Tumor-infiltrating lymphocytes and the PD-L1/PD-1 signaling axis in breast cancer: biology and clinical implications

Cancer immunotherapy and especially immune checkpoint blockade therapy has revolutionized cancer treatment in many tumor types including breast cancer (BC). The expression and specific role of Programmed Death-1 (PD-1)/Programmed Death-Ligand 1 (PD-L1) signaling axis, immune-related gene signatures and immune cell subpopulations needs to be fully elucidated while the identification of prognostic and predictive biomarkers in BC is of utmost importance. The overall aim of the thesis was to investigate the expression, prognostic/predictive implications and regulatory mechanisms of the PD-1/PD-L1 checkpoints and immune infiltrate in early and advanced BC. In paper I, a comprehensive analysis of PD-L1 expression patterns and its prognostic implications was performed in early BC. PD-L1 protein expression was mostly expressed in immune cells and was most abundant in the TNBC subtype. PD-L1 expression in tumor cells was a poor prognostic factor whereas PD-L1 expression in immune cells was correlated with improved survival outcomes in TNBC, as revealed in a trial-level meta-analysis including 38 studies. PD-L1 gene expression was associated with improved prognosis in the entire population as revealed in a pooled transcriptomic analysis of 39 publicly available datasets. PD-L1 expression can therefore represent a promising clinically relevant biomarker of good prognosis which can also select appropriate candidates for immunotherapy. In paper II, PD-L1 expression was evaluated both at the protein and mRNA level in the same retrospective early BC patient cohort. Both protein (stained with the antibody clone SP263) and gene expression predicted improved outcome in early BC and also correlated with enhanced T-cell infiltration (CD3+ IHC and in silico estimation of CD8+/CD4+ T-cells). Of note, PD-L1 mRNA expression can add significant prognostic value to the prospectively validated 21- and 70-gene gene signatures in ER+/HER2- disease. Upon validation, this finding might improve prognostication capacity of the current gene signatures. In paper III, we investigated the role of STAT3 as a regulator of PD-L1 expression and immune response in BC in vitro, in vivo and in BC patient samples. A positive correlation between STAT3 and PD-L1 was observed at the protein and gene expression level while a transcriptional STAT3-mediated regulation of PD-L1 was demonstrated in BC cells. Of note, STAT3 modulated antitumor immune response mainly through macrophage phenotype shift and accumulation of NK cells rather than via cytotoxic T-cell infiltration in a murine BC model. Furthermore, pro-tumoral macrophages were correlated with PD-L1 expression in BC patient tumors, thus providing insights on the tumor-immune cell interactions and potential clinical implications. In paper IV, a multi-level study of the prognostic capacity of PD-1 expression in early BC was performed including: a) a study cohort with IHC and GEP data, b) systematic review and trial- level IHC-based meta-analysis of 15 studies and c) pooled analysis of publicly available transcriptomic datasets). PD-1 protein and gene expression were correlated with improved OS in the entire population of the study cohort. In the pooled analyses PD-1 protein and gene expressions were correlated with better survival outcomes in TNBC/basal-like patients and therefore is a promising biomarker which merits further validation. In paper V the role of relevant gene signatures as predictors of response to chemotherapy was investigated in patients (n=109) with advanced BC participating in the translational sub-study of the phase III TEX trial. Fine-needle aspiration biopsies were used for gene expression profiling and for TILs enumeration. Immune-related gene signatures predicted better response to chemotherapy in ER+ and luminal BC patients which was further confirmed through an independent gene set enrichment and other in silico analyses. The lymphocytic abundance was low and predicted no effect to chemotherapy. These results may pave the way for the development of immune-based drivers of chemosensitivity in the least immunogenic luminal tumors

The interplay between eosinophils and T cells in breast cancer immunotherapy

Treatment with immune checkpoint inhibitors (ICI) has revolutionized cancer management for multiple tumor types, including breast cancer. However, not all patients respond to ICI, and unraveling the determinants and mechanisms of response still remains an unmet need. A recent study has uncovered the critical role of eosinophils in mediating immunotherapy effect in breast cancer, mainly by stimulating the activation of CD8+ T‐cells. Furthermore, the intratumoral eosinophil recruitment was directed by CD4+ T cells and the interleukins IL‐5 and IL‐33, thus providing the rationale for targeting eosinophils to enhance ICI response

