Therapies that boost the anti-tumor responses of cytotoxic T lymphocytes (CTLs) have shown promise; however, clinical responses to the immunotherapeutic agents currently available vary considerably, and the molecular basis of this is unclear. We performed transcriptomic profiling of tumor-infiltrating CTLs from treatment-naive patients with lung cancer to define the molecular features associated with the robustness of anti-tumor immune responses. We observed considerable heterogeneity in the expression of molecules associated with activation of the T cell antigen receptor (TCR) and of immunological-checkpoint molecules such as 4-1BB, PD-1 and TIM-3. Tumors with a high density of CTLs showed enrichment for transcripts linked to tissue-resident memory cells (TRM cells), such as CD103, and CTLs from CD103hi tumors displayed features of enhanced cytotoxicity. A greater density of TRM cells in tumors was predictive of a better survival outcome in lung cancer, and this effect was independent of that conferred by CTL density. Here we define the ‘molecular fingerprint’ of tumor-infiltrating CTLs and identify potentially new targets for immunotherapy.

Objective-Hereditary hemorrhagic telangiectasia is a genetic disorder characterized by visceral and mucocutaneous arteriovenous malformations (AVMs). Clinically indistinguishable hereditary hemorrhagic telangiectasia 1 and hereditary hemorrhagic telangiectasia 2 are caused by mutations in ENG and ALK1, respectively. In this study, we have compared the development of visceral and mucocutaneous AVMs in adult stages between Eng-and Alk1-inducible knockout (iKO) models. Approach and Results-Eng or Alk1 were deleted from either vascular endothelial cells (ECs) or smooth muscle cells in adult stages using Scl-CreER and Myh11-CreER lines, respectively. Latex perfusion and intravital spectral imaging in a dorsal skinfold window chamber system were used to visualize remodeling vasculature during AVM formation. Global Eng deletion resulted in lethality with visceral AVMs and wound-induced skin AVMs. Deletion of Alk1 or Eng in ECs, but not in smooth muscle cells, resulted in wound-induced skin AVMs. Visceral AVMs were observed in EC-specific Alk1-iKO but not in Eng-iKO. Intravital spectral imaging revealed that Eng-iKO model exhibited more dynamic processes for AVM development when compared with Alk1-iKO model. In this study, we examined the effect of wounding in the development of mucocutaneous telangiectases and investigated the cellular origin of de novo AVMs in both HHT1 and HHT2 mouse models. Conclusions-Both Materials and MethodsMaterials and Methods are available in the online-only Supplement. ResultsTo investigate the role of ENG in adult mice, Eng was globally deleted using the tamoxifen-inducible ROSA26 CreER mouse strain (R26 CreER ). 10 We found that 3 consecutive day injection of tamoxifen at 2.5 mg/25 g body weight was the most effective regimen for R26CreER/+ ;Eng 2f/2f mice (Figure I in the online-only Data Supplement). In 3 to 4 days after the first tamoxifen injection, the R26 CreER/+ ;Eng 2f/2f mutant mice displayed signs of illness, such as slow movement, diarrhea, and dehydration, and died around day 4 to 10 (n>30). To analyze the subdermal vessels in the back skin, vascular casting with blue latex was performed at days 5 to 8 by infusing it into the left ventricle. Subdermal vessels were unaffected in the intact back skins of tamoxifen-injected adult R26CreER/+ ;Eng 2f/2f mice (data not shown) and also wounded skin of tamoxifen-injected adult R26CreER/+ ;Eng 2f/+ mice ( Figure IIA in the online-only Data Supplement; n=6). However, areas of wounds in mid-dorsum and ear of tamoxifen-injected R26 CreER/+ ;Eng 2f/2f mice showed dilated and tortuous vessels, and the latex dye was found in both arteries and veins, indicating the presence of AV shunts ( Figure IIB in the online-only Data Supplement; n=13). However, blood vessels away from the wound in Eng-iKOs had normal morphology and latex only in arterial branches.To determine the vascular cell type where ENG plays a critical role for the development of the vascular network at adult stages, we used 2 cell-type-specific inducible Cre lines: Scl-CreER 11 fo...