Immune profile of patients with peritoneal carcinomatosis selected for CRS-HIPEC therapy.

Cytoreductive surgery (CRS) combined with hyperthermic intraperitoneal chemotherapy (HIPEC) is a treatment option for peritoneal carcinomatosis (PC) from colorectal cancer (CRC), which is otherwise a terminal stage of disease. Nevertheless, survival outcomes are only marginally superior to other treatments. This fact highlights the need for better strategies to control intra-abdominal disease recurrence after CRS-HIPEC, including the complementary use of immunotherapies. The aim of this study was therefore to investigate the immune phenotype of T cells in patients with PC. Fifty three patients with CRC (34 patients with PC and 19 patients without PC) were enrolled in a prospective study (clinicaltrials.gov: NCT04108936). Peripheral blood and omental fat were collected to isolate peripheral blood mononuclear cells (PBMCs) and adipose tissue mononuclear cells (ATMCs). These cells were analysed by flow cytometry using a panel focused upon T cell memory differentiation and exhaustion markers. We found a more naïve profile for CD8+ T cells in peripheral blood and intra-abdominal fat of PC patients compared to comparator group (CG) patients. Furthermore, there was an over-representation of CD4+ T cells expressing inhibitory receptors in adipose tissue of PC patients, but not in blood. Our description of intraperitoneal T cell subsets gives us a better understanding of how peritoneal carcinomatosis shapes local immune responses.

External validation of biomarkers for immune-related adverse events after immune checkpoint inhibition.

Immune checkpoint inhibitors have revolutionized treatment of advanced melanoma, but commonly cause serious immune-mediated complications. The clinical ambition of reserving more aggressive therapies for patients least likely to experience immune-related adverse events (irAE) has driven an extensive search for predictive biomarkers. Here, we externally validate the performance of 59 previously reported markers of irAE risk in a new cohort of 110 patients receiving Nivolumab (anti-PD1) and Ipilimumab (anti-CTLA-4) therapy. Alone or combined, the discriminatory value of these routine clinical parameters and flow cytometry biomarkers was poor. Unsupervised clustering of flow cytometry data returned four T cell subsets with higher discriminatory capacity for colitis than previously reported populations, but they cannot be considered as reliable classifiers. Although mechanisms predisposing some patients to particular irAEs have been described, we are presently unable to capture adequate information from pre-therapy flow cytometry and clinical data to reliably predict risk of irAE in most cases.

Initial Steps for the Development of a Phage-Mediated Gene Replacement Therapy Using CRISPR-Cas9 Technology.

p53 gene (TP53) replacement therapy has shown promising results in cancer gene therapy. However, it has been hampered, mostly because of the gene delivery vector of choice. CRISPR-Cas9 technology (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) can knock out the mutated TP53 (mutTP53), but due to its large size, many viral vectors are not suitable or require implemented strategies that lower the therapeutic efficiency. Here, we introduced a bacteriophage or phage-based vector with the ability to target cancer cells and aimed to investigate the feasibility of using this vector to deliver CRISPR-Cas9 transgene in human lung adenocarcinoma cells. First, we produced a tumour-targeted bacteriophage carrying a CRISPR-Cas9 transgene cassette. Next, we investigated any negative impact on vector titers via quantitative polymerase chain reaction (qPCR) and colony-forming agar plate. Last, we combined Western blot analysis and immunofluorescence staining to prove cell transduction in vitro. We showed that the tumour-targeted bacteriophage can package a large-size vector genome, ~10 kb, containing the CRISPR-Cas9 sequence without any negative impact on the active or total number of bacteriophage particles. Then, we detected expression of the Cas9 in human lung adenocarcinoma cells in a targeted and efficient manner. Finally, we proved loss of p53 protein expression when a p53 gRNA was incorporated into the CRISPR-Cas9 phage DNA construct. These proof-of-concept findings support the use of engineered bacteriophage for TP53 replacement therapy in lung cancer.