Brisk Tumor-infiltrating Lymphocytes Research Articles

Systemic Inflammation, the Peripheral Blood Transcriptome, and Primary Melanoma

Peripheral blood transcriptomes from 383 newly-diagnosed melanoma patients were subjected to differential gene expression analysis. The hypotheses were that impaired systemic immunity is associated with poorer prognosis (thicker tumors and fewer tumor infiltrating lymphocytes (TILs)) and evidence of systemic inflammation (high-sensitivity C-reactive protein (hsCRP) and fibrinogen levels). Higher fibrinogen levels were associated with thicker primaries. In single gene analysis hsCRP levels were significantly associated with higher blood CD274 expression, (coding for PD-L1), but each was independently prognostic, with hsCRP associated with increased mortality, and higher CD274 protective, independent of age. Pathway analysis identified downregulation of immune cell signalling pathways in the blood of people with thicker tumors and notable upregulation of STAT1 in people with brisk TILs. Transcriptomic data provided evidence for increased NFB signalling with higher inflammatory markers but with reduction in expression of HLA class II molecules and higher CD274 suggesting that aberrant systemic inflammation is a significant mediator of reduced immune function in melanoma. In summary, transcriptomic data revealed evidence of reduced immune function in patients with thicker tumors and fewer TILs, at diagnosis. Inflammatory markers were associated with thicker primaries and independently with death from melanoma suggesting that systemic inflammation contributes to that reduced immune function.

Tumor-Associated Macrophages in Merkel Cell Carcinoma: Old Balances, New Checks.

A recent report sheds light on the tumor-associated macrophages (TAM) in Merkel cell carcinoma (MCC), and the association of S100A8-expressing TAMs with resistance to anti-PD-(L)1 inhibitors. These data improve our understanding about why some tumors with brisk tumor-infiltrating lymphocytes do not respond to immunotherapy and provide a compelling rationale to target myeloid checkpoints in MCC. See related article by Tabachnick-Cherny et al., p. xxxx.

Correlation of patient characteristics and tumor histology in patients (pts) with locally advanced cutaneous squamous cell carcinoma (LACSCC) receiving immune checkpoint inhibitor treatment (ICI): A retrospective analysis.

e21553 Background: Cutaneous squamous cell carcinoma (CSCC) arising from epidermal keratinocytes is highly curable with surgery. Functional anti-cancer immunity is protective of CSCC and high mortality due to local or systemic dissemination is seen in pts. with immune deficiency. ICI treatment is a logical FDA approved option that result in improved outcomes in affected pts. We report our experience in LACSCC pts. receiving ICI treatment and describe correlation between pt. characteristics, tumor histology and treatment outcome. Methods: Following Institutional Review Board approval, we did a retrospective analysis of LACSCC pts. receiving ICI therapy at the Neag Cancer Center who received at least one dose of ICI between 2018-2023. Analysis includes pt. demographics, relevant comorbidities, medications, and tumor histology and tumor infiltrating lymphocytes (TILs) of tumors reviewed with dermatopathologist. Results: Between 2018-2023, 12 pts. with LACSCC met the criteria for analysis. Median age 77.5 years (yrs.), (range 65-102), 9 males, 3 females, tumors sites - head and neck region 8 pts., torso 2 pts., legs 2 pts. Relevant past medical history included treatment of prostate cancer 3, chronic lymphocytic Leukemia 1, psoriasis 2 (one received anti TNF treatment), and MGUS, Parkinson’s disease and dementia in 1 each. Majority of pts. had significant smoking history (9/12), lymphopenia in 8 out of 12 pts. Tumors were poorly differentiated in 6 out of 12 pts. Brisk TILs was present in 7 and non-brisk TILs in 5 pts. 3 pts. with brisk TILs achieved complete remission (CR), one very good partial response (VGPR), 1 partial response (PR) and 2 not evaluable for response. Out of 5 pts. with non-brisk TILS, one achieved CR, one achieved VGPR, 1 PR, 1 stable disease (SD), and 1 progressive disease (PD) in a pt. who received anti TNF treatment of psoriasis. 9 patients are alive between 1 week and 50 months after initiation of treatment. 3 pts. died, one of PD, one of bacterial sepsis and one of Covid. Conclusions: High risk LACSCC affected males with smoking histories in the majority. Durable responses following ICI treatment occurred irrespective of age, while resistant disease was noted in 1 pt. with previous anti TNF treatment. Brisk TILs predicted high response rate and durable responses although some pts. with non-brisk TILs also showed favorable outcomes. Association with previous cancer and macular degeneration may reflect underlying immune deficiency. Three pts. developed infection within two weeks of ICI treatment (urinary tract infection, wound infection, Covid). More studies will shed light at factors compounding immune deficiency and safety of this treatment in a high-risk population.

Sinonasal Mucosal Melanoma: Role of Tumor Proliferative Indices and Pathological Factors in Survival.

