Background: T cell density in colorectal cancer (CRC) has proven to be of high prognostic importance. Here, we evaluated the influence of a hyperfractionated preoperative short-term radiation protocol (25 Gy) on immune cell density in tumor samples of rectal cancer (RC) patients and on patient survival. In addition, we assessed spatial tumor heterogeneity by comparison of analogue T cell quantification on full tissue sections with digital T cell quantification on a virtually established tissue microarray (TMA). Methods: A total of 75 RC patients (60 irradiated, 15 treatment-naïve) were defined for retrospective analysis. RC samples were processed for immunohistochemistry (CD3, CD8, PD-1, PD-L1). Analogue (score 0–3) as well as digital quantification (TMA: 2 cores vs. 6 cores, mean T cell count) of marker expression in 2 areas (central tumor, CT; invasive margin, IM) was performed. Survival was estimated on the basis of analogue as well as digital marker densities calculated from 2 cores (Immunoscore: CD3/CD8 ratio) and 6 cores per tumor area. Results: Irradiated RC samples showed a significant decrease in CD3 and CD8 positive T cells, independent of quantification mode. T cell densities of 6 virtual cores approximated to T cell densities of full tissue sections, independent of individual core density or location. Survival analysis based on full tissue section quantification demonstrated that CD3 and CD8 positive T cells as well as PD-1 positive tumor infiltrating leucocytes (TILs) in the CT and the IM had a significant impact on disease-free survival (DFS) as well as overall survival (OS). In addition, CD3 and CD8 positive T cells as well as PD-1 positive TILs in the IM proved as independent prognostic factors for DFS and OS; in the CT, PD-1 positive TILs predicted DFS and CD3 and CD8 positive T cells as well as PD-1 positive TILs predicted OS. Survival analysis based on virtual TMA showed no impact on DFS or OS. Conclusion: Spatial tumor heterogeneity might result in inadequate quantification of immune marker expression; however, if using a TMA, 6 cores per tumor area and patient sample represent comparable amounts of T cell densities to those quantified on full tissue sections. Consistently, the tissue area used for immune marker quantification represents a crucial factor for the evaluation of prognostic and predictive biomarker potential.

AF1q impairs survival in hematologic and solid malignancies. AF1q expression is associated with tumor progression, migration and chemoresistance and acts as a transcriptional co-activator in WNT and STAT signaling. This study evaluates the role of AF1q in patients with resectable esophageal cancer (EC). A total of 278 patients operated on for EC were retrospectively included and the expression of AF1q, CD44 and pYSTAT3 was analyzed following immunostaining. Quantified data were processed to correlational and survival analysis. In EC tissue samples, an elevated expression of AF1q was associated with the expression of CD44 (p = 0.004) and pYSTAT3 (p = 0.0002). High AF1q expression in primary tumors showed high AF1q expression in the corresponding lymph nodes (p = 0.016). AF1q expression was higher after neoadjuvant therapy (p = 0.0002). Patients with AF1q-positive EC relapsed and died earlier compared to patients with AF1q-negative EC (disease-free survival (DFS), p = 0.0005; disease-specific survival (DSS), p = 0.003); in the multivariable Cox regression model, AF1q proved to be an independent prognostic marker (DFS, p = 0.01; DSS, p = 0.03). AF1q is associated with WNT and STAT signaling; it impairs and independently predicts DFS and DSS in patients with resectable EC. Testing AF1q could facilitate prognosis estimation and provide a possibility of identifying the patients responsive to the therapeutic blockade of its oncogenic downstream targets.

Thyroid hormone levels may be associated with disease outcome in head and neck squamous cell carcinoma (HNSCC). µ-Crystallin (CRYM), a thyroid hormone binding protein, blocks intracellular binding of the thyroid hormone T3 to its receptors. In this study, we aimed to analyze the association of CRYM levels with disease outcome in HNSCC patients. We retrospectively assessed immunohistochemical CRYM expression in 121 head and neck cancer patients. Preoperative thyrotropin levels could be extracted for 50 patients. Patients with low thyrotropin levels had a worse prognosis compared to euthyroid patients (5-year overall survival TSH low 20% vs. TSH norm 58%). We observed an association of CRYM+ patients with improved overall survival (5-year overall survival for CRYM+ 78.6% vs. CRYM− 56%). Interaction analysis between CRYM and HPV revealed that this effect was limited to HPV− patients (CRYM+|HPV− HR 0.12, 95% CI 0.01–0.87, p = 0.036). These results were replicated in an independent dataset. CRYM expression identified patients with favorable disease progression for HPV− HNSCC patients and could serve as a useful biomarker in this patient population. This study further confirms a correlation of thyroid hormone levels with adverse disease outcome in HNSCC patients, which could be potentially exploited as a therapeutic target.

