SummaryLymphoid nodules are a normal component of the mucosa of the rectum, but little is known about their function and whether they contribute to the host immune response in malignancy. In rectal cancer specimens from patients with local (n = 18), regional (n = 12) and distant (n = 10) disease, we quantified T cell (CD3, CD25) and dendritic cell (CD1a, CD83) levels at the tumour margin as well as within tumour-associated lymphoid nodules. In normal tissue CD3+ , but not CD25 + , T cells are concentrated at high levels within lymphoid nodules, with significantly fewer cells found in surrounding normal mucosa (P = 0·001). Mature (CD83), but not immature (CD1a), dendritic cells in normal tissue are also found clustered almost exclusively within lymphoid nodules (P = < 0·0001). In rectal tumours, both CD3+ T cells (P = 0·004) and CD83+ dendritic cells (P = 0·0001) are also localized preferentially within tumour-associated lymphoid nodules. However, when comparing tumour specimens to normal rectal tissue, the average density of CD3 + T cells (P = 0·0005) and CD83+ dendritic cells (P = 0·0006) in tumourassociated lymphoid nodules was significantly less than that seen in lymphoid nodules in normal mucosa. Interestingly, regardless of where quantified, T cell and dendritic cell levels did not depend upon the stage of disease. Increased CD3 + T cell infiltration of tumour-associated lymphoid nodules predicted improved survival, independent of stage (P = 0·05). Other T cell (CD25) markers and different levels of CD1a + or CD83 + dendritic cells did not predict survival. Tumour-associated lymphoid nodules, enriched in dendritic cells and T cells, may be an important site for antigen presentation and increased T cell infiltration may be a marker for improved survival.

Objective: The purpose of this study was to investigate whether somatic nonsynonymous variants in tumor tissue can potentially be identified in circulating cellfree DNA (cfDNA) of head and neck oropharyngeal squamous cell carcinoma (OPSCC) patients using next-generation sequencing and can predict recurrence or persistence disease. Methods: A total of 22 OPSCC patients with tumor tissue and respective plasma samples were included in this study. Matching cfDNA and tumor tissues were processed, and DNA sequencing was conducted using the MiSeq platform. Variants were identified using Biomedical Genomic Workbench and Genialis's online data analysis platform for Swift Biosciences' Accel-amplicon panels. Results: Among 11 nonresponders, 6 matched mutations were detected in 5 patients suggesting a predictive factor for patients with likelihood of recurrence. The matched variants and their allele frequencies identified in the nonresponder group were (tumor DNA/cfDNA in %): TP53 G325fs (27/0.62), TP53 R282W (48/1.74), TP53 R273C (39/2.17), FBXW7 R505G (30/0.6), FBXW7 R505L (31/0.65), and TP53 Q331H (56.5/0.52). Interestingly, the matched somatic mutations were only detected in patients who did not respond to therapy or had persistent disease. Conclusions: Somatic nonsynonymous variants in tumor tissue can potentially be identified in cfDNA of OPSCC patients using NGS. The likelihood of variant detection in cfDNA is greater in nonresponders, especially in human papillomavirus-negative nonresponders, rendering it beneficial as a less invasive detection method for disease persistence/recurrence and prognosis.

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