor model. CSPG4 expression in RDEB-cSCC and non-RDEB cSCC tumors was analyzed via immunohistochemistry and single-cell RNA sequencing (scRNA-seq), respectively. RESULTS: Inhibiting CSPG4 expression reduced invasive potential in multiple RDEB-cSCC cell lines and altered membrane-proximal TGFß signal activation through changes in SMAD3 phosphorylation. CSPG4 expression was uniformly localized to basal-layer keratinocytes in fibrotic RDEB skin and tumor cells at the tumor/stroma interface at the invasive front in RDEB-cSCC tumors in vivo. Analysis of published scRNA-seq data revealed that CSPG4 expression was correlated with an enhanced EMT transcriptomic signature in cells at the tumor/stroma interface of non-RDEB cSCC tumors. Cytopathological metrics, like nucleus:cell area ratio, were influenced by CSPG4 expression in in vitro tumor models. CONCLUSIONS: We determined that CSPG4 expression in RDEB-cSCC cell lines enhanced invasive potential. Mechanistically, CSPG4 was found to enhance membrane-proximal TGFß-stimulated signaling through SMAD3, which is a key mediator of EMT in RDEB-cSCC. The implication of these studies is that CSPG4 may represent a therapeutic target that can be leveraged for clinical management in patients with RDEB-cSCC.

Metastatic Tonsil Squamous Cell Carcinoma: An Important Consideration in the Differential Diagnosis of Malignant Basaloid Neoplasms in the Skin: Case Report and Review of the Literature.

ABSTRACT: Malignant basaloid neoplasms of the skin are frequent, and their accurate diagnosis holds paramount importance for treatment and prognosis. However, these neoplasms can present diagnostic challenges because of their extensive differential diagnosis, which encompasses cutaneous metastasis among many other possibilities. We present a case of a 74-year-old man with a history of p16-positive palatine tonsil squamous cell carcinoma (SCC) treated with surgery and adjuvant radiation with no prior evidence of recurrence who presented to the dermatologist with 2 chin papules. The initial histopathologic evaluation of these lesions showed poorly differentiated malignant basaloid neoplasms. Subsequently, these biopsies were compared with the previous biopsies from his tonsil and lymph node, which showed similar findings including positive p16 staining and positive molecular testing for human papillomavirus-16, confirming the diagnosis of cutaneous metastasis from his previously diagnosed human papillomavirus-related tonsil SCC. Additional imaging studies found metastases to internal organs including the brain, and he was started on chemotherapy, immunotherapy, and radiation therapy. Cutaneous metastases from tonsil SCC are exceedingly rare, and only 5 cases have been described. Furthermore, this is the first case confirming the presence of high-risk human papillomavirus by molecular studies within the cutaneous metastases. The presented case underscores the importance of recognizing this unusual manifestation of tonsil SCC metastatic to the skin along with a good clinical patient history, ensuring accurate and prompt diagnosis and treatment of this condition.

Apoptosis Pathway-Associated Proteins Are Frequently Expressed in Melanoma: A Study of 32 Cases With Focus on Acral Lentiginous Melanoma.

ABSTRACT: Acral lentiginous melanoma (ALM) is an aggressive type of cutaneous melanoma (CM) that arises on palms, soles, and nail units. ALM is rare in White population, but it is relatively more frequent in dark-skinned populations. There is an unmet need to develop new personalized and more effective treatments strategies for ALM. Increased expression of antiapoptotic proteins (ie, BCL2, MCL1) has been shown to contribute to tumorigenesis and therapeutic resistance in multiple tumor types and has been observed in a subset of ALM and mucosal melanoma cell lines in vivo and in vitro. However, little is known about their expression and clinical significance in patients with ALM. Thus, we assessed protein expression of BCL2, MCL1, BIM, and BRAF V600E by immunohistochemistry in 32 melanoma samples from White and Hispanic populations, including ALM and non-ALM (NALM). BCL2, MCL1, and BIM were expressed in both ALM and NALM tumors, and no significant differences in expression of any of these proteins were detected between the groups, in our relatively small cohort. There were no significant associations between protein expression and BRAF V600E status, overall survival, or ethnicity. In summary, ALM and NALM demonstrate frequent expressions of apoptosis-related proteins BCL2, MCL1, and BIM. Our findings suggest that patients with melanoma, including ALM, may be potential candidates for apoptosis-directed therapies.