The objective of this study is to determine the association of proliferation indices and pathologic biomarkers on overall and recurrence/metastasis-free survival (OS and RMFS) in patients with sinonasal mucosal melanoma (SNMM) and to assess the genetic mutational landscape of SNMM. This is a retrospective cohort study of 45 SNMM patients without neoadjuvant therapy who underwent surgical therapy with curative intent and had tumor tissue available for histopathologic review, molecular analysis, and genetic mutational assessment. The OS and RMFS were assessed for associations with numerous tumor and patient-related factors. Among proliferative indices, higher Ki67 and mitotic rates were associated with worsened OS and RMFS (Ki67: p=0.0007 and p < 0.0001; mitotic rate: p=0.005 and p=0.0009, respectively). The presence of brisk tumor-infiltrating lymphocytes (TILs) was associated with improved RMFS (p=0.007) and the presence of lymphovascular invasion was associated with worsened OS and RMFS (p=0.02 and p=0.04, respectively). Patients with amelanotic tumors were more likely to have higher T-stage (p=0.046), less likely to have brisk TILs (p=0.02) and had worsened RMFS (p=0.03). Patients on immunotherapy with tumor Ki67 < 40% had better 3-year OS compared to those with higher Ki67 index (p=0.004). Actionable genetic mutations such as BRAF V600E are rare and present in only 1 of 20 patients tested. In SNMM patients, pathologic and proliferation markers such as Ki67, mitotic rate and brisk TILs are associated with survival and may be considered in future staging systems. Clinical response to immunotherapy appears to correlate with the Ki67 index. Given the distinct genetic profile of SNMM, targeted therapies against the MAPK kinase pathway have limited utility. 3 Laryngoscope, 132:2350-2358, 2022.

Survival outcomes following immune checkpoint inhibitor therapy of advanced cutaneous melanoma by age and tumor infiltrating lymphocytes in the primary cutaneous site.

e21514 Background: Tumor infiltrating lymphocytes (TILs) in the tumor microenvironment generate anti-tumor immunity and are associated with response to immune checkpoint inhibitor (ICI) therapy of melanoma. However, likelihood of response to ICI therapy depends upon intensity of TILs, proportion of effector and regulatory T cells and age. Methods: In a research study approved by the Institutional Review Board of University of Connecticut, we retrospectively analyzed outcome of patients diagnosed with cutaneous melanoma between 2009 and 2019 and developed unresectable stage III or stage IV disease requiring ICI treatment. Patients were stratified by age (&lt; 75 and &gt; 75 years old) and TILs at the primary cutaneous melanoma site categorized as brisk (present throughout vertical phase), non-brisk (present in one of more foci of vertical phase) or absent. Patient’s age at the time of metastases, as well as 12-month and 24-month overall survival (OS) following ICI treatment was recorded. Results: A total of 93 patients were screened, and 68 were included for analysis of survival outcome. Patients had to receive at least one dose of ICI treatment at UConn Health and had available data on TILs of primary cutaneous melanoma. In patients over 75 years of age, those with brisk TILs had an increased 24-month survival compared to those with non-brisk or absent TILs (100% vs. 45%, p = 0.04). In patients &lt; 75 years of age, the 24 month survival was similar in those with brisk and non-brisk or absent TILs (67% vs. 69%, p = .91). There was no significant difference in overall survival between patients &lt; 75 or &gt; 75 years with brisk TILs (67% vs. 100%, p = .19). Additionally, in patients with non-brisk or absent TILs, there was a not quite significant difference in survival between those &gt; 75 vs &lt; 75 years (45% vs. 69%, p = .09). Conclusions: Our findings suggest that intensity of TILs in primary cutaneous melanoma is predictive of outcome following ICI treatment of advanced melanoma in patients over 75 years of age. These findings support possible interaction between qualitative differences of TILs and patient age. Our findings may help identify older melanoma patients who may benefit from ICI treatment.

Immunometabolic predictive factors in Merkel cell carcinoma (MCC) patients treated with avelumab.

e21525 Background: Merkel cell carcinoma (MCC) is a rare and aggressive skin cancer, associated with a worse prognosis. The link between MCC and immune suppression is well demonstrated. The population of patients with MCC is frequently elderly and frail, making it essential to determine whether the results of clinical trial can be replicated in a real-world setting. Despite the introduction of immune-checkpoint inhibitors (ICIs) has provided great benefit for some patients with advanced MCC (aMCC), it remains a subsets of patients who are refractory to ICIs or develop acquired resistance over time. Thus, there is a clinical need for predictive factors of ICI response. Methods: Twenty patients with aMCC treated with avelumab were included. The treatment was administered as I or II line. Clinical-pathological characteristics, Body Mass Index (BMI), and response to avelumab were analyzed from a MCC System database prospectively collected. An explorative analysis was performed, for available samples, to investigate: i) the plasma levels of soluble PD-1 (sPD-1), and PD-L1 (sPD-L1) collected at baseline, measured using homemade ELISA assays not yet commercially available, and designed according to investigator specifications; ii) IHC for PD-L1 in tumor samples; iii) the presence/absence/class (brisk vs no-brisk) of tumor-infiltrating lymphocytes (TILs) in tumor samples. The primary outcome investigated was the Time to Treatment Failure (TTF). Results: From February 2019 to January 2022, twenty (20) patients were included in the study. The median age was 74 (range 56-83); 10 patients were men (50%) and 10 were women (50%). Seventeen (17) patients (85%) were treated with avelumab as I line, and 3 patients (15%) as II line. The overall response rate was 65% (70.6% in I line patients). One (1) patient (5%) had a complete response (CR), 13 patients (65%) partial response (RP), 4 patients (20%) stable disease (SD), and 2 patients (10%) had a progression disease (PD). Overall median TTF was 22 months (95% confidence interval [CI]: -13.0-30.9). At the time of data analyses, a total of 9 events (progression or death) occurred (45%). Notably, a BMI ≥ 30 was significantly associated with longer TTF (p = 0.004) and objective response rate (p = 0.01). In the explorative biomarker analysis, preliminary data on 6 tumor and plasma samples, showed that plasma sPD-1 &gt; 3.8 ng/ml, and the presence of PD-L1 and brisk TILs on tumor samples, were associated to longer TTF. Conclusions: These finding highlight the complex immune-metabolic interplay in the immunotherapy response. These data extends the previous finding on “obesity paradox” and the role of BMI as predictive factors of ICIs. The data on biomarker analysis warrants further prospective validation.

