The probabilistic model of Alzheimer disease: the amyloid hypothesis revised.

The current conceptualization of Alzheimer disease (AD) is driven by the amyloid hypothesis, in which a deterministic chain of events leads from amyloid deposition and then tau deposition to neurodegeneration and progressive cognitive impairment. This model fits autosomal dominant AD but is less applicable to sporadic AD. Owing to emerging information regarding the complex biology of AD and the challenges of developing amyloid-targeting drugs, the amyloid hypothesis needs to be reconsidered. Here we propose a probabilistic model of AD in which three variants of AD (autosomal dominant AD, APOE ε4-related sporadic AD and APOE ε4-unrelated sporadic AD) feature decreasing penetrance and decreasing weight of the amyloid pathophysiological cascade, and increasing weight of stochastic factors (environmental exposures and lower-risk genes). Together, these variants account for a large share of the neuropathological and clinical variability observed in people with AD. The implementation of this model in research might lead to a better understanding of disease pathophysiology, a revision of the current clinical taxonomy and accelerated development of strategies to prevent and treat AD.

Clonal expansion of intra-epithelial T cells in breast cancer revealed by spatial transcriptomics.

The spatial distribution of tumor-infiltrating lymphocytes (TIL) predicts breast cancer outcome and response to systemic therapy, highlighting the importance of an intact tissue structure for characterizing tumors. Here, we present ST-FFPE, a spatial transcriptomics method for the analysis of formalin-fixed paraffin-embedded samples, which opens the possibility of interrogating archival tissue. The method involves extraction, exome capture and sequencing of RNA from different tumor compartments microdissected by laser-capture, and can be used to study the cellular composition of tumor microenvironment. Focusing on triple-negative breast cancer (TNBC), we characterized T cells, B cells, dendritic cells, fibroblasts and endothelial cells in both stromal and intra-epithelial compartments. We found a highly variable spatial distribution of immune cell subsets among tumors. This analysis revealed that the immune repertoires of intra-epithelial T and B cells were consistently less diverse and more clonal than those of stromal T and B cells. T-cell receptor (TCR) sequencing confirmed a reduced diversity and higher clonality of intra-epithelial T cells relative to the corresponding stromal T cells. Analysis of the top 10 dominant clonotypes in the two compartments showed a majority of shared but also some unique clonotypes both in stromal and intra-epithelial T cells. Hyperexpanded clonotypes were more abundant among intra-epithelial than stromal T cells. These findings validate the ST-FFPE method and suggest an accumulation of antigen-specific T cells within tumor core. Because ST-FFPE is applicable for analysis of previously collected tissue samples, it could be useful for rapid assessment of intratumoral cellular heterogeneity in multiple disease and treatment settings.

Linking unfolded protein response to ovarian cancer cell fusion.

BACKGROUND: Polyploid giant cancer cells (PGCCs) have been observed in epithelial ovarian tumors. They can resist antimitotic drugs, thus participating in tumor maintenance and recurrence. Although their origin remains unclear, PGCC formation seems to be enhanced by conditions that trigger the unfolded protein response (UPR) such as hypoxia or chemotherapeutic drugs like paclitaxel. Hypoxia has been shown to promote the formation of ovarian PGCCs by cell fusion. We thus hypothesized that the UPR could be involved in EOC cell fusion, possibly explaining the occurrence of PGCCs and the aggressiveness of EOC. METHODS: The UPR was induced in two ovarian cancer cell lines (SKOV3 and COV318). The UPR activation was assessed by Western blot and polyploidy indexes were calculated. Then, to confirm the implication of cell fusion in PGCC formation, two populations of SKOV3 cells were transfected with plasmids encoding for two distinct nuclear fluorescent proteins (GFP and mCherry) associated with different antibiotic resistance genes, and the two cell populations were mixed in co-culture. The co-culture was submitted to a double-antibiotic selection. The resulting cell population was characterized for its morphology, cyclicity, and proliferative and tumorigenic capacities, in addition to transcriptomic characterization. RESULTS: We demonstrated that cell fusion could be involved in the generation of ovarian PGCCs and this process was promoted by paclitaxel and the UPR activation. Double-antibiotic treatment of PGCCs led to the selection of a pure population of cells containing both GFP- and mCherry-positive nuclei. Interestingly, after 3 weeks of selection, we observed that these cells were no longer polynucleated but displayed a single nucleus positive for both fluorescent proteins, suggesting that genetic material mixing had occurred. These cells had reinitiated their normal cell cycles, acquired an increased invasive capacity, and could form ovarian tumors in ovo. CONCLUSIONS: The UPR activation increased the in vitro formation of PGCCs by cell fusion, with the newly generated cells further acquiring new properties. The UPR modulation in ovarian cancer patients could represent an interesting therapeutic strategy to avoid the formation of PGCCs and therefore limit cancer relapse and drug resistance.

