The prognostic value of the Merkel cell polyomavirus serum antibody test: A dual institutional observational study.

BACKGROUND: Merkel cell carcinoma (MCC) is an aggressive cancer with often poor outcomes. Limited biomarkers exist for predicting clinical outcomes. The Merkel cell polyomavirus (MCPyV) serum antibody test (AMERK) has shown potential for indicating better recurrence-free survival in a single-institution study. The study aimed to evaluate the link between initial AMERK serostatus and survival. Secondary objectives included examining the relationship between initial AMERK titer levels and tumor burden. METHODS: A retrospective cohort study across two institutions analyzed patients tested with AMERK within 90 days of MCC diagnosis. Regression models assessed the association of survival outcomes with serostatus, considering various factors. The relationship between AMERK titer and tumor burden indicators was evaluated using ANOVA. Significance testing was exploratory, without a fixed significance level. RESULTS: Of 261 MCC patients tested, 49.4% were initially seropositive (titer ≥75). Multivariable analysis showed that seropositivity improved recurrence, event-free, overall, and MCC-specific survival rates. Strong associations were found between initial AMERK titer and clinical, tumor, and nodal stages, tumor size, and disease extent. Notably, improved survival with seropositivity was observed only in patients with localized disease at initial presentation. CONCLUSION: Circulating antibodies to MCPyV oncoproteins, as indicated by the AMERK test, are linked with better survival in MCC patients with localized disease at presentation. This could enhance patient risk profiling and treatment personalization. The study's retrospective nature and exploratory analysis are key limitations. PLAIN LANGUAGE SUMMARY: Merkel cell carcinoma (MCC) is a potentially aggressive skin cancer, and tools to predict patient outcomes are limited. A blood test called anti-Merkel cell panel (AMERK), which checks for specific antibodies related to this cancer, might give us some clues. In this study, we looked at 261 MCC patients who took the AMERK test within 90 days of diagnosis. We found that patients with an initial positive AMERK result tended to have better outcomes, especially if their cancer was in the early stages. However, it is important to note that this study has limitations, including using retrospective data and exploratory analyses.

Patterns of initial distant metastases in 151 patients undergoing surveillance for treated Merkel cell carcinoma.

BACKGROUND: Merkel cell carcinoma (MCC) is associated with high rates of recurrence and distant metastatic progression. Current guidelines for surveillance imaging are not evidence based. Better characterization of the pattern of distant metastatic spread will better inform surveillance and facilitate earlier detection of metastases. OBJECTIVES: This retrospective study aimed to assess potential relationships between primary tumour site and site of initial distant metastasis, time to distant metastasis, overall survival (OS) and MCC-specific death (MSD). METHODS: Patients with local or regional (Stage I-III) disease who were treated with curative intent and progressed to Stage IV were included in this study (n = 151). Fisher's exact test was used to assess differences in patterns of initial distant metastases based on primary tumour site. Time to initial distant metastasis was calculated from date of MCC diagnosis. OS and MSD were calculated from date of initial distant metastasis to date of death from any or MCC-related causes, respectively. RESULTS: Of 151 patients included in analysis, 89 (58.9%) had a single initial distant metastatic site, and 62 (41.1%) had multiple sites. Patients with upper limb primary tumours were significantly less likely to develop distant lymph node or liver metastases (p = 0.02 and 0.04, respectively). Median time to distant metastasis was 11 months (IQR 6.7-17.9 months). Median OS was 15.3 months, and was shorter for patients with liver (7.0 months, p = 0.0004) or bone metastases (8.9 months, p < 0.0001). Using skin/soft tissue metastasis as a reference group, patients with multiple metastatic sites had significantly higher hazards of MSD (HR = 3.46 univariate, 3.77 multivariate analysis). Time to distant metastasis, OS and MSD did not differ by viral status. CONCLUSION: Sites of initial distant metastasis are related to primary tumour sites and survival outcomes. Because patients often have multiple initial metastases, full-body cross-sectional rather than region-specific imaging may facilitate earlier detection of metastatic disease.

Neoadjuvant-Intent Immunotherapy in Advanced, Resectable Cutaneous Squamous Cell Carcinoma.