Phase II study of nivolumab (nivo) with relatlimab (rela) in patients (pts) with first-line advanced melanoma: Early on-treatment major pathologic response on biopsy.

9514 Background: A phase II study of nivo and rela was designed to evaluate the antitumor activity and mechanism of this combination and components for first-line treatment of pts with advanced melanoma. Pts received lead-in treatment with 1 cycle of nivo (480mg IV q4wk), rela (160mg IV q4wk), or nivo-rela followed by combination therapy. We assessed the effect of each lead-in treatment on immune-related pathological response (irPR) at 4-wk biopsy to develop early biomarkers of antitumor response. Methods: Core biopsy of an index lesion was performed at baseline and after 4 wk on-treatment. Immune characteristics of pathological response were assessed on H&amp;E sections, including presence of tumor-infiltrating lymphocytes (TIL), neovascularization, proliferative fibrosis, plasma cells, and lymphoid aggregates. irPR score was calculated as described by Stein JE et al Ann Oncol 2019, from 0 (no irPR features) to 3 (major pathologic response on biopsy [MPRbx], ≤10% residual viable tumor). We assessed the association between irPR and radiological response (RECIST v1.1) at 4-wk evaluations. Results: The current cohort includes 22 pts, median age = 67, male = 13. Pts were randomized to nivo = 7, rela = 7, and nivo-rela = 8 lead-in groups. Two pts had no irPR evaluation due to early progression and unscorable tumor. Among 20 evaluable pts, proliferative fibrosis, neovascularization, plasma cells, brisk TIL, and lymphoid aggregates were identified in 50%, 35%, 26.3%, 25%, and 5% of cases, respectively. Lead-in nivo (n = 2/6), rela (n = 0/6), and nivo-rela (n = 3/8) resulted in irPR = 3 in 25% of pts. Radiological response was identified as partial response (PR) = 1/22 (4.5%), stable disease (SD) = 12/22 (54.5%), and progressive disease (PD) = 9/22 (41%). Among pts with PD, 44% received rela-, 33% nivo-, and 22% nivo-rela- lead-in. Pts with irPR score = 3 had radiological PR = 1, SD = 3, and PD = 1 at 4wks. No association was found between MPRbx and radiological response at 4 wks. Conclusions: Four-wk MPRbx may serve as an early biomarker of treatment response in advanced melanoma. Lead-in treatment resulted in MPRbx of 25% and was greatest with nivo-rela lead-in. Correlations between 4 wk MPRbx and later radiological responses, survival and other endpoints will be made at completion of trial accrual. Clinical trial information: NCT03743766. [Table: see text]

Clinical-Pathological Evaluation and Prognostic Analysis of 228 Merkel Cell Carcinomas Focusing on Tumor-Infiltrating Lymphocytes, MCPYV Infection and ALK Expression

Merkel cell carcinoma is a rare and aggressive primary neuroendocrine carcinoma of the skin, whose pathogenesis can be traced back to UV radiation damage or Merkel cell polyomavirus (MCPyV) infection. Despite some improvements on the characterization of the disease partly due to its increased incidence, crucial pathogenetic and prognostic factors still need to be refined. A consecutive series of 228 MCC from three hospitals in Turin was collected with the aim of both analyzing the apparent increase in MCC incidence in our area and investigating the distribution and prognostic role of clinical-pathological parameters, with a focus on MCPyV status, ALK tumor expression and tumor infiltrating lymphocytes (TILs). Review of morphology and conventional immunohistochemical staining was possible in 191 cases. In 50 cases, the expression of the novel neuroendocrine marker INSM1 was additionally assessed. Fourteen cases of MCC of unknown primary skin lesion were identified and separately analyzed. While confirming an exponential trend in MCC incidence in the last decades and providing a description of histological and cytological features of a large series of MCC, the present study concludes that 1) INSM1 is a highly sensitive marker in both skin and lymph node primary MCC; 2) positive MCPyV status, brisk TILs and lower tumor size and thickness are independent positive prognostic parameters, and the combination of the former two may provide a novel tool for prognostic stratification; 3) ALK is expressed 87% of MCC and associated with positive viral status, and could represent a prognostic biomarker, if validated in larger series.