Study protocol of a phase II study to evaluate safety and efficacy of neo-adjuvant pembrolizumab and radiotherapy in localized rectal cancer.

BACKGROUND: Reshaping the tumor microenvironment by novel immunotherapies represents a key strategy to improve cancer treatment. Nevertheless, responsiveness to these treatments is often correlated with the extent of T cell infiltration at the tumor site. Remarkably, microsatellite stable rectal cancer is characterized by poor T cell infiltration and, therefore, does not respond to immune checkpoint blockade. To date, the only available curative option for these patients relies on extensive surgery. With the aim to broaden the application of promising immunotherapies, it is necessary to develop alternative approaches to promote T cell infiltration into the tumor microenvironment of these tumors. In this regard, recent evidence shows that radiotherapy has profound immunostimulatory effects, hinting at the possibility of combining it with immunotherapy. The combination of long-course chemoradiotherapy and immune checkpoint inhibition was recently shown to be safe and yielded promising results in rectal cancer, however short-course radiotherapy and immune checkpoint inhibition have never been tested in these tumors. METHODS: Our clinical trial investigates the clinical and biological impact of combining pembrolizumab with short-course radiotherapy in the neo-adjuvant treatment of localized rectal cancer. This phase II non-randomized study will recruit 25 patients who will receive short-course preoperative radiotherapy (5 Gy × 5 days) and four injections of pembrolizumab starting on the same day and on weeks 4, 7 and 10. Radical surgery will be performed three weeks after the last pembrolizumab injection. Our clinical trial includes an extensive translational research program involving the transcriptomic and proteomic analysis of tumor and blood samples throughout the course of the treatment. DISCUSSION: Our study is the first clinical trial to combine short-course radiotherapy and immune checkpoint inhibition in rectal cancer, which could potentially result in a major breakthrough in the treatment of this cancer. Additionally, the translational research program will offer insights into immunological changes within the tumor and blood and their correlation with patient outcome. Taken together, our work will help optimizing future treatment combinations and, possibly, better selecting patients. TRIAL REGISTRATION: This study was registered with www. CLINICALTRIAL: gov : NCT04109755 . Registration date: June, 2020.

Patients' experiences with cancer care in Switzerland: Results of a multicentre cross-sectional survey.

OBJECTIVES: The objectives were to describe patients' experiences of cancer care in Switzerland and explore the variation of these experiences by type of cancer. METHODS: The Swiss Cancer Patient Experiences (SCAPE) study was a cross-sectional, multicentre survey conducted in 2018. Adult patients (n = 7145) with breast, prostate, lung, colorectal, skin or haematological cancer from four large hospitals in French-speaking Switzerland were invited to complete a survey. Logistic regressions were used to assess whether experiences varied according to cancer type, adjusting for confounders. RESULTS: Of the 3121 persons who returned the survey (44% response rate), 2755 reporting an eligible cancer were included in the analyses. Participants' average score for overall care was 8.5 out of a maximum score of 10. Higher rates of positive experiences were found for nurse consultations (94%), diagnostic tests (85%) and inpatient care (82%). Lower positive responses were reported for support for people with cancer (70%), treatment decisions (66%), diagnosis (65%) and home care (55%). We observed non-systematic differences in experiences of care by cancer type. CONCLUSIONS: This large study identified that cancer patient experiences can be improved in relation to communication, information and supportive care aspects. Improvement efforts should target these areas of care to enhance responsiveness of cancer care.