Importance: In clinical trials, preoperative immune checkpoint inhibitors (ICIs) have shown clinical activity in advanced cutaneous squamous cell carcinoma (cSCC). However, these studies excluded patients with relevant comorbidities. Objective: To evaluate radiologic and pathologic response rates to neoadjuvant-intent programed cell death protein 1 (PD-1) ICIs in a clinical population. Design, Setting, and Participants: This cohort study of patients who were treated with neoadjuvant cemiplimab or pembrolizumab for advanced cSCC from January 2018 to January 2023 was conducted at 2 academic institutions in Boston, Massachusetts. Median follow-up was 9.5 months (range, 1.2-40.5). Exposures: Cemiplimab or pembrolizumab. Main Outcomes and Measures: Primary outcomes were radiologic and pathologic response rates. Secondary outcomes were 1-year recurrence-free survival, progression-free survival, disease-specific survival, and overall survival. Results: This cohort study included 27 patients (including 9 patients [33.3%] with a history of lymphoma). Most patients were male (18 of 27 [66.7%]), with a median age of 72 years (range, 53-87 years). Most primary tumors were located on the head/neck (21 of 27 [77.8%]). There were no unexpected delays in surgery. The median number of doses before surgery was 3.5 (range, 1.0-10.0). Five patients (18.5%) ultimately declined to undergo planned surgery due to clinical responses or stability, and 1 (3.7%) did not undergo surgery due to progressive disease. The overall pathologic response rate (pathological complete response [pCR] or major pathological response) was 47.4% (9 of 19), and the overall radiologic response rate (radiologic complete response or partial response) was 50.0% (8 of 16). The pCR rate (7 of 19 [36.8%]) was higher than the radiologic complete response rate (2 of 16 [12.5%]). The pCR rate among patients with cSCC and concomitant lymphoma was 25.0%. The 1-year recurrence-free survival rate was 90.9% (95% CI, 50.8%-98.7%), progression-free survival was 83.3% (95% CI, 27.3%-97.5%), disease-specific survival was 91.7% (95% CI, 53.9%-98.8%), and overall survival was 84.6% (95% CI, 51.2%-95.9%). Conclusions and Relevance: The results of this cohort study support the reproducibility of neoadjuvant-intent immunotherapy for cSCC in the clinical setting, including for patients with a history of lymphoma. Outside of clinical trials, it is not infrequent for patients to opt out of surgery for regressing tumors. The inclusion of higher-risk patients and preference for nonsurgical treatment are 2 factors that might explain the numerically lower pathologic response rate in this institutional experience.

Tumor cell-intrinsic PD-1 promotes Merkel cell carcinoma growth by activating downstream mTOR-mitochondrial ROS signaling.

Merkel cell carcinoma (MCC) is a rare and aggressive skin cancer. Inhibitors targeting the programmed cell death 1 (PD-1) immune checkpoint have improved MCC patient outcomes by boosting antitumor T cell immunity. Here, we identify PD-1 as a growth-promoting receptor intrinsic to MCC cells. In human MCC lines and clinical tumors, RT-PCR-based sequencing, immunoblotting, flow cytometry, and immunofluorescence analyses demonstrated PD-1 gene and protein expression by MCC cells. MCC-PD-1 ligation enhanced, and its inhibition or silencing suppressed, in vitro proliferation and in vivo tumor xenograft growth. Consistently, MCC-PD-1 binding to PD-L1 or PD-L2 induced, while antibody-mediated PD-1 blockade inhibited, protumorigenic mTOR signaling, mitochondrial (mt) respiration, and ROS generation. Last, pharmacologic inhibition of mTOR or mtROS reversed MCC-PD-1:PD-L1-dependent proliferation and synergized with PD-1 checkpoint blockade in suppressing tumorigenesis. Our results identify an MCC-PD-1-mTOR-mtROS axis as a tumor growth-accelerating mechanism, the blockade of which might contribute to clinical response in patients with MCC.

Liquid biopsy epigenomic profiling for cancer subtyping.