PD01-12 PAN-CANCER CHARACTERIZATION OF THE TUMOR MICROENVIRONMENT REVEALS STROMA AND IMMUNE ENRICHED SUBTYPE WITH DISTINCT MOLECULAR, CLINICAL, AND PROGNOSTIC FEATURES IN DIFFERENT GENITOURINARY CANCERS

You have accessJournal of UrologyCME1 May 2022PD01-12 PAN-CANCER CHARACTERIZATION OF THE TUMOR MICROENVIRONMENT REVEALS STROMA AND IMMUNE ENRICHED SUBTYPE WITH DISTINCT MOLECULAR, CLINICAL, AND PROGNOSTIC FEATURES IN DIFFERENT GENITOURINARY CANCERS Mark Farha, Brittney Cotta, Srinivas Nallandhighal, Kevin Hu, Steven Goldenthal, Aaron Udager, and Simpa Salami Mark FarhaMark Farha More articles by this author , Brittney CottaBrittney Cotta More articles by this author , Srinivas NallandhighalSrinivas Nallandhighal More articles by this author , Kevin HuKevin Hu More articles by this author , Steven GoldenthalSteven Goldenthal More articles by this author , Aaron UdagerAaron Udager More articles by this author , and Simpa SalamiSimpa Salami More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002516.12AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Transcriptomic data driven tumor microenvironment (TME) deconvolution tools have enabled robust analyses correlating TME subtypes with prognosis and therapeutic response. Here, we performed a pan-cancer analysis of tumors included in the TCGA to assess the impact of the TME on oncological outcome. METHODS: Data was downloaded from the TIMEx portal. TIMEx utilizes scRNA-seq signatures from 16 solid cancers to classify 37 cell types and activation states, including malignant, benign tissue, immune, and stromal cells. Unsupervised hierarchical clustering was performed to divide the TIMEx deconvoluted TCGA data (31 cancers, n=9848) into three clusters. Prevalence of each cancer type was calculated for each of the clusters. Progression free survival (PFS) was assessed by cluster for the entire cohort and for genitourinary (GU) cancers in a sub-analysis – Bladder (BLCA), Prostate (PRAD), Kidney [Clear Cell (KIRC), Papillary (KIRP), and Chromophobe (KICH)]. Tumor infiltrating lymphocyte (TIL) architecture data was collected from Saltz et al. (2018) in which “computational staining” was performed on the TCGA pathology slide archive. RESULTS: Three clusters with similar fractions of malignant and native tissue cells, but different immune and stromal enrichment were identified: CL1ImmuneLow/StromaLow, CL2ImmuneLow/StromaMod, and CL3ImmuneHi/StromaHi. While BLCA was heterogeneous, the majority of PRAD tumors were in CL1 (65%, p<2.2x10-16). The majority of KIRC tumors were in CL3 (80%, p<2.2x10-16), while KICH were mostly made up of CL1 (52%, p<2.2x10-16), and most KIRP were in CL2 (62%, p=2.2x10-13). Overall across cancers, CL3 had a longer PFS compared to grouped CL1 and CL2 combined (100.5 mo (95% CI 83.9-NR) vs. 85.7 mo (95% CI 74.4-101 mo), p=0.016). In a survival analysis of GU cancers, CL3 was associated with a longer PFS in BLCA (p=0.03) but shorter PFS in KIRC (p=0.05). Notably, BLCA CL3 patients had fewer FGFR mutations (p=0.007), brisk TIL spatial pattern (p=1.5x10-5), and more Basal/Squamous tumors by the Kamoun classifier (p=3.6x10-8) compared to CL1 and CL2. KIRC CL3 patients on the other hand, showed no difference in detection of VHL, PBRM1, SETD2, or TCEB1 alterations compared to CL1 and CL2. CONCLUSIONS: Our analyses reveal the heterogeneous composition and prognostic implications of the TME across various cancers. Further studies are needed to explore the capacity of this classification scheme to guide prognostication and therapeutic decisions. Source of Funding: A.M.U., S.S.S. are supported by the University of Michigan Health System-Peking University Health Science Center (UMHS-PUHSC) Joint Institute (JI). S.S.S. is supported by the Robert Wood Johnson Foundation as part of the Harold Amos Medical Faculty Development Program (AMFDP). S.S.S. is supported in part by the Urology Care Foundation Rising Stars in Urology Research Award Program and Astellas, Inc. S.S.S. is supported by P30CA046592. A.M.U. is supported by the Department of Defense (W81XWH-19-1-0407) and National Institutes of Health (U01 CA232931and R37 CA222829). A.M.U. and S.S.S. are supported by the National Institutes of Health (P50 CA186786) © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e34 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Mark Farha More articles by this author Brittney Cotta More articles by this author Srinivas Nallandhighal More articles by this author Kevin Hu More articles by this author Steven Goldenthal More articles by this author Aaron Udager More articles by this author Simpa Salami More articles by this author Expand All Advertisement PDF DownloadLoading ...