ER-mitochondria contacts control surface glycan expression and sensitivity to killer lymphocytes in glioma stem-like cells.

Glioblastoma is a highly heterogeneous aggressive primary brain tumor, with the glioma stem-like cells (GSC) being more sensitive to cytotoxic lymphocyte-mediated killing than glioma differentiated cells (GDC). However, the mechanism behind this higher sensitivity is unclear. Here, we found that the mitochondrial morphology of GSCs modulates the ER-mitochondria contacts that regulate the surface expression of sialylated glycans and their recognition by cytotoxic T lymphocytes and natural killer cells. GSCs displayed diminished ER-mitochondria contacts compared to GDCs. Forced ER-mitochondria contacts in GSCs increased their cell surface expression of sialylated glycans and reduced their susceptibility to cytotoxic lymphocytes. Therefore, mitochondrial morphology and dynamism dictate the ER-mitochondria contacts in order to regulate the surface expression of certain glycans and thus play a role in GSC recognition and elimination by immune effector cells. Targeting the mitochondrial morphology, dynamism, and contacts with the ER could be an innovative strategy to deplete the cancer stem cell compartment to successfully treat glioblastoma.

Identification of Tumor Antigens Among the HLA Peptidomes of Glioblastoma Tumors and Plasma.

Glioblastoma multiforme (GBM) is the most aggressive brain tumor with poor prognosis to most patients. Immunotherapy of GBM is a potentially beneficial treatment option, whose optimal implementation may depend on familiarity with tumor specific antigens, presented as HLA peptides by the GBM cells. Further, early detection of GBM, such as by a routine blood test, may improve survival, even with the current treatment modalities. This study includes large-scale analyses of the HLA peptidome (immunopeptidome) of the plasma-soluble HLA molecules (sHLA) of 142 plasma samples, and the membranal HLA of GBM tumors of 10 of these patients' tumor samples. Tumor samples were fresh-frozen immediately after surgery and the plasma samples were collected before, and at multiple visits after surgery. In total, this HLA peptidome analysis involved 52 different HLA allotypes and resulted in the identification of more than 35,000 different HLA peptides. Strong correlations were observed in the signal intensities and in the repertoires of identified peptides between the tumors and plasma-soluble HLA peptidomes of the individual patients, whereas low correlations were observed between these HLA peptidomes and the tumors' proteomes. HLA peptides derived from Cancer/Testis Antigens (CTAs) were selected based on their presence among the HLA peptidomes of the patients and absence of expression of their source genes from any healthy and essential human tissues, except from immune-privileged sites. Additionally, peptides were selected as potential biomarkers if their levels in the plasma-sHLA peptidome were significantly reduced after the removal of tumor mass. The CTAs identified among the analyzed HLA peptidomes provide new opportunities for personalized immunotherapy and for early diagnosis of GBM.

Identification of Tumor Antigens Among the HLA Peptidomes of Glioblastoma Tumors and Plasma.