Although circulating tumor DNA (ctDNA) assays are increasingly used to inform clinical decisions in cancer care, they have limited ability to identify the transcriptional programs that govern cancer phenotypes and their dynamic changes during the course of disease. To address these limitations, we developed a method for comprehensive epigenomic profiling of cancer from 1 ml of patient plasma. Using an immunoprecipitation-based approach targeting histone modifications and DNA methylation, we measured 1,268 epigenomic profiles in plasma from 433 individuals with one of 15 cancers. Our assay provided a robust proxy for transcriptional activity, allowing us to infer the expression levels of diagnostic markers and drug targets, measure the activity of therapeutically targetable transcription factors and detect epigenetic mechanisms of resistance. This proof-of-concept study in advanced cancers shows how plasma epigenomic profiling has the potential to unlock clinically actionable information that is currently accessible only via direct tissue sampling.

No difference in survival for primary cutaneous Merkel cell carcinoma after Mohs micrographic surgery and wide local excision.

BACKGROUND: The preferred treatment for clinically node-negative Merkel cell carcinoma (MCC) is surgical excision in conjunction with sentinel lymph node biopsy. There is limited large-scale research on survival outcomes by surgical approach for management of the primary tumor. OBJECTIVE: To compare overall and MCC-specific survival outcomes in clinically and pathologically, node-negative MCC patients treated with wide-local excision (WLE) and Mohs micrographic surgery (MMS) in a nationally representative sample. METHODS: Overall and MCC-specific survival outcomes for primary MCC tumors contained in the SEER (Surveillance, Epidemiology, and End Results)-18 database from 1989 to 2015 were stratified by surgical modality and analyzed via competing risk analysis. RESULTS: A total of 2359 US adults with MCC were included in the analysis. For overall and MCC-specific survival, there was no significant difference in survival outcomes between WLE and MMS on multivariable analysis (hazard ratio, 1.04 [95% CI, 0.88-1.22]; subdistribution hazard ratio, 0.76 [95% CI, 0.53-1.09]). Sentinel lymph node biopsy was associated with improved overall survival and MCC-specific survival. LIMITATIONS: Retrospective design of SEER and the lack of covariates such as comorbidities and immunostaining. CONCLUSION: There is no survival disadvantage for MMS compared to WLE as the surgical modality for primary cutaneous MCC. Sentinel lymph node biopsy should be coordinated prior to MMS.

YAP1 and WWTR1 expression inversely correlates with neuroendocrine markers in Merkel cell carcinoma.

BackgroundMerkel cell carcinoma (MCC) is an aggressive neuroendocrine (NE) skin cancer caused by severe UV-induced mutations or expression of Merkel cell polyomavirus (MCPyV) large and small T antigens (LT and ST). Despite deep genetic differences between MCPyV-positive and -negative subtypes, current clinical diagnostic markers are indistinguishable, and the expression profile of MCC tumors is, to our knowledge, unexplored.MethodsHere, we leveraged bulk and single-cell RNA-Seq of patient-derived tumor biopsies and cell lines to explore the underlying transcriptional environment of MCC.ResultsStrikingly, MCC samples could be separated into transcriptional subtypes that were independent of MCPyV status. Instead, we observed an inverse correlation between a NE gene signature and the Hippo pathway transcription factors Yes1-associated transcriptional regulator (YAP1) and WW domain-containing transcriptional regulator 1 (WWTR1). This inverse correlation was broadly present at the transcript and protein levels in the tumor biopsies as well as in established and patient-derived cell lines. Mechanistically, expression of YAP1 or WWTR1 in a MCPyV-positive MCC cell line induced cell-cycle arrest at least in part through TEA domain-dependent (TEAD-dependent) transcriptional repression of MCPyV LT.ConclusionThese findings identify what we believe to be a previously unrecognized heterogeneity in NE gene expression within MCC and support a model of YAP1/WWTR1 silencing as essential for the development of MCPyV-positive MCC.FundingUS Public Health Service grants R35CA232128, P01CA203655, and P30CA06516.

Reversal of viral and epigenetic HLA class I repression in Merkel cell carcinoma.