Theragnostic significance of tumor-infiltrating lymphocytes and Ki67 in BRAFV600-mutant metastatic melanoma (BRIM-3 trial)

We conducted a retrospective tumor tissue analysis as part of the BRIM3 trial to evaluate the theragnostic significance of tumor-infiltrating lymphocytes (TILs) and melanoma cell proliferation. Using manual semi-quantitative analyses, we assessed the density of TILs by pathology review of tissue sections stained with hematoxylin and eosin (H&E TIL score) and by immunohistochemistry (IHC) with an anti–CD8 antibody (CD8 TIL score); also, the melanoma cell proliferation by IHC with an anti–Ki67 antibody. Three hundred and fifty-three, 280, and 172 patients’ tumor tissue samples were available for H&E, CD8, and Ki67 IHC analysis, respectively. There was no association between high (2+, 3+) peritumoral and intratumoral H&E and/or TIL CD8 score or high Ki67 proliferation index (>15%) with serum LDH level and stage IV melanoma. Neither high Ki67 proliferation, nor high peritumoral and/or intratumoral TIL score was significantly associated with objective antitumor response in any treatment arm. High intratumoral and high peritumoral CD8 TIL score was significantly associated with progression-free survival (PFS) only in DTIC-treated patients (P = 0.002 and 0.037, respectively); in vemurafenib-treated patients, high intratumoral and/or peritumoral CD8 TIL score was not significant (log-rank P = 0.053 and 0.062, respectively). Nevertheless, a high peritumoral CD8 TIL score was a significant predictor of PFS and overall survival after adjustment for age, sex, serum LDH, ECOG performance status, and treatment arm in a Cox regression model. Vemurafenib does not only benefit patients bearing brisk TILs; even vemurafenib-treated patients with absent and/or non–brisk TILs tend to have longer PFS compared to DTIC-treated patients with brisk TILs. High peritumoral CD8 TIL score is a favorable prognostic factor independent of well-established AJCC staging factors.

Is the presence of tumor-infiltrating lymphocytes predictive of outcomes in patients with melanoma?

The significance of tumor-infiltrating lymphocytes (TILs) in melanoma is debated. This article presents a multicenter, retrospective study assessing the predictive and prognostic value of TILs. The Sentinel Lymph Node Working Group database was queried from 1993 to 2018 for cases with known TIL data. TILs were categorized as absent or present, which included nonbrisk (NB), brisk (B), and present but unspecified TIL levels. Clinicopathologic factors were correlated with TILs, sentinel lymph node (SLN) status, and melanoma-specific survival (MSS). Overall, 3203 patients were included. The median thickness was 1.5 mm, and 469 cases had SLN metastases. TILs were present in 2458 cases (76.7%), with NB, B, and unspecified TILs seen in 1691 (68.8%), 691 (28.1%), and 76 (3.1%), respectively. Multivariable analysis showed that the presence of TILs significantly predicted a negative SLN biopsy (P < .05). The median follow-up was 25.2 months. MSS was significantly better for cases with TILs than cases without TILs (P < .001). According to multivariable analysis, age, gender, thickness, mitotic rate, ulceration, lymphovascular invasion, and SLN status were significantly prognostic of MSS (all P values < .05). Although TILs were not prognostic of MSS, when multiple imputation was used and the SLN status was excluded, the presence of TILs was significantly prognostic of improved MSS (hazard ratio, 0.78; 95% confidence interval, 0.64-0.95; P = .0154). TILs are a favorable marker because their presence significantly predicts a negative SLN, and the absence of TILs may be a prognostic marker of worse survival in patients with a positive SLN but not a negative SLN. TILs may also serve as a prognostic marker of survival when the SLN status is not considered.

Assessing the Prognostic Significance of Tumor-Infiltrating Lymphocytes in Patients With Melanoma Using Pathologic Features Identified by Natural Language Processing

Although tumor-infiltrating lymphocytes (TILs) are an important histopathologic characteristic reflecting host immune response in patients with melanoma, their prognostic value remains controversial. Because manual review of medical records is labor intensive, a survival analysis using a large patient cohort with comprehensive clinical and histopathologic characteristics is lacking. To assess the prognostic significance of TILs among patients with cutaneous melanoma using a large cohort established through natural language processing (NLP) algorithms. This retrospective cohort study analyzed the medical records of 14 436 patients with cutaneous melanoma at Brigham and Women's Hospital between June 1, 2004, and December 31, 2019. Patients were followed up to death or censored at their last clinical visit. The primary outcome was overall survival (OS). Survival analysis was conducted using Kaplan-Meier curves, the log-rank test, and Cox proportional hazards regression analysis. A total of 14 436 patients with cutaneous melanoma were identified in the institution's pathology information system. Using NLP, we established a study cohort of 2624 patients (1462 men [55.7%]; median age, 61 years [interquartile range, 50-72 years]) who had vertical growth phase melanoma with TIL status scored. Absent TILs were identified in 434 patients (16.5%), nonbrisk TILs in 1916 patients (73.0%), and brisk TILs in 274 patients (10.4%). The 5-year survival rate was 71.0% (95% CI, 65.5%-76.9%) among patients with an absence of TILs, 73.8% (95% CI, 71.1%-76.5%) among patients with nonbrisk TILs, and 85.2% (95% CI, 80.0%-90.7%) among patients with brisk TILs. Brisk TILs were significantly associated with improved OS (adjusted hazard ratio, 0.63; 95% CI, 0.42-0.95; P = .03; 14.2% OS advantage at 5 years), and nonbrisk TILs were not associated with improved OS (adjusted hazard ratio, 0.87; 95% CI, 0.68-1.11; P = .25), compared with the absence of TILs. This study provides evidence based on a large patient cohort from a single institution that suggests that brisk TILs represent an independent prognostic factor for OS among patients with primary cutaneous melanoma. The study also suggests that NLP is a highly efficient tool to facilitate large-scale analyses that involve free-text clinical data.