Glioblastoma multiforme (GBM) is the most aggressive brain tumor with poor prognosis to most patients. Immunotherapy of GBM is a potentially beneficial treatment option, whose optimal implementation may depend on familiarity with tumor specific antigens, presented as HLA peptides by the GBM cells. Furthermore, early detection of GBM, such as by a routine blood test, may improve survival, even with the current treatment modalities. This study includes large-scale analyses of the HLA peptidome (immunopeptidome) of the plasma-soluble HLA molecules (sHLA) of 142 plasma samples, and the membranal HLA of GBM tumors of 10 of these patients' tumor samples. Tumor samples were fresh-frozen immediately after surgery and the plasma samples were collected before, and at multiple visits after surgery. In total, this HLA peptidome analysis involved 52 different HLA allotypes and resulted in the identification of more than 35,000 different HLA peptides. Strong correlations were observed in the signal intensities and in the repertoires of identified peptides between the tumors and plasma-soluble HLA peptidomes of the individual patients, whereas low correlations were observed between these HLA peptidomes and the tumors' proteomes. HLA peptides derived from Cancer/Testis Antigens (CTAs) were selected based on their presence among the HLA peptidomes of the patients and absence of expression of their source genes from any healthy and essential human tissues, except from immune-privileged sites. Additionally, peptides were selected as potential biomarkers if their levels in the plasma-sHLA peptidome were significantly reduced after the removal of tumor mass. The CTAs identified among the analyzed HLA peptidomes provide new opportunities for personalized immunotherapy and for early diagnosis of GBM.

[Oncology: navigating the COVID-19 Pandemic and Steer the Course]. / Oncologie†: naviguer la pandémie de COVID-19 et garder le cap.

Medical oncologists are steering a difficult course during the COVID-19 pandemic between three opposing forces : revisiting optimal standards of cancer care, facing constantly evolving shortages as some resources are being redirected, and acknowledging the paradoxical need to keep patients away from the health care facility. This article compiles recommendations fr om cancer societies and expert opinions to provide guidance and practical solutions for the oncology clinic. We propose that optimal standards of care be upheld, and short-term safety concerns due to exposure to SARS-CoV-2 be weighed against a long-term compromise in cancer prognosis when deciding on adjustments in cancer care. Proper mitigation strategies in the clinic and use of less resource-heavy but equivalent treatment alternatives often allow optimal cancer care. The magnitude of benefit of cancer treatments needs to be systematically considered.

Current strategies for vaccination in glioblastoma.

PURPOSE OF REVIEW: Immunotherapy is viewed as a promising approach for glioblastoma and, in particular, therapeutic vaccines are being intensively studied. Here, we review results provided by recent clinical trials of glioblastoma vaccination and discuss the required strategies to optimize such approaches. RECENT FINDINGS: Two studies showed the feasibility of generating mutation-derived personalized vaccines in the short time frame given by the fast course of disease in glioblastoma. However, one of these demonstrated lack of mutation-derived cell surface presented MHC class I or II peptides in tumors with low mutational burden. SUMMARY: Whereas glioblastoma vaccines are well tolerated, impact on patient survival has yet to be proven. Combinations with immune checkpoint inhibitors are being tested, but strategies aiming at targeting the tumor microenvironment should be implemented as well. Finally, accurate immunomonitoring should be promoted in order to identify the best vaccine strategies, alone or in combination.

Identification of a novel population of highly cytotoxic c-Met-expressing CD8+ T lymphocytes.

CD8+ cytotoxic T lymphocytes (CTLs) are critical mediators of anti-tumor immunity, and controlling the mechanisms that govern CTL functions could be crucial for enhancing patient outcome. Previously, we reported that hepatocyte growth factor (HGF) limits effective murine CTL responses via antigen-presenting cells. Here, we show that a fraction of murine effector CTLs expresses the HGF receptor c-Met (c-Met+ CTLs). Phenotypic and functional analysis of c-Met+ CTLs reveals that they display enhanced cytolytic capacities compared to their c-Met- CTL counterparts. Furthermore, HGF directly restrains the cytolytic function of c-Met+ CTLs in cell-mediated cytotoxicity reactions in vitro and in vivo and abrogates T-cell responses against metastatic melanoma in vivo Finally, we establish in three murine tumor settings and in human melanoma tissues that c-Met+ CTLs are a naturally occurring CD8+ T-cell population. Together, our findings suggest that the HGF/c-Met pathway could be exploited to control CD8+ T-cell-mediated anti-tumor immunity.

Primordial germ cells as a potential shared cell of origin for mucinous cystic neoplasms of the pancreas and mucinous ovarian tumors.