Cancers avoid immune surveillance through an array of mechanisms, including perturbation of HLA class I antigen presentation. Merkel cell carcinoma (MCC) is an aggressive, HLA-I-low, neuroendocrine carcinoma of the skin often caused by the Merkel cell polyomavirus (MCPyV). Through the characterization of 11 newly generated MCC patient-derived cell lines, we identified transcriptional suppression of several class I antigen presentation genes. To systematically identify regulators of HLA-I loss in MCC, we performed parallel, genome-scale, gain- and loss-of-function screens in a patient-derived MCPyV-positive cell line and identified MYCL and the non-canonical Polycomb repressive complex 1.1 (PRC1.1) as HLA-I repressors. We observed physical interaction of MYCL with the MCPyV small T viral antigen, supporting a mechanism of virally mediated HLA-I suppression. We further identify the PRC1.1 component USP7 as a pharmacologic target to restore HLA-I expression in MCC.

A Retrospective Study of Ipilimumab Plus Nivolumab in Anti-PD-L1/PD-1 Refractory Merkel Cell Carcinoma.

Merkel cell carcinoma (MCC) is a very rare but highly aggressive cutaneous neuroendocrine carcinoma and is associated with chronic exposure to ultraviolet light and the Merkel cell polyoma virus. The incidence rate of MCC is increasing and MCC is associated with high rates of recurrence and mortality. Immune checkpoint inhibitors (ICIs) offer durable responses and significant clinical benefit with 2 agents-avelumab (anti-PD-L1) and pembrolizumab (anti-PD-1)-currently approved by the U.S. Food and Drug Administration for the treatment of advanced MCC. Despite the advances in systemic therapy options for MCC, ~50% of patients with advanced MCC treated with ICI progress on therapy. There is a paucity of studies assessing second-line systemic therapy following primary/acquired resistance to ICIs. Current management in this setting remains a clinical challenge especially in trial ineligible patients. We evaluated objective response to ipilimumab plus nivolumab in metastatic MCC refractory to anti-PD-(L)1 therapy. Thirty-one percent of patients experienced a grade III or grade IV immune-related adverse event (irAE) due to ipilimumab plus nivolumab. No patients (0/13) achieved a complete or partial response via RECISTv1.1/irRECIST. Twenty-three percent (3/13) of patients achieved stable disease as the best overall response but progressed shortly thereafter. The median progression-free survival was 1.3 months (90% CI 1.1-1.5) from the initiation of ipi-nivo. The median overall survival was 4.7 months (95% CI 3-17). This study suggests limited, if any, clinical benefit of ipi-nivo in patients with advanced anti-PD-L1/anti-PD-1 refractory MCC.

Characterization of clinical outcomes after shorter course hypofractionated and standard-course radiotherapy for stage I-III curatively-treated Merkel cell carcinoma.

BACKGROUND: Limited data exists regarding the efficacy of curative hypofractionated radiotherapy (hypo-RT) regimens compared to conventionally-fractionated radiotherapy (conv-RT) for Merkel cell carcinoma (MCC). METHODS: A retrospective analysis of 241 patients diagnosed with non-metastatic MCC from 2005-2021 and who received RT at Dana-Farber/Brigham & Women's Cancer Center. The primary outcome was cumulative incidence of in-field locoregional relapse using Gray's test with competing risks of death and isolated out-of-field recurrence. Secondary outcomes included overall survival (OS) and MCC-specific survival using log-rank tests, and risk factors of recurrence using Cox-proportional hazards regression. RESULTS: There were 50 (20.6 %) and 193 (79.4 %) courses of hypo-RT and conv-RT, respectively. The hypo-RT cohort was older (≥73 years at diagnosis: 78.0 % vs 41.5 %, p < 0.01), and received a lower equivalent total RT dose in 2 Gy per fraction (<50 Gy: 58.0 % vs 5.2 %, p < 0.01). Median follow-up was 65.1 months (range: 1.2-194.5) for conv-RT and 25.0 months (range: 1.6-131.3) for hypo-RT cohorts. Two-year cumulative incidence of in-field locoregional relapse was low in both groups (1.1 % conv-RT vs 4.1 % hypo-RT, p = 0.114). While two-year OS was lower for the hypo-RT group (62.6 % vs 84.4 %, p = 0.0008), two-year MCC-specific survival was similar (84.7 % vs 86.6 %, p = 0.743). On multivariable analysis, immunosuppression, clinical stage III disease, and lymphovascular invasion were associated with any-recurrence when controlling for sex, age, and hypo-RT. CONCLUSIONS AND RELEVANCE: There was no difference in cumulative incidence of in-field locoregional relapse or MCC-specific survival between hypo-RT and conv-RT. Prospective studies are needed to confirm hypo-RT as an efficacious treatment option for MCC.