The proteins sodium iodide symporter (NIS), μ-crystallin (CRYM), and thyroid hormone receptor beta (THRB) have been associated with prognosis in various cancer entities. While NIS and THRB may serve as possible therapeutic targets, the role of CRYM in cancer is still unclear. Protein levels of 44 patients with adenoid cystic carcinoma of the head and neck were analyzed using immunohistochemistry and correlated with clinicopathological data and outcome. NIS was positive in 72%, CRYM was positive in 55%, and THRB was positive in 39% of the patients. CRYM-positive adenoid cystic carcinomas were associated with a better cause-specific survival. Thus, our data indicate that CRYM might be a suitable positive prognostic marker in adenoid cystic carcinoma of the head and neck. Furthermore, expression of NIS was present in most patients and therefore evaluation of the use of radioiodine treatment is recommended.

Background/Objectives: Fibroblast activation protein (FAP) has gained tremendous traction as a target for tumor imaging and cancer treatment, while also playing a key role in fibrosis. Our study aimed to evaluate [⁶⁸Ga]Ga-DATA^5m.SA.FAPi for PET imaging of replacement fibrosis following myocardial infarction (MI) or interstitial fibrosis associated with hypertrophy. Methods: MI or transverse aortic constriction (TAC)-induced hypertrophy was induced in C57BL/6 mice, with sham-operated animals serving as controls. At multiple time points during disease progression (1, 2, and 6 weeks post-surgery), [⁶⁸Ga]Ga-DATA^5m.SA.FAPi PET/CT scans were performed, followed by ex vivo investigations. Additionally, in vitro cell uptake experiments simulating hypertrophy were conducted. Results: Cardiac uptake of [⁶⁸Ga]Ga-DATA5m.SA.FAPi significantly increased two weeks after MI induction (MI: 2.1 ± 0.2%ID/g, n = 7 vs. SHAM: 1.1 ± 0.2%ID/g, n = 5; p = 0.002), confirmed by ex vivo autoradiography. No significant difference was observed at six weeks post-MI (MI: 1.1 ± 0.1%ID/g, n = 4 vs. SHAM: 0.8 ± 0.0%ID/g, n = 3), indicating infarct healing completion. In contrast, TAC mice showed increased uptake after six weeks (TAC: 1.8 ± 0.2%ID/g, n = 6; p = 0.007), related to interstitial fibrosis progression. Consistently, high-stretched cardiac fibroblasts demonstrated a higher uptake compared to low-stretched conditioned ones, suggesting the stretch mediates regulation of FAP. Conclusions: This study demonstrated the efficacy of [⁶⁸Ga]Ga-DATA^5m.SA.FAPi for longitudinal imaging of cardiac fibrosis in response to different cardiac injuries. In vivo FAP imaging during cardiac remodeling may serve as a valuable tool for diagnosing and predicting disease progression, ultimately aiding in the clinical management of patients.

Colorectal cancer liver metastasis (CRLM) is a major clinical problem. The regulators of immunosurveillance of CRLM could hold potential for developing therapeutic strategies to prevent or treat metastasis. In this study, using a murine colorectal cancer organoid-based transplantation model, we identified TYK2 as a key factor controlling CRLM. Evaluation of the effects of Tyk2 deletion in different subsets of immune cells and in colorectal cancer cells demonstrated that TYK2 was not required in cancer cells, macrophages, NK cells, T cells, or Kupffer cells. Instead, TYK2 controlled CRLM via a dendritic cell–dependent mechanism that relied on MHC-I–mediated cross-presentation of antigens to CD8+ T cells. Analysis of single-cell RNA sequencing data from primary colorectal cancer and CRLM revealed that TYK2 was predominantly expressed in a dendritic cell population destined to present antigens in tumor-draining lymph nodes. Treatment with the TYK2 inhibitor deucravacitinib, which is approved by the FDA for treating plaque psoriasis and is under clinical investigation for other autoimmune diseases, promoted CRLM. Together, these data demonstrate that TYK2 controls CRLM immunosurveillance, which should be carefully considered when treating patients with TYK2 inhibitors. Significance: TYK2 restricts the metastasis of colorectal tumors to the liver by supporting dendritic cell-dependent induction of antitumor CD8+ T cells, which could impact the use of TYK2 inhibitors in patients.