There is increasing evidence that tertiary lymphoid structures (TLS) control not only local adaptive B cell responses at melanoma tumor sites but also the cellular composition and function of other immune cells. In human melanoma, however, a comprehensive analysis of TLS phenotypes, density and spatial distribution at different disease stages is lacking. Here we used 7-color multiplex immunostaining of whole tissue sections from 103 human melanoma samples to characterize TLS phenotypes along the expression of established TLS-defining molecular and cellular components. TLS density and spatial distribution were determined by referring TLS counts to the tissue area within defined intra- and extratumoral perimeters around the invasive tumor front. We show that only a subgroup of primary human melanomas contains TLS. These TLS rarely formed germinal centers (GCs) and mostly located intratumorally within 1 mm distance to the invasive tumor front. In contrast, melanoma metastases had a significantly increased density of secondary follicular TLS. They appeared preferentially in stromal areas within an extratumoral 1 mm distance to the invasive tumor front and their density varied over time and site of metastasis. Interestingly, secondary follicular TLS in melanoma often lacked BCL6+ lymphatic cells and GC polarity. Our work provides an integrated qualitative and quantitative analysis of TLS in human melanoma and shows disease progression- and site-associated changes in TLS phenotypes, density and spatial distribution. The frequent absence of GC polarity in melanoma TLS highlights the induction of TLS maturation as a potential additive to future immunotherapy studies. Given variable evaluation strategies in former TLS studies, this report demonstrates a highly reproducible, standardized quantitative evaluation approach.

ESDR265 - A standardized analysis of tertiary lymphoid structures in human melanoma: disease progression- and tumor site-associated changes with germinal center alteration

50th Annual Meeting of the European Society for Dermatological Research. VIRTUAL MEETING