Sentinel lymph node assessment in melanoma: current state and future directions.

Assessment of sentinel lymph node status is an important step in the evaluation of patients with melanoma for both prognosis and therapeutic management. Pathologists have an important role in this evaluation. The methodologies have varied over time, from the evaluation of dimensions of metastatic burden to determination of the location of the tumour deposits within the lymph node to precise cell counting. However, no single method of sentinel lymph node tumour burden measurement can currently be used as a sole independent predictor of prognosis. The management approach to sentinel lymph node-positive patients has also evolved over time, with a more conservative approach recently recognised for selected cases. This review gives an overview of past and current status in the field with a glimpse into future directions based on prior experiences and clinical trials.

Primary mucosal melanomas of the head and neck are characterised by overexpression of the DNA mutating enzyme APOBEC3B.

AIMS: Primary head/neck mucosal melanomas (MMs) are rare and exhibit aggressive biologic behaviour and elevated mutational loads. The molecular mechanisms responsible for high genomic instability observed in head/neck MMs remain elusive. The DNA cytosine deaminase APOBEC3B (A3B) constitutes a major endogenous source of mutation in human cancer. A3B-related mutations are identified through C-to-T/-G base substitutions in 5'-TCA/T motifs. Herein, we present immunohistochemical and genomic data supportive of a role for A3B in head/neck MMs. METHODS AND RESULTS: A3B protein levels were assessed in oral (n = 13) and sinonasal (n = 13) melanomas, and oral melanocytic nevi (n = 13) by immunohistochemistry using a custom rabbit α-A3B mAb (5210-87-13). Heterogeneous, selective-to-diffuse, nuclear only, A3B immunopositivity was observed in 12 of 13 (92.3%) oral melanomas (H-score range = 9-72, median = 40) and 8 of 13 (62%) sinonasal melanomas (H-score range = 1-110, median = 24). Two cases negative for A3B showed prominent cytoplasmic staining consistent with A3G. A3B protein levels were significantly higher in oral and sinonasal MMs than intraoral melanocytic nevi (P < 0.0001 and P = 0.0022, respectively), which were A3B-negative (H-score range = 1-8, median = 4). A3B levels, however, did not differ significantly between oral and sinonasal tumours (P > 0.99). NGS performed in 10 sinonasal MMs revealed missense NRAS mutations in 50% of the studied cases and one each KIT and HRAS mutations. Publicly available whole-genome sequencing (WGS) data disclosed that the number of C-to-T mutations and APOBEC3 enrichment score were markedly elevated in head/neck MMs (n = 2). CONCLUSION: The above data strongly indicate a possible role for the mutagenic enzyme A3B in head/neck melanomagenesis, but not benign melanocytic neoplasms.

Erythema nodosum after azacitidine in a patient with acute myeloid leukemia.

INTRODUCTION: Nodular skin lesions in patients with acute myeloid leukemia (AML) raise clinical suspicion for leukemia cutis versus fungal infections. Here, we report a rare case of treatment-related erythema nodosum (EN) in a patient with AML. CASE REPORT: Approximately 5 weeks after the initiation of sorafenib and one week after azacitidine initiation, a 32-year-old man with primary refractory AML presented with several painful red nodules on the lower extremities. Histological examination established a diagnosis of EN. MANAGEMENT AND OUTCOME: Treatment with topical and oral steroids led to complete resolution of the nodules. However, once the dose of steroids was reduced, the lesions rapidly recurred. Higher dose steroids were reinitiated, again with a resolution of the nodules, confirming steroid responsiveness of the underlying process. DISCUSSION: Given the onset of lesions one week after the initiation of azacitidine and 5 weeks after the initiation of sorafenib, azacitidine was considered the more likely culprit. Only 2 cases of EN-like eruption after azacitidine and 1 case after sorafenib have been reported. Although fungal infections and leukemia cutis are the top differentials considered for skin nodules in a patient with AML, EN should be considered as an alternative diagnosis. Correct diagnosis is critical because it will guide treatment.