Prognostic Value of Tumor-Infiltrating Lymphocytes in Sinonasal Mucosal Melanoma.

Tumor-infiltrating lymphocytes (TILs) predict better outcome in several types of cancers. However, the prognostic value of TILs in sinonasal mucosal melanoma (SNMM) is uncertain. Here, we investigated whether TILs can be used as a prognostic indicator for survival in SNMM. Retrospective cohort study. Patient history and histologic specimens from 27 patients with primary SNMM were retrospectively analyzed. TIL grade was determined and associations between TILs and AJCC tumor stage, overall survival, and recurrence-free survival were analyzed. Patients with TILs in the primary tumor classified as brisk or non-brisk survived significantly longer than patients with SNMMs lacking lymphocyte infiltrates. Brisk TILs were associated with the lower T3 stage and increased recurrence-free and 5-year survival. Our results indicate that TIL density is a strong prognostic factor for better survival in SNMM. Prospective studies with larger case numbers are warranted to determine whether TILs should be included in future AJCC staging guidelines. 3 Laryngoscope, 132:1334-1339, 2022.

A Retrospective, Observational Analysis of Tumor Infiltrating Lymphocytes and Tumor Regression in Melanoma

Tumor infiltrating lymphocytes (TILs) and regression are thought to be distinct markers of the immune response to melanoma. This study sought to analyze the relationship of TIL grade and presence of regression to each other and to other prognostic histopathologic and clinical values in melanoma. A retrospective analysis was conducted using patients diagnosed with melanoma between 2013 and 2019 whose complete histopathologic reports were available. Regression was seen in 48.9%, 30.1% and 37.9% of patients with brisk, non-brisk, and absent TILs respectively (P=0.019). Melanoma tumors with brisk TILs were found to have a lower Breslow thickness than those with non-brisk or absent (P= 0.001). Tumors with regression were also found to have lower Breslow thickness (P<0.001). Neither TIL grade nor regression were protective of nodal metastasis or associated with improved survival. Brisk TILs have a positive association with thinner tumors and the presence of tumor regression relative to non-brisk or absent TILs. This may suggest a more robust immune response in tumors with brisk TILs. Further exploration of the interplay between TIL grade, lymphocyte cell subtype and lymphocyte density may help explain this finding.

Transcriptomic profiling of peripheral blood mononuclear cells in patients with multiple primary melanoma.

e21558 Background: Patients (PTs) with cutaneous melanoma (MEL) are at increased risk of developing subsequent MEL. The primary objective of the present study was to determine whether the peripheral blood gene expression profile (GEP) of PTs with multiple primary melanoma (MPM) differs from that of patients with single primary melanoma (SPM) and healthy controls (HCs). A secondary objective is to assess if the peripheral blood immune transcriptional profile varies in relation with clinical and immunopathological characteristics of MEL. Methods: Cases with MPM, SPM and HCs were retrieved from the University of Pittsburgh Human Biological Sample and Nevus Image Banking and Analysis Repository. NanoString PanCancer Immune Profiling of 770-immune related genes was used to define GEP in peripheral blood mononuclear cells (PBMCs). Quality control (QC), data normalization, and differential expression analyses were performed using the ROSALIND Analysis System. Genes with fold change ≥1.5 and p-value &lt; 0.05 were considered as differentially-expressed between cohorts. Results: In total, 47 PTs including 10 with SPM, 40 with MPM, and 10 HCs, were included. GEP data from 3 cases did not meet QC requirements and were discarded. The majority of PTs had stage I/II disease (87.2%). The median Breslow thickness was 1 mm, ulceration was present in 6 (12.2%) MELs. Tumor-infiltrating lymphocytes (TILs) were absent, non-brisk, and brisk in 20%, 65%, 15% of cases, respectively. The expression of IFIT1, MX1, ISG15 was decreased in PTs with MPM in comparison to HCs. In comparison to SPMs, PTs with MPM demonstrated downregulation of B-cell receptor signaling pathway genes (CD19, CD22, CD79A, BLK). 14 genes involved in TNF-alpha effects on cytokine activity, cell motility, and apoptosis, and 12 genes in the IL18 signaling pathway were differentially expressed in PTs with Breslow thickness of &gt;1mm vs ≤1mm. The top downregulated genes in &gt;1mm thick MELs were CD83, VEGFA and CXCL3. Ulcerated MEL had decreased expression of CCL2, CCL3, CCL3L1, CCL4. Stage III MEL had decreased expression of GZMH, MHC Class II genes HLA-DMB, HLA-DRB3, lectin receptor family genes KLRC2 and KLRD1, as well as Type-1 immune-associated biomarkers IRF1, SYK, TBX21 in comparison to Stage I/II cases. Tumors deficient in TILs exhibited decreased expression of Granzyme A and B, T-box 21, and IL18RAP (enhances IL18R-mediated signaling) in peripheral blood. Tumors with brisk TILs had increased expression of the B-cell associated transcripts CD19, CD22, CD79A in peripheral blood. Conclusions: The GEP of PBMC's from PTs with MPM was similar overall to that of PTs with SPM/HC, with only a subset of genes exhibiting differentially expression in PTs with MPM vs. PTs with SPM and HCs. GEP varied in relation to Breslow thickness, stage, ulceration, and TIL content in MELs.