Predictors of immunotherapy benefit in Merkel cell carcinoma.

Merkel cell carcinoma is a rare cancer for which immune checkpoint blockade is standard-of-care for recurrent/metastatic disease. However, not all patients benefit from immunotherapy. A greater understanding of molecular mechanisms and predictive biomarkers are unmet needs. We retrospectively analyzed electronic health records and next-generation sequencing data of 45 patients treated at our institution from 2013 to 2020 to understand clinical and genomic correlates of benefit from immunotherapy. Our cohort predominantly included individuals with stage III disease at primary disease diagnosis and individuals with stage IV disease at recurrent/metastatic disease diagnosis. Most received immunotherapy as first-line treatment. 43% experienced objective response (median duration of response 24.2 months, 95% confidence interval 8.8-not reached). Median overall survival was 15.5 months (95% confidence interval 9.0-28.7) (median follow-up 25.2 months). Less advanced stage at primary disease diagnosis and shorter disease-free interval between completion of initial treatment and recurrence were each associated with greater odds of response (odds ratio of 0.06, p = 0.04 for stage; odds ratio 0.75, p = 0.05 for disease-free interval). Single-nucleotide variants in ARID2 and NTRK1 were associated with response (p = 0.05, without Bonferroni correction), while none of Merkel cell polyomavirus status, total mutational burden, ultraviolet mutational signatures, and copy-number alterations predicted outcomes. Patients with shorter disease-free interval may be particularly suitable immunotherapy candidates. Our molecular findings point to ARID2 and NTRK1 as potential predictive markers and/or therapeutic targets (e.g., with Trk inhibitors), although this association needs to be confirmed in a larger sample.

Real-world outcomes treating patients with advanced cutaneous squamous cell carcinoma with immune checkpoint inhibitors (CPI).

BACKGROUND: Immunotherapy has revolutionised the treatment of advanced cutaneous squamous cell carcinoma (cSCC). It is important to understand both safety and efficacy in a real-world and trial-ineligible cSCC population. We aimed to evaluate safety, efficacy and molecular insights among a broader cSCC population, including immunosuppressed patients, treated with immune checkpoint inhibitors (CPI). METHODS: We present a cohort of advanced cSCC patients (n = 61) treated from 2015 to 2020 evaluating the best overall response (BOR) (RECISTv1.1) to CPI therapy, immune-related adverse events (irAEs) and tumour mutational burden (TMB) to correlate with outcomes. A validated geriatric scoring index (CIRS-G) was utilised to assess comorbidities among patients ≥75. These data were compared with published clinical trial results among the broader cSCC population. RESULTS: BOR to CPI was lower among the entire cohort when compared with trial data (31.5 vs. 48%, P < 0.01), with higher rates of progression (59 vs. 16.5%, P < 0.01), regardless of immunosuppression history or age. Grade 3+ irAEs were more common among responders (P = 0.02), while pre-treatment lymphocyte count and TMB predicted response (P = 0.02). CONCLUSIONS: We demonstrate comparatively lower response rates to CPI among real-world cSCC patients not explained by older age or immunosuppression history alone. Immune-related toxicity, absolute lymphocyte count and TMB predicted CPI response.

Association of Programmed Death 1 Protein Ligand (PD-L1) Expression With Prognosis in Merkel Cell Carcinoma.