Sentinel lymph node tumour burden using digital cell count estimation predicts outcomes in melanoma.

BACKGROUND AND AIMS: Cutaneous melanoma often metastasises in primis to sentinel lymph nodes (SLNs). Currently, there is no standardized method of characterizing the micrometastatic tumour burden in SLN biopsies for melanoma. Different criteria have been developed to evaluate SLN biopsies, yet none consider the number of cells identified. Here, we used software analysis to digitally quantify metastatic tumour burden within SLNs and correlated these data with clinicopathological and prognostic information. METHODS AND RESULTS: We identified 246 cases of SLN biopsies, including 63 positive (26%) and183 (74%) negative for metastatic melanoma. Digital cell counting was performed within the greatest metastatic focus and the entire metastatic tumour burden within the same SLN. Increasing cell count in the largest metastatic deposit correlated with the previously described Rotterdam [Spearman's r = 0.91; 95% confidence interval (CI) = 0.84, 0.94], Starz (Spearman's r = 0.78; 95% CI = 0.68, 0.87) and Dewar criteria (P < 0.01), validating our method of using cell count to define SLN tumour burden. Additionally, increasing cell count was associated with decreased metastasis-free survival (hazard ratio = 2.29; 95% CI = 1.22, 4.31). CONCLUSION: These data support the use of computerized cell count analysis for prognostication of outcomes in patients undergoing SLN biopsy.

The mixed spongiotic and interface reaction pattern: A study of clinical and histopathologic findings.

BACKGROUND: Categorization of biopsy specimens into inflammatory reaction patterns is central to dermatopathologic assessment. Mixed inflammatory patterns are poorly characterized and may represent clinicopathologic challenges. The purpose of this study was to identify clinical and histopathologic findings associated with the mixed spongiotic-interface dermatitis (SID) histopathologic pattern. METHODS: Fifty-one institutional biopsy specimens of SID were identified over a 2-year period by retrospective natural language search. Histopathologic and clinical features were identified. RESULTS: The most common histopathologic features associated with SID were mild spongiosis (51%), focal vacuolar interface change (72%), lymphocytic exocytosis (92%), and superficial-dermal lymphocytic infiltrate (94%) with variable eosinophils (61%). Clinically, 80% of subjects presented with a symmetric morbilliform eruption. Polypharmacy (94%), immunosuppression (47%), and history of malignancy (47%) were common. The most common diagnoses were drug reaction (37%), possible drug reaction (12%), and viral exanthem (12%). Drug reaction with eosinophilia and systemic symptoms represented 25% of all confirmed cutaneous adverse drug reactions (CADR). Average time from drug initiation to symptom initiation was 20 days (SD: 22.3, range: 0-90); median disease duration was 25.5 days. Spongiotic vesicles and Langerhans cells were less common in patients with a strong clinicopathologic diagnosis of drug reaction compared to non-drug eruptions (p = 0.04). CONCLUSIONS: The mixed SID pattern is commonly encountered in CADR but may represent a more subacute course, implying consideration for inciting medication(s) started before the typical 7- to 14-day window.

Diagnostic utility of PRAME in distinguishing proliferative nodules from melanoma in giant congenital melanocytic nevi.