Intratumoral T-Lymphocyte Subsets in Canine Oral Melanoma and Their Association With Clinical and Histopathological Parameters.

Canine oral melanoma is a common, aggressive tumor with limited treatment options. Tumor-infiltrating lymphocytes (TILs) are important in antitumor immunity. This study used histopathology and immunophenotyping by flow cytometry to evaluate the presence and distribution of TILs in canine oral melanoma, including the frequency of CD8+ T cells, CD4+ T cells, and regulatory T cells. Fifty samples of oral melanoma from 45 dogs that did not receive treatment prior to surgery were included in the study. The distribution of TILs in the tissue (brisk, nonbrisk, and absent) was evaluated in 48 samples. Twenty-eight (58%) samples had a brisk distribution pattern, 10 (21%) samples had a nonbrisk pattern, and 10 (21%) samples had an absent TIL pattern. Comparing the histological evaluation and the immunophenotyping data, it was observed that samples with a brisk TIL pattern had a higher frequency of CD8+ T lymphocytes (P = .05) and a lower frequency of CD4+/CD25+/FoxP3+ Tregs (P = .03), compared to the samples with nonbrisk and absent infiltrate patterns. Patients with a higher survival rate had higher TIL scores (P = .002), a brisk or nonbrisk TIL pattern (P = .001), and an increased frequency of CD8+ T lymphocytes infiltrating the tumor (P = .003). Our analysis suggests that the evaluation of TILs in canine oral melanoma is relevant to predict tumor aggressiveness and patient prognosis.

Brain Tumor Microenvironment and Angiogenesis in Melanoma Brain Metastases.

BackgroundHigh tumor-infiltrating lymphocytes (TILs) and hemorrhage are important prognostic factors in patients who have undergone craniotomy for melanoma brain metastases (MBM) before 2011 at the University of Pittsburgh Medical Center (UPMC). We have investigated the prognostic or predictive role of these histopathologic factors in a more contemporary craniotomy cohort from the University of North Carolina at Chapel Hill (UNC-CH). We have also sought to understand better how various immune cell subsets, angiogenic factors, and blood vessels may be associated with clinical and radiographic features in MBM.MethodsBrain tumors from the UPMC and UNC-CH patient cohorts were (re)analyzed by standard histopathology, tumor tissue imaging, and gene expression profiling. Variables were associated with overall survival (OS) and radiographic features.ResultsThe patient subgroup with high TILs in craniotomy specimens and subsequent treatment with immune checkpoint inhibitors (ICIs, n=7) trended to have longer OS compared to the subgroup with high TILs and no treatment with ICIs (n=11, p=0.059). Bleeding was significantly associated with tumor volume before craniotomy, high melanoma-specific expression of basic fibroblast growth factor (bFGF), and high density of CD31+αSMA- blood vessels. Brain tumors with high versus low peritumoral edema before craniotomy had low (17%) versus high (41%) incidence of brisk TILs. Melanoma-specific expression of the vascular endothelial growth factor (VEGF) was comparable to VEGF expression by TILs and was not associated with any particular prognostic, radiographic, or histopathologic features. A gene signature associated with gamma delta (gd) T cells was significantly higher in intracranial than same-patient extracranial metastases and primary melanoma. However, gdT cell density in MBM was not prognostic.ConclusionsICIs may provide greater clinical benefit in patients with brisk TILs in MBM. Intratumoral hemorrhage in brain metastases, a significant clinical problem, is not merely associated with tumor volume but also with underlying biology. bFGF may be an essential pathway to target. VEGF, a factor principally associated with peritumoral edema, is not only produced by melanoma cells but also by TILs. Therefore, suppressing low-grade peritumoral edema using corticosteroids may harm TIL function in 41% of cases. Ongoing clinical trials targeting VEGF in MBM may predict a lack of unfavorable impacts on TIL density and/or intratumoral hemorrhage.

Genotype-phenotype associations in colorectal adenocarcinomas and their matched metastases