The outcome-based iCAN! / theyCAN! feedback paradigm differentiates strong and weak learning outcomes, learner diversity, and the learning outcomes of each learner: A shift to metacognitive assessment

Can learning outcomes be transformed in useful tools revealing strong and weak learning outcomes, learners, teachers; reporting student self-assessment overestimation; informing formative feedback and summative examinations

Beyond PD-1/PD-L1 Inhibition: What the Future Holds for Breast Cancer Immunotherapy

Cancer immunotherapy has altered the management of human malignancies, improving outcomes in an expanding list of diseases. Breast cancer - presumably due to its perceived low immunogenicity - is a late addition to this list. Furthermore, most of the focus has been on the triple negative subtype because of its higher tumor mutational load and lymphocyte-enriched stroma, although emerging data show promise on the other breast cancer subtypes as well. To this point the clinical use of immunotherapy is limited to the inhibition of two immune checkpoints, Programmed Cell Death Protein 1 (PD-1) and Cytotoxic T-lymphocyte-associated Protein 4 (CTLA-4). Consistent with the complexity of the regulation of the tumor − host interactions and their lack of reliance on a single regulatory pathway, combinatory approaches have shown improved efficacy albeit at the cost of increased toxicity. Beyond those two checkpoints though, a large number of co-stimulatory or co-inhibitory molecules play major roles on tumor evasion from immunosurveillance. These molecules likely represent future targets of immunotherapy provided that the promise shown in early data is translated into improved patient survival in randomized trials. The biological role, prognostic and predictive implications regarding breast cancer and early clinical efforts on exploiting these immune-related therapeutic targets are herein reviewed

Plasmablastic Lymphoma with Coexistence of Chronic Lymphocytic Leukemia in an Immunocompetent Patient: A Case Report and Mini-Review

Background. Plasmablastic lymphoma (PBL) is a rare, aggressive B-cell lymphoma with poor prognosis usually found in the oral cavity of HIV-positive patients. Chronic lymphocytic leukemia (CLL) is an indolent B-cell lymphoma with a variable clinical course. Transformation of CLL to PBL as Richter’s syndrome is rare while coexistence of CLL and PBL at diagnosis is even rarer. Case Report. We describe a case of a male immunocompetent patient with an ileum-cecum valve mass and a soft tissue mass at the left humerus with histologic evidence of PBL with coexistence of CLL in the bone marrow and peripheral blood. Amputation of the patient’s left arm was inevitable, and the patient was started on bortezomib and dexamethasone. However, prolonged hospitalization was complicated by aspiration pneumonia, and the patient passed away. Conclusions. No standard of care exists for patients with PBL, and prognosis remains dismal. Concomitant presentation of hematological malignancies becomes increasingly recognized, and further insight is needed in order to delineate whether they originate from the same clone or from different ones

Sense of Coherence and Defense Style Predict Sleep Difficulties in Early Non-metastatic Colorectal Cancer

BACKGROUND: Sleep disturbances are common in cancer patients, but little is known about the complex interplay between the background psychological profile, coping with health stressors capacities and psychological distress in the formation of sleep difficulties in colorectal cancer. AIMS: To study the course and to identify psychological predictors of sleep difficulties in early non-metastatic colorectal cancer patients over a one-year period. METHODS: In this 1-year prospective study, we assessed in 84 early non-metastatic colorectal cancer patients the association of psychological distress (SCL-90-R), sense of coherence (SOC-29), and defense styles (Defense Style Questionnaire) with sleep difficulties (SCL-90-R) in multiple regression models. Eighty-two patients with breast cancer and 50 patients with cancer of unknown primary site served as disease controls, and 84 matched for age and sex alleged healthy individuals served as healthy controls. RESULTS: Colorectal cancer patients presented more sleep difficulties compared to healthy participants but fewer than patients with breast cancer and cancer of unknown primary site. Colorectal cancer patients' trouble falling asleep (p = 0.033) and wakening up early in the morning (p < 0.001) deteriorated over time. Sleep that was restless or disturbed was independently associated with low SOC (p = 0.046) and maladaptive defenses (p = 0.008). Anxiety symptoms (p < 0.001) predicted deterioration in trouble falling asleep, while depressive symptoms (p = 0.022) and self-sacrificing defense style (p = 0.049) predicted deterioration in wakening up early in the morning. CONCLUSIONS: Psychological parameters and coping with health stressors capacities are independently associated with sleep difficulties in colorectal cancer patients, indicating the need for psychological interventions aiming at improving adjustment to the disease