We describe a case of a melanocytic proliferation arising in a giant congenital melanocytic nevus (CMN) and outline the potential utility of an immunohistochemical study with PReferentially expressed Antigen in MElanoma (PRAME) in distinguishing benign proliferative nodules (PN) from melanoma in this context. A 15-day-old girl presented with a fibrotic nodule clinically suspicious for melanoma within a giant CMN. Histopathological examination showed a predominantly intradermal melanocytic nevus with congenital features intermixing with an ill-defined proliferation of larger melanocytes demonstrating mild-to-moderate cytologic atypia and increased mitotic activity. Anti-PRAME was diffusely positive within the congenital nevus while negative within the larger proliferating cells. Chromosomal microarray analysis revealed whole chromosomal gains and losses only, consistent with a PN arising in a giant CMN. To our knowledge, PRAME expression in giant CMN, PN, and pediatric melanomas has not been previously described. Based on our experience with this case, we propose that differential patterns of PRAME expression may be present in these three lesions, allowing PRAME immunohistochemistry to potentially serve as a helpful adjunct diagnostic tool for laboratories that do not readily have access to molecular testing in rendering a diagnosis for atypical melanocytic proliferations arising in giant CMN.

Cutaneous adnexal carcinosarcoma: Immunohistochemical and molecular evidence of epithelial mesenchymal transition.

Cutaneous carcinosarcomas are rare biphenotypic tumors that simultaneously show epithelial and mesenchymal differentiation. The most common carcinomatous components in skin carcinosarcomas are basal cell carcinoma and squamous cell carcinoma; adnexal carcinomas are rarely encountered. We report a case of an adnexal carcinoma with ductal and squamous differentiation and spindle cell component, which is interpreted as carcinosarcoma. Loss of immunohistochemical expression of E-cadherin and ß-catenin detected in the sarcomatous component suggested epithelial mesenchymal transition (EMT). RNA sequencing analysis identified several gene mutations and alterations such as translocations and upregulations/downregulations, either shared by the two components of the tumor or differentially present in the carcinoma or the sarcoma parts. Thus, mutations in genes, such as TP53, were found in both components of the tumor while mutations in PDGFRA and RB1 (a pathogenic missense mutation) were exclusively present in the sarcomatous areas, further supporting EMT. EMT is a dynamic process by which tumors acquire mesenchymal phenotype while simultaneously losing epithelial properties. Although the pathways involved in EMT have been extensively studied, this phenomenon still needs to be investigated in cutaneous tumors of adnexal origin for a better understanding of their pathogenesis. These molecular changes may represent promising targets for personalized therapies.

Delayed-onset cutaneous eruption associated with lenalidomide in setting of multiple myeloma.

Lenalidomide (LEN) is increasingly being used for the treatment of multiple myeloma (MM). Adverse cutaneous reactions to LEN are common and present almost exclusively within one month of initiating therapy. We report a case of delayed-onset LEN-associated eruption presenting over three years after starting treatment. Histopathologic findings are also described, which are infrequently reported for LEN-associated eruptions. Our case serves as a reminder that proper recognition and management of LEN-associated eruption is important in the treatment of MM. Dermatologists should be aware of the potential for delayed presentations of adverse cutaneous reactions to LEN, even years after initiation.

The Role of MET in Melanoma and Melanocytic Lesions.

Melanoma is the leading cause of death due to cutaneous malignancy and its incidence is on the rise. Several signaling pathways, including receptor tyrosine kinases, have been recognized to have an etiopathogenetic role in the development and progression of precursor melanocytic lesions and malignant melanoma. Among those, the hepatocyte growth factor/MET (HGF/MET) axis is emerging as a critical player not only in the tumor itself but also in the immune microenvironment in which the tumor grows and advances in its development. Moreover, the activation of this pathway has emerged as a paradigm of tumor resistance to modern targeted therapies, and the assessment of its expression in patients' samples may be a valuable biomarker of tumor progression and response to targeted therapy. Here we summarize our current understanding of this important receptor tyrosine kinase in normal melanocyte proliferation/motility, in tumor progression and metastasis, its genetic alterations in certain subtype of melanocytic lesions, and how its pathway has been explored for the development of selective inhibitors.

Analyses of molecular and histopathologic features and expression of PRAME by immunohistochemistry in mucosal melanomas.