Mucinous ovarian tumors (MOTs) morphologically and epidemiologically resemble mucinous cystic neoplasms (MCNs) of the pancreas, sharing a similar stroma and both occurring disproportionately among young females. Additionally, MOTs and MCNs share similar clinical characteristics and immunohistochemical phenotypes. Exome sequencing has revealed frequent recurrent mutations in KRAS and RNF43 in both MOTs and MCNs. The cell of origin for these tumors remains unclear, but MOTs sometimes arise in the context of mature cystic teratomas and other primordial germ cell (PGC) tumors. We undertook the present study to investigate whether non-teratoma-associated MOTs and MCNs share a common cell of origin. Comparisons of the gene expression profiles of MOTs [including both the mucinous borderline ovarian tumors (MBOTs) and invasive mucinous ovarian carcinomas (MOCs)], high-grade serous ovarian carcinomas, ovarian surface epithelium, Fallopian tube epithelium, normal pancreatic tissue, pancreatic duct adenocarcinomas, MCNs, and single-cell RNA-sequencing of PGCs revealed that both MOTs and MCNs are more closely related to PGCs than to either eutopic epithelial tumors or normal epithelia. We hypothesize that MCNs may arise from PGCs that stopped in the dorsal pancreas during their descent to the gonads during early human embryogenesis, while MOTs arise from PGCs in the ovary. Together, these data suggest a common pathway for the development of MCNs and MOTs, and suggest that these tumors may be more properly classified as germ cell tumor variants. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Prolonged Versus Short-Duration Use of Nasogastric Tubes in Patients with Head and Neck Cancer During Radiotherapy Alone or Combined Chemoradiotherapy.

To compare safety and effectiveness of prolonged (>28 days) versus short duration (≤28 days) use of nasogastric tube for enteral nutrition and weight loss prevention during curative radiotherapy with or without concurrent chemotherapy or cetuximab for head and neck cancer patients. We performed a retrospective study and database review of all patients at our center, treated with radiotherapy for head and neck cancer receiving enteral nutrition by nasogastric tube. Type of treatment, weight and body mass index changes, and related complications (gastroesophageal reflux, pneumonia, ulcer, feeding tube obstruction, or dislocation) were documented. Comparison between patients with prolonged (>28 days, group A) and short duration (≤28 days, group B) of EN through nasogastric tube was performed. Data expressed as mean ± SD or median (min; max) values as appropriate, and analyzed by ANOVA repeated measures and Kaplan-Meier estimates. We identified 114 patients who fulfilled the inclusion criteria. Among them, 10% were treated with radiotherapy alone, while 90% received concurrent chemotherapy or cetuximab. Ninety-four patients (82%, group A) had a nasogastric tube in place for a period >28 days and 20 (18%, group B) for ≤28 days during treatment. Patients were mainly men (86 patients, 75%), with a median age of 61 years (range 49-73) and advanced stage IV disease in most cases (87 patients, 76%) without differences between both groups (p = 0.53, 0.47, and 0.30, respectively). Treatment discontinuation did not occur within both groups. Fifty-six patients (49%) developed complications, without a significant difference between both groups (P = 0.23). Body weight and BMI changes did not differ during EN (-0.8 ± 4.5 and -0.3 ± 1.6), the oncological treatment (-5.3 ± 4.0 and -1.8 ± 1.4), or 6 months after the end of treatment (-0.6 ± 4.4 and -0.2 ± 1.5). Our findings suggest that prolonged enteral nutrition by nasogastric tube is safe and effective in preventing weight loss during curative radiotherapy or radio-chemotherapy for head and neck cancer.

[« Doctor, I am fasting during chemotherapy ¼ : illusion or new clinical facts ?] / « Docteur, je jeûnelors de la chimiothérapie ¼ : illusion ou nouvelle réalité clinique ?

Fasting concomitantly with oncology treatments (chemotherapy mainly) induces a growing interest among patients following overmediatisation of recent discoveries. The goal of this article is to provide updated information about this approach. According to preclinical studies, fasting may be a way to increase the therapeutic index of major oncology treatments. However, clinical data is based on small exploratory studies only and the results of larger scale studies are not yet available. The approach of fasting during chemotherapy can and should neither be recommended nor implemented in standard care. However, further scientific and clinical investigation may contribute to a better understanding of the metabolic aspects of cancer.