SETD2 mutation in renal clear cell carcinoma suppress autophagy via regulation of ATG12

Inactivating mutations in the SETD2 gene, encoding for a nonredundant histone H3 methyltransferase and regulator of transcription, is a frequent molecular feature in clear cell renal cell carcinomas (ccRCC). SETD2 deficiency is associated with recurrence of ccRCC and bears low prognostic values. Targeting autophagy, a conserved catabolic process with critical functions in maintenance of cellular homeostasis and cell conservation under stress condition, is emerging as a potential therapeutic strategy to combat ccRCC. Epigenetics-based pathways are now appreciated as key components in the regulation of autophagy. However, whether loss of function in the SETD2 histone modifying enzyme occurring in ccRCC cells may impact on their ability to undergo autophagy remained to be explored. Here, we report that SETD2 deficiency in RCC cells is associated with the aberrant accumulation of both free ATG12 and of an additional ATG12-containing complex, distinct from the ATG5-ATG12 complex. Rescue of SETD2 functions in the SETD2 deficiency in RCC cells, or reduction of SETD2 expression level in RCC cells wild type for this enzyme, demonstrates that SETD2 deficiency in RCC is directly involved in the acquisition of these alterations in the autophagic process. Furthermore, we revealed that deficiency in SETD2, known regulator of alternative splicing, is associated with increased expression of a short ATG12 spliced isoform at the depend of the canonical long ATG12 isoform in RCC cells. The defect in the ATG12-dependent conjugation system was found to be associated with a decrease autophagic flux, in accord with the role for this ubiquitin-like protein conjugation system in autophagosome formation and expansion. Finally, we report that SETD2 and ATG12 gene expression levels are associated with favorable respective unfavorable prognosis in ccRCC patients. Collectively, our findings bring further argument for considering the SETD2 gene status of ccRCC tumors, when therapeutic interventions, such as targeting the autophagic process, are considered to combat these kidney cancers

Prognostic implications of PD-L1 expression in breast cancer: Systematic review and meta-analysis of immunohistochemistry and pooled analysis of transcriptomic data

Purpose: Conflicting data have been reported on the prognostic value of PD-L1 protein and gene expression in breast cancer. Experimental Design: Medline, Embase, Cochrane Library, and Web of Science Core Collection were searched, and data were extracted independently by two researchers. Outcomes included pooled PD-L1 protein positivity in tumor cells, immune cells, or both, per subtype and per antibody used, and its prognostic value for disease-free and overall survival. A pooled gene expression analysis of 39 publicly available transcriptomic datasets was also performed. Results: Of the initial 4,184 entries, 38 retrospective studies fulfilled the predefined inclusion criteria. The overall pooled PD-L1 protein positivity rate was 24% (95% CI, 15%-33%) in tumor cells and 33% (95% CI, 14%- 56%) in immune cells. PD-L1 protein expression in tumor cells was prognostic for shorter overall survival (HR, 1.63; 95% CI, 1.07-2.46; P = 0.02); there was significant heterogeneity (I2 = 80%, Pheterogeneity < 0.001). In addition, higher PD-L1 gene expression predicted better survival in multivariate analysis in the entire population (HR, 0.82; 95% CI, 0.74- 0.90; P < 0.001 for OS) and in basal-like tumors (HR, 0.64; 95% CI, 0.52-0.80; P < 0.001 for OS; Pinteraction 0.005). Conclusions: The largest to our knowledge meta-analysis on the subject informs on PD-L1 protein positivity rates and its prognostic value in breast cancer. Standardization is needed prior to routine implementation. PD-L1 gene expression is a promising prognostic factor, especially in basal-like breast cancer. Discrepant prognostic information might be related to PD-L1 gene expression in the stroma.SCOPUS: ar.jinfo:eu-repo/semantics/publishe