Mucosal melanomas are rare, and less is known about the biomarkers of this subtype in comparison to cutaneous or uveal melanomas. Preferentially expressed antigen in melanoma (PRAME) has been studied as a tool for prognostication of uveal melanomas, and immunotherapy against PRAME-expressing tumor cells has already shown promise. Our goal was to retrospectively analyze 29 cases of mucosal melanomas at our institution to determine if any molecular and histopathologic prognosticators could be identified, as well as to study PRAME expression and its association with prognosis. We found that the majority of mucosal melanomas expressed PRAME and a high PRAME expression score predicted a poor prognosis. There was no association between prognosis and the histomorphologic features analyzed, such as presence of spindle cell or epithelioid predominance. BRAF mutations were absent in 16 of 16 cases tested. Pathogenic NRAS mutations were detected in 3 of 11 cases tested and were associated with shorter overall survival compared to those without NRAS alterations, but the presence of NRAS mutations did not correlate with PRAME expression. In conclusion, an increase in PRAME expression and the presence of a pathogenic NRAS were both associated with a worse prognosis in mucosal melanomas.

Diagnosis of epidermodysplasia verruciformis: Two cases highlighting the role of direct HPV L1 gene sequencing.

Epidermodysplasia verruciformis (EV) is a rare skin disease characterized by the development of multiple flat warts with the potential for malignant transformation. Patients are susceptible to human papillomavirus (HPV) infection that develops in a background of either a genetic or acquired immunodeficiency predisposing patients to infection with specific HPV types that are ubiquitous but generally non-pathogenic in healthy individuals. There is no standard clinical methodology for determining the causative HPV from patients with suspected EV. Here, we report the diagnostic workup of two EV cases and describe the use of L1 gene Sanger sequencing as a specific method to accurately identify the causative HPV genotype and confirm the diagnosis of EV.

Primary cutaneous diffuse large B-cell lymphoma presenting as chronic non-healing ulcer.

Primary cutaneous diffuse large B-cell lymphoma is an uncommon and aggressive lymphoproliferative disorder with a rapid growth rate and dismal prognosis. We present the case of a 91-year-old female with an unusual manifestation of primary cutaneous diffuse large B-cell lymphoma, mimicking other more prevalent diseases like chronic non-healing venous ulceration. Dermatopathologic evaluation rendered the correct diagnosis. A discussion of this rare presentation is important for clinician consideration to prevent misdiagnosis and prolongation of proper management in patients with chronic non-healing leg ulcers.

Preliminary evaluation of urinary soluble Met as a biomarker for urothelial carcinoma of the bladder.

BACKGROUND: Among genitourinary malignancies, bladder cancer (BCa) ranks second in both prevalence and cause of death. Biomarkers of BCa for diagnosis, prognosis and disease surveillance could potentially help prevent progression, improve survival rates and reduce health care costs. Among several oncogenic signaling pathways implicated in BCa progression is that of hepatocyte growth factor (HGF) and its cell surface receptor, Met, now targeted by 25 experimental anti-cancer agents in human clinical trials. The involvement of this pathway in several cancers is likely to preclude the use of urinary soluble Met (sMet), which has been correlated with malignancy, for initial BCa screening. However, its potential utility as an aid to disease surveillance and to identify patients likely to benefit from HGF/Met-targeted therapies provide the rationale for this preliminary retrospective study comparing sMet levels between benign conditions and primary BCa, and in BCa cases, between different disease stages. METHODS: Normally voided urine samples were collected from patients with BCa (Total: 183; pTa: 55, pTis: 62, pT1: 24, pT2: 42) and without BCa (Total: 83) on tissue-procurement protocols at three institutions and sMet was measured and normalized to urinary creatinine. Normalized sMet values grouped by pathologic stage were compared using non-parametric tests for correlation and significant difference. ROC analyses were used to derive classification models for patients with or without BCa and patients with or without muscle-invasive BCa (MIBCa or NMIBCa). RESULTS: Urinary sMet levels accurately distinguished patients with BCa from those without (p<0.0001, area under the curve (AUC): 0.7008) with limited sensitivity (61%) and moderate specificity (76%), and patients with MIBCa (n=42) from those with NMIBCa (n=141; p<0.0001, AUC: 0.8002) with moderate sensitivity and specificity (76% and 77%, respectively) and low false negative rate (8%). CONCLUSIONS: Urinary sMet levels distinguish patients with BCa from those without, and patients with or without MIBCa, suggesting the potential utility of urinary sMet as a BCa biomarker for surveillance following initial treatment. Further studies are warranted to determine its potential value for prognosis in advanced disease, predicting treatment response, or identifying patients likely to benefit from Met-targeted therapies.