Background: Merkel cell carcinoma (MCC) is a rare neuroendocrine skin cancer. Prior to the advent of immunotherapy, treatment options were limited. In our study, we evaluate the impact of tumor cell PD-L1 expression and tumor immune microenvironment on survival in MCC patients who were not treated with immune checkpoint inhibitors. Methods: Clinical data and tissue samples were collected from 78 patients with confirmed MCC treated at Dana-Farber Cancer Institute. Specimens were analyzed for the distribution of PD-L1 by immunohistochemistry staining (IHC) and standardized analysis. Results were correlated with survival data. Results: In this study, membrane and cytoplasmic MCC tumor cell staining for PD-L1 was detected in 22.4% (15 of 67) of cases and PD-L1 staining of intratumoral microvessels and PD-L1 positive immune cells at the infiltrative margins of the tumor in 92.5% (62 of 67) of cases. In patients untreated with immune checkpoint inhibitors, median overall survival was not different for patients based on PD-L1 expression (PD-L1+ 64 months vs. PD-L1- not reached; HR = 1.26, 95% CI: 0.46-3.45; p = 0.60). Conclusion: PD-L1 expression is frequently detected in MCC tumor cells and tumor microenvironment. PD-L1 expression did not affect prognosis in this cohort that had not received PD-1/L1 blockade.

Clinical and molecular characterization of virus-positive and virus-negative Merkel cell carcinoma.

BACKGROUND: Merkel cell carcinoma (MCC) is a highly aggressive neuroendocrine carcinoma of the skin caused by either the integration of Merkel cell polyomavirus (MCPyV) and expression of viral T antigens or by ultraviolet-induced damage to the tumor genome from excessive sunlight exposure. An increasing number of deep sequencing studies of MCC have identified significant differences between the number and types of point mutations, copy number alterations, and structural variants between virus-positive and virus-negative tumors. However, it has been challenging to reliably distinguish between virus positive and UV damaged MCC. METHODS: In this study, we assembled a cohort of 71 MCC patients and performed deep sequencing with OncoPanel, a clinically implemented, next-generation sequencing assay targeting over 400 cancer-associated genes. To improve the accuracy and sensitivity for virus detection compared to traditional PCR and IHC methods, we developed a hybrid capture baitset against the entire MCPyV genome and software to detect integration sites and structure. RESULTS: Sequencing from this approach revealed distinct integration junctions in the tumor genome and generated assemblies that strongly support a model of microhomology-initiated hybrid, virus-host, circular DNA intermediate that promotes focal amplification of host and viral DNA. Using the clear delineation between virus-positive and virus-negative tumors from this method, we identified recurrent somatic alterations common across MCC and alterations specific to each class of tumor, associated with differences in overall survival. Finally, comparing the molecular and clinical data from these patients revealed a surprising association of immunosuppression with virus-negative MCC and significantly shortened overall survival. CONCLUSIONS: These results demonstrate the value of high-confidence virus detection for identifying molecular mechanisms of UV and viral oncogenesis in MCC. Furthermore, integrating these data with clinical data revealed features that could impact patient outcome and improve our understanding of MCC risk factors.

Chromosome 3q arm gain linked to immunotherapy response in advanced cutaneous squamous cell carcinoma.

AIMS: The activity that the immune checkpoint inhibitor (ICI) cemiplimab has recently demonstrated has led to a paradigm shift in the management of patients with advanced cutaneous squamous cell carcinoma (cSCC). To identify predictive biomarkers of response to ICIs in advanced cSCC, we studied 33 patients who received ICI therapy at the Dana-Farber/Harvard Cancer Center (DF/HCC) and analysed sequencing data for a subset of these patients. METHODS: We collected clinical data using electronic health records and genomic data using the institutional OncoPanel platform of the DF/HCC. We compared tumour genomics with data from previously sequenced cSCC cohorts. RESULTS: We observed high tumour mutational burden regardless of smoking status and response to ICI and longer median overall survival among those patients who achieved an ICI response. We compared the genetic data from our cohort with data from other cohorts that included fewer patients with distant metastatic disease. Although our cohort had a similar genetic landscape to those of comparator cohorts, mutations in PIK3C2B were more common in our study. In our cohort, copy number alterations (CNAs) in the 3q chromosomal arm appeared to predict response to ICI therapy. CONCLUSION: CNAs in the 21-27 bands of chromosome arm 3q, a region that includes PIK3CA, ETV5 and BCL6, may represent predictors of response to ICI and may be candidates for drug targeting in combination or sequence with ICI agents.