Although primary colorectal carcinomas (CRCs) frequently share genetic alterations with their metastases, morphologic surrogates reflecting the genotype contexture of metastases remain largely unknown. We investigated phenotype/genotype associations in paired primary and metastatic colorectal adenocarcinomas from 75 patients. Thirty-three (44%) metastatic lesions were synchronous and 42 (56%) were metachronous. Tumor budding, micronecrosis, and tumor-infiltrating lymphocyte (TIL) density were compared with matched next-generation sequencing genotypes. Micronecrosis in the primary were significantly associated with nodal status (P=0.0054) and with micronecrosis in metastatic sites (P=0.0216), particularly in metachronous metastases (P=0.0033). With a 57-gene panel, one or more mutations were identified in 64 (85.3%) cases. In metastases, high (brisk) TILs were associated with overall mutational burden (P=0.0058) and with mutations in EGF (P=0.0325), RAS genes (P=0.0043), and MMR genes (P=0.0069), whereas high-level micronecrosis correlated with mutations in APC (P=0.0004) and MSH6 (P=0.0385) genes. Genomic alterations were shared in 90.1% of primary/metastatic pairs, but clonality of the same mutation was shared in only 57.1% of paired lesions. Compared with synchronous, metachronous metastases had more private clonal alterations (P=0.0291); in this group, clonal alterations coincided with brisk TILs (P=0.0334) and high micronecrosis (P=0.0133). High TILs in metastatic lesions were predictive of favorable overall survival (log-rank P=0.044). The observed phenotype/genotype associations favor the clonal evolution model in CRC metastases that seems accompanied by intense host immune response. If the role of micronecrosis and brisk TILs in metachronous metastases is validated in larger studies, these histologic parameters will be worth adding in the armamentarium for the evaluation of metastatic CRC.

Impact of Tumor-Infiltrating Lymphocytes on Overall Survival in Merkel Cell Carcinoma.

Merkel cell carcinoma (MCC) is a rare and aggressive neuroendocrine carcinoma of the skin. As the clinical course can be variable, prognostic markers are needed to better stratify patients. Prior literature, composed of small series with limited sample size, has demonstrated that tumor-infiltrating lymphocytes (TILs) are an important prognostic marker in MCC. To validate these findings on a population level, we sought to analyze and report the prognostic value of TILs in a large national data set. A retrospective observational cohort study was conducted of patients with nonmetastatic MCC from 2010 to 2015 using the National Cancer Database. Individual variables trending toward significance using a univariable analysis were included in a multivariable Cox proportional hazards model to assess their independent effect on overall survival (OS). TILs were subclassified into none, nonbrisk, and brisk and the survival analysis was performed. Propensity score-weighted multivariable analysis (PS MVA) was performed to adjust for additional confounding. A total of 2,182 patients met inclusion criteria: 611 (28.0%) were identified as having TILs present, and 1,571 (72.0%) had TILs absent in the tumor. On MVA, subdivision of TIL status into nonbrisk (hazard ratio [HR], 0.750; 95% confidence interval [CI], 0.602-0.933) and brisk (HR, 0.499; 95% CI, 0.338-0.735) was associated with incrementally improved OS compared with no TILs. The association of nonbrisk and brisk TILs with improved OS was retained on PS MVA (Nonbrisk: HR, 0.720; 95% CI, 0.550-0.944; Brisk: HR, 0.483; 95% CI, 0.286-0.814). The presence of nonbrisk and brisk TILs is associated with incrementally improved OS in patients with nonmetastatic MCC in a large national data set. This pathologic feature can aid with risk stratification, estimation of prognosis, and, importantly, decision-making with respect to treatment intensification in high-risk patients. Merkel cell carcinoma (MCC) is an aggressive neuroendocrine cutaneous malignancy with variable clinical course. Prognostic markers are needed to better risk stratify patients. We present the largest retrospective observational cohort study of patients with nonmetastatic MCC using the National Cancer Database. Our analysis demonstrates an association between increasing degrees of tumor-infiltrating lymphocytes and incrementally improved survival. These conclusions improve pathologic risk stratification, and decision-making with respect to treatment intensification. Intensification may include adjuvant radiation therapy to the primary site after wide excision despite small tumor size, to the nodal basin in sentinel lymph node-negative patients, or offering closer follow-up.

Tumor infiltrating lymphocyte grade in Merkel cell carcinoma: relationships with clinical factors and independent prognostic value

Introduction Surrogate markers of the host immune response are not currently included in AJCC staging for Merkel cell carcinoma (MCC), and have not been consistently associated with clinical outcomes. We performed an analysis of a large national database to investigate tumor infiltrating lymphocyte (TIL) grade as an independent predictor of overall survival (OS) for patients with MCC and to characterize the relationship between TIL grade and other clinical prognostic factors. Material and methods The NCDB was queried for patients with resected, non-metastatic MCC with known TIL grade (absent, non-brisk and brisk). Multivariable Cox regression modeling was performed to define TIL grade as a predictor of OS adjusting for other relevant clinical factors. Multinomial, multivariable logistic regression was performed to characterize the relationship between TIL grade and other clinical prognostic factors. Multiple imputation was performed to account for missing data bias. Results Both brisk (HR 0.55, CI 0.36–0.83) and non-brisk (HR 0.77, CI 0.60–0.98) were associated with decreased adjusted hazard of death relative to absent TIL grade. Adverse clinical factors such as 1–3 positive lymph nodes, lymphovascular invasion (LVI) and immunosuppression were associated with increased likelihood of non-brisk TIL relative to absent TIL grade (p values <.05). Extracapsular extension (ECS) was associated with decreased likelihood of brisk TIL relative to absent TIL grade (p<.05). Discussion Histopathologic TIL grade was independently predictive for OS in this large national cohort. Significant differences in the likelihood of non-brisk or brisk TIL relative to absent grade were present with regards to LVI, ECS and immune status. TIL grade may be a useful prognostic factor to consider in addition to more granular characterization of TIL morphology and immunophenotype.