Spitz melanoma with MAP3K8::ABLIM1 rearrangement: a case report with review of the literature.

BACKGROUND: Spitz tumors are relatively uncommon melanocytic lesions, typically affecting a relatively younger population but can be encountered at any age. They are characterized by a proliferation of melanocytes with epithelioid and/or spindled cytomorphology features, and interpretation is often challenging. The majority of these tumors are driven by kinase fusions or HRAS mutations. MAP3K8 fusions, although rare, are characteristic genomic events in Spitz tumors, especially in more atypical or malignant lesions. CASE PRESENTATION: Here, we present the case of a 43-year-old woman with a clinically cystic mass in her right groin, histologically characterized as a spindle and epithelioid cell malignant tumor. Immunohistochemistry revealed diffuse expression of S100 protein, tyrosinase and SOX10, patchy weak PRAME, HMB45 and Melan-A reactivity, and negative staining for BRAF V600E. Next-generation sequencing analysis revealed the presence of a MAP3K8::ABLIM1 fusion gene, as well as GRIN2A and TERT promoter mutations. The morphology, immunohistochemistry and molecular analysis confirmed Spitz melanoma with molecular features suggesting a worse prognosis. CONCLUSION: This case introduces a novel fusion partner of MAP3K8 in the context of Spitz melanoma and expands the morphologic and molecular spectrum of Spitz melanoma.

Radiation and systemic immunotherapy for metastatic uveal melanoma: a clinical retrospective review.

Introduction: Metastatic uveal melanoma (mUM) is a difficult to treat disease. The liver is the primary site of metastasis in most patients, though uveal melanoma spreads widely in advanced disease. The only FDA approved immunotherapy medication for metastatic uveal melanoma is the HLA-A02:01 restricted bispecific T cell engager drug, Tebentafusp. Checkpoint inhibitor strategies and combination approaches have been tried with some limited success. We describe our experience treating patients at the University of Minnesota. Methods: Patients were included if they had biopsy-confirmed mUM. Twenty-five (25) patients meeting the criteria were identified. Medical records were reviewed and data extracted for patient baseline characteristics and response to treatments. Results: Median time to metastasis from the time of local therapy to the eye was 14.2 months (IQR; 9.3-22.0), and first site of metastasis was liver in 92% of patients. Two patients (8%) did not receive systemic therapy or radiation therapy for metastatic disease. Twenty-three (92%) patients received systemic therapy, 13 patients (52%) received ipilimumab-nivolumab as the first-line, while 4 patients (16%) received pembrolizumab. Landmark survival analysis by receipt of systemic therapy and radiation therapy treatments within 6 months of biopsy confirmed diagnosis is shown. Twenty patients (80%) received systemic therapy within 6 months of mUM diagnosis. Thirteen patients (52%) received liver directed radiation therapy within 6 months of mUM diagnosis. Discussion: Within our cohort, there was no overall survival benefit for patients receiving treatment of metastatic disease within 6 months of mUM diagnosis, versus those electing later or no treatment at all. There was remarkable clinical activity of ipilimumab and nivolumab in a subset of patients with mUM, in agreement with prior studies, and metastatic PD-L1 positive tumors were associated with a prolonged survival.