Jeûner au cours des traitements oncologiques (de la chimiothérapie en particulier) génère un intérêt croissant auprès des patients suite à la surmédiatisation de récentes découvertes. Le but de cet article est de faire un état des lieux sur les bases scientifiques de cette approche. Selon des études précliniques, le jeûne pourrait augmenter l'index thérapeutique des principaux traitements oncologiques. Mais les données cliniques manquent cruellement, malgré quelques données émanant de petites études exploratoires. Les études à plus grande échelle sont en cours. Aujourd'hui, l'approche du jeûne pendant la chimiothérapie ne peut et ne doit être ni recommandée ni introduite dans des protocoles de soins. Son étude peut par contre améliorer la compréhension des aspects métaboliques du cancer.

[The care of the sexuality among oncologic patients - A research project within the University hospitals of Geneva]. / Prise en charge de la sexualité chez les patients oncologiques: Un projet de recherche au sein des Hôpitaux universitaires de Genève.

Cancer affects sexual function, sexual health and relationships as well as mental health and quality of life. Due to the importance of sexual health for each individual and in order to ensure optimal and quality care, we have assessed the demand for sexual health and advise not only among cancer patients but also among professionals in this area and nursing staff. The results showed that sexuality was important both for patients and caregivers. Also we did find out that carers want to create a better professional network with the aim of redirecting patients to specialists according to their specific needs. Carers have also expressed a desire to have relevant training in sexology and sexual health to achieve this.

Les cancers affectent la fonction sexuelle, la santé sexuelle, la relation de couple, ainsi que la santé mentale et la qualité de vie. Pour assurer une prise en charge et une qualité de soins optimales, en raison de l'importance de la santé sexuelle pour chaque individu, nous avons évalué la demande concernant la sexualité et la santé sexuelle chez les patients atteints d'un cancer et les besoins de connaissances professionnelles dans ce domaine du personnel soignant. Les résultats ont montré que la sexualité avait une place importante chez les patients et les soignants. De plus, les soignants souhaitent créer un meilleur réseau professionnel, dans le but de rediriger les patients vers des spécialistes, en fonction de leurs besoins spécifiques. Enfin les soignants ont exprimé une volonté de mieux se former en sexologie et en santé sexuelle.

[Cancer and sexual medicine: a hand to the human]. / Médecine sexuelle et cancers : une main tendue vers l'humain.

Cancer itself and its treatments can result in physical and/or psychological adverse events which include disorders of sexual health. Both clinicians and patients are reluctant to talk about sexual issues, body image or intimacy between partners. Barriers from the clinician's perspective include feeling inadequately trained, lack of time or privacy. On the patients' side, embarrassment or belief that their sexual health are an untreatable complication of their disease and its treatment are the main barriers. Concerned by this problematic, a multidisciplinary group was created to address the question of sexual health in cancer patients and to set up a specialized training for the clinicians, with the overall goal to integrate this field in the standard management of cancer patients.

Le cancer et ses traitements peuvent induire de nombreux effets secondaires. Parmi eux, les problèmes liés à la santé sexuelle sont souvent laissés de côté, tant par les cliniciens que par les patients eux-mêmes. Du côté des cliniciens, les principaux obstacles évoqués sont le sentiment d'une formation inadéquate, ou le manque de temps ou d'intimité. Du côté des patients, la gêne ou l'impression que les problèmes sexuels liés au cancer sont incurables, sont souvent retrouvées. Devant cette problématique, un groupe multidisciplinaire a été créé pour traiter de la santé sexuelle des patients oncologiques et mettre en place une formation spécialisée pour les cliniciens, avec comme objectif global d'intégrer la problématique sexuelle dans la prise en charge globale de ces patients.

