Prostate cancer (PCa) has become the most common cancer among males in Europe and the USA. Adoptive immunotherapy appears a promising strategy to control the advanced stages of the disease by specifically targeting the tumor, in particular through chimeric antigen receptor T (CAR-T) cell therapy. Despite the advancements of CAR-T technology in the treatment of hematological malignancies, solid tumors still represent a challenge. To overcome current limits, other cellular effectors than T lymphocytes are under study as possible candidates for CAR-engineered cancer immunotherapy. A novel approach involves the NK-92 cell line, which mediates strong cytotoxic responses against a variety of tumor cells but has no effect on non-malignant healthy counterparts. Here, we report a novel therapeutic approach against PCa based on engineering of NK-92 cells with a CAR recognizing the human prostate-specific membrane antigen (PSMA), which is overexpressed in prostatic neoplastic cells. More importantly, the potential utility of NK-92/CAR cells to treat PCa has not yet been explored. Upon CAR transduction, NK-92/CAR cells acquired high and specific lytic activity against PSMA-expressing prostate cancer cells in vitro, and also underwent degranulation and produced high levels of IFN-γ in response to antigen recognition. Lethal irradiation of the effectors, a safety measure requested for the clinical application of retargeted NK-92 cells, fully abrogated replication but did not impact on phenotype and short-term functionality. PSMA-specific recognition and antitumor activity were retained in vivo, as adoptive transfer of irradiated NK-92/CAR cells in prostate cancer-bearing mice restrained tumor growth and improved survival. Anti-PSMA CAR-modified NK-92 cells represent a universal, off-the-shelf, renewable, and cost-effective product endowed with relevant potentialities as a therapeutic approach for PCa immunotherapy.

Cytokine-Induced Killer (CIK) cells share several functional and phenotypical properties of both T and natural killer (NK) cells. They represent an attractive approach for cell-based immunotherapy,...

While interferon alpha (IFNα) is used in several viral and cancer contexts, its efficacy against ovarian cancer (OC) is far from being incontrovertibly demonstrated and, more importantly, is hindered by heavy systemic side effects. To overcome these issues, here we propose a strategy that allows a targeted delivery of the cytokine, by conjugating IFNα2a with an aldehyde-modified form of hyaluronic acid (HA). The resulting HA-IFNα2a bioconjugate was biochemically and biologically characterized. The conjugation with HA did not substantially modified both the antiviral function and the anti-proliferative activity of the cytokine. Moreover, the induction of STAT1 phosphorylation and of a specific gene expression signature in different targets was retained. In vivo optical imaging biodistribution showed that the i.p.-injected HA-IFNα2a persisted into the peritoneal cavity longer than IFNα2a without being toxic for intraperitoneal organs, thus potentially enhancing the loco-regional therapeutic effect. Indeed, in OC xenograft mouse models bioconjugate significantly improved survival as compared to the free cytokine. Overall, HA-IFNα2a bioconjugate disclosed an improved anticancer efficacy, and can be envisaged as a promising loco-regional treatment for OC.

Background: Patients affected by aggressive B-cell malignancies who are resistant to primary or salvage chemoimmunotherapy have an extremely poor prognosis and limited therapeutic options. Promising therapeutic success has been achieved with the infusion of CD19 chimeric antigen receptor-T cells, but several limits still restrain the administration to a limited proportion of patients. This unmet clinical need might be fulfilled by an adoptive immunotherapy approach that combines cytokine-induced killer (CIK) cells and monoclonal antibodies (mAb) to the CD20 antigen. Indeed, CIK cells are an effector population endowed with antitumor activity, which can be further improved and antigen-specifically redirected by clinical-grade mAb triggering antibody-dependent cell-mediated cytotoxicity. Methods: CIK cells were generated from peripheral blood of patients affected by different B-cell malignancies using a blinatumomab-based cell culture protocol. Effector cells were combined with the anti-CD20 mAb obinutuzumab and their therapeutic activity was assessed both in vitro and in vivo. Results: CIK cells were successfully expanded in clinically relevant numbers, starting from small volumes of peripheral blood with extremely low CD3⁺ counts and high tumor burden. This relied on the addition of blinatumumab in culture, which leads to the simultaneous expansion of effector cells and the complete elimination of the neoplastic component. Moreover, CIK cells were highly cytotoxic in vitro against both B-cell tumor cell lines and autologous neoplastic targets, and had a significant therapeutic efficacy against a B-cell malignancy patient-derived xenograft on in vivo transfer. Conclusions: The combination of an easily expandable CIK cell effector population with a mAb already in clinical use establishes a tumor antigen-specific redirection strategy that can be rapidly translated into clinical practice, providing an effective therapeutic alternative for B-cell malignancies without any need for genetic modifications. Additionally, the approach can be potentially applied to an extremely vast array of different tumors by simply substituting the targeting mAb.

Cytokine-Induced (CIK) cells represent an attractive approach for cell-based immunotherapy, as they show several advantages compared with other strategies. Here we describe an original serum-free protocol for CIK cell expansion that employs G-Rex devices and compare the resulting growth, viability, phenotypic profile and cytotoxic activity with conventional culture in tissue flasks. CIK cells were obtained from buffy coats, seeded in parallel in G-Rex and tissue flasks, and stimulated with clinical-grade IFN-γ, anti-CD3 antibody and IL-2. G-Rex led to large numbers of CIK cells, with a minimal need for technical interventions, thus reducing the time and costs of culture manipulation. CIK cells generated in G-Rex showed a less differentiated phenotype, with a significantly higher expression of naive-associated markers such as CD62L, CD45RA and CCR7, which correlates with a remarkable expansion potential in culture and could lead to longer persistence and a more sustained anti-tumor response in vivo. The described procedure can be easily translated to large-scale production under Good Manufacturing Practice. Overall, this protocol has strong advantages over existing procedures, as it allows easier, time-saving and cost-effective production of CIK effector cells, fostering their clinical application.

The high mutation rate of SARS-CoV-2 leads to the emergence of multiple variants, some of which are resistant to vaccines and drugs targeting viral elements. Targeting host dependency factors, e.g. cellular proteins required for viral replication, would help prevent the development of resistance. However, it remains unclear whether different SARS-CoV-2 variants induce conserved cellular responses and exploit the same core host factors. To this end, we compared three variants of concern and found that the host transcriptional response was conserved, differing only in kinetics and magnitude. Clustered regularly interspaced short palindromic repeats screening identified host genes required for each variant during infection. Most of the genes were shared by multiple variants. We validated our hits with small molecules and repurposed the US Food and Drug Administration-approved drugs. All the drugs were highly active against all the tested variants, including new variants that emerged during the study (Delta and Omicron). Mechanistically, we identified reactive oxygen species production as a key step in early viral replication. Antioxidants such as N-acetyl cysteine (NAC) were effective against all the variants in both human lung cells and a humanized mouse model. Our study supports the use of available antioxidant drugs, such as NAC, as a general and effective anti-COVID-19 approach.