Management of central nervous system involvement in chronic lymphocytic leukaemia: a retrospective cohort of 30 patients.

Central nervous system involvement (CNSi) is a rare and poorly reported complication of chronic lymphocytic leukaemia (CLL). Establishing cause and effect between the CLL and the neurological symptoms remains challenging. We have analysed a retrospective cohort of 30 CLL patients with CNSi, documented by lymphocytic infiltration either by flow cytometry of the cerebrospinal fluid (CSF; n = 29) or CNS biopsy (n = 1). Neurological symptoms were heterogeneous. At the time of CNSi, less than half of the patients had a progressive CLL and 20 had never been treated for CLL. Initial treatment with fludarabine-based immuno-chemotherapy, with or without intra-CSF therapy, led to durable response in eight out of nine untreated patients. In contrast, 50% patients receiving various prior treatments needed additional therapy within a median of 4 months (1-16). Ibrutinib led to complete response in 4/4 heavily pre-treated patients. From CNSi, 5-year overall survival was 72% and 48% for treatment-naïve and previously treated patients respectively (P = 0·06); 5-year progression-free survival (PFS) was 43% and 0% (P = 0·125). 17p deletion was significantly associated with poor PFS (P = 0·006). CNSi may be the only sign of progression of CLL and should be considered an initiation criterion of systemic treatment. Prognosis seemed to be related to CLL characteristics rather than to CNSi itself.

Enhancing Antitumor Immune Responses by Optimized Combinations of Cell-penetrating Peptide-based Vaccines and Adjuvants.

Cell penetrating peptides (CPPs) from the protein ZEBRA are promising candidates to exploit in therapeutic cancer vaccines, since they can transport antigenic cargos into dendritic cells and induce tumor-specific T cells. Employing CPPs for a given cancer indication will require engineering to include relevant tumor-associated epitopes, administration with an appropriate adjuvant, and testing for antitumor immunity. We assessed the importance of structural characteristics, efficiency of in vitro transduction of target cells, and choice of adjuvant in inducing the two key elements in antitumor immunity, CD4 and CD8 T cells, as well as control of tumor growth in vivo. Structural characteristics associated with CPP function varied according to CPP truncations and cargo epitope composition, and correlated with in vitro transduction efficiency. However, subsequent in vivo capacity to induce CD4 and CD8 T cells was not always predicted by in vitro results. We determined that the critical parameter for in vivo efficacy using aggressive mouse tumor models was the choice of adjuvant. Optimal pairing of a particular ZEBRA-CPP sequence and antigenic cargo together with adjuvant induced potent antitumor immunity. Our results highlight the irreplaceable role of in vivo testing of novel vaccine constructs together with adjuvants to select combinations for further development.

Phenotypic switch of CD8(+) T cells reactivated under hypoxia toward IL-10 secreting, poorly proliferative effector cells.

CD8(+) T cells controlling pathogens or tumors must function at sites where oxygen tension is frequently low, and never as high as under atmospheric culture conditions. However, T-cell function in vivo is generally analyzed indirectly, or is extrapolated from in vitro studies under nonphysiologic oxygen tensions. In this study, we delineate the role of physiologic and pathologic oxygen tension in vitro during reactivation and differentiation of tumor-specific CD8(+) T cells. Using CD8(+) T cells from pmel-1 mice, we observed that the generation of CTLs under 5% O2, which corresponds to physioxia in lymph nodes, gave rise to a higher effector signature than those generated under atmospheric oxygen fractions (21% O2). Hypoxia (1% O2) did not modify cytotoxicity, but decreasing O2 tensions during CTL and CD8(+) tumor-infiltrating lymphocyte reactivation dose-dependently decreased proliferation, induced secretion of the immunosuppressive cytokine IL-10, and upregulated the expression of CD137 (4-1BB) and CD25. Overall, our data indicate that oxygen tension is a key regulator of CD8(+) T-cell function and fate and suggest that IL-10 release may be an unanticipated component of CD8(+) T cell-mediated immune responses in most in vivo microenvironments.