Preclinical Development of CAR T Cells with Antigen-Inducible IL18 Enforcement to Treat GD2-Positive Solid Cancers.

PURPOSE: Cytokine-engineering of chimeric antigen receptor-redirected T cells (CAR T cells) is a promising principle to overcome the limited activity of canonical CAR T cells against solid cancers. EXPERIMENTAL DESIGN: We developed an investigational medicinal product, GD2IL18CART, consisting of CAR T cells directed against ganglioside GD2 with CAR-inducible IL18 to enhance their activation response and cytolytic effector functions in the tumor microenvironment. To allow stratification of patients according to tumor GD2 expression, we established and validated immunofluorescence detection of GD2 on paraffin-embedded tumor tissues. RESULTS: Lentiviral all-in-one vector engineering of human T cells with the GD2-specific CAR with and without inducible IL18 resulted in cell products with comparable proportions of CAR-expressing central memory T cells. Production of IL18 strictly depends on GD2 antigen engagement. GD2IL18CART respond to interaction with GD2-positive tumor cells with higher IFNγ and TNFα cytokine release and more effective target cytolysis compared with CAR T cells without inducible IL18. GD2IL18CART further have superior in vivo antitumor activity, with eradication of GD2-positive tumor xenografts. Finally, we established GMP-compliant manufacturing of GD2IL18CART and found it to be feasible and efficient at clinical scale. CONCLUSIONS: These results pave the way for clinical investigation of GD2IL18CART in pediatric and adult patients with neuroblastoma and other GD2-positive cancers (EU CT 2022-501725-21-00).

IRF4 downregulation improves sensitivity and endurance of CAR T cell functional capacities.

Chimeric antigen receptor (CAR) modified T cells can induce complete remissions in patients with advanced hematological malignancies. Nevertheless, the efficacy is mostly transient and remains so far poor in the treatment of solid tumors. Crucial barriers to long-term CAR T cell success encompass loss of functional capacities known as "exhaustion", among others. To extend CAR T cell functionality, we reduced interferon regulatory factor 4 (IRF4) levels in CAR T cells using a one-vector system encoding a specific short-hairpin (sh) RNA along with constitutive CAR expression. At baseline, CAR T cells with downregulated IRF4 showed equal cytotoxicity and cytokine release compared to conventional CAR T cells. However, under conditions of repetitive antigen encounter, IRF4low CAR T cells displayed enhanced functionality with superior cancer cell control in the long-term compared with conventional CAR T cells. Mechanistically, the downregulation of IRF4 in CAR T cells resulted in prolonged functional capacities and upregulation of CD27. Moreover, IRF4low CAR T cells were more sensitive to cancer cells with low levels of target antigen. Overall, IRF4 downregulation capacitates CAR T cells to recognize and respond to target cells with improved sensitivity and endurance.

CAR Triggered Release of Type-1 Interferon Limits CAR T-Cell Activities by an Artificial Negative Autocrine Loop.

The advent of chimeric antigen receptor (CAR) T cells expedited the field of cancer immunotherapy enabling durable remissions in patients with refractory hematological malignancies. T cells redirected for universal cytokine-mediated killing (TRUCKs), commonly referred to as "fourth generation" CAR T-cells, are designed to release engineered payloads upon CAR-induced T-cell activation. Building on the TRUCK technology, we aimed to generate CAR T-cells with a CAR-inducible artificial, self-limiting autocrine loop. To this end, we engineered CAR T-cells with CAR triggered secretion of type-1 interferons (IFNs). At baseline, IFNα and IFNß CAR T-cells showed similar capacities in cytotoxicity and cytokine secretion compared to conventional CAR T-cells. However, under "stress" conditions of repetitive rounds of antigen stimulation using BxPC-3 pancreas carcinoma cells as targets, anti-tumor activity faded in later rounds while being fully active in destructing carcinoma cells during first rounds of stimulation. Mechanistically, the decline in activity was primarily based on type-1 IFN augmented CAR T-cell apoptosis, which was far less the case for CAR T-cells without IFN release. Such autocrine self-limiting loops can be used for applications where transient CAR T-cell activity and persistence upon target recognition is desired to avoid lasting toxicities.

Modification of genital kidney nephrons for sperm transport in a plethodontid salamander, Eurycea longicauda.

Salamanders possess kidneys with two distinct regions: a caudal pelvic portion and cranial genital portion. Nephrons of the pelvic region are responsible for urine formation and transport. Nephrons of the genital region transport sperm from testes to Wolffian ducts; however, nephrons of the genital region possess all the same functional regions found in pelvic kidney nephrons that are involved with urine formation and transport (renal corpuscles, proximal tubules, distal tubules, and collecting ducts). Morphological similarities between pelvic and genital regions stimulated past researchers to hypothesize that nephrons of genital kidneys possess dual function; that is, sperm transport and urine formation/transport. Considering size of glomeruli is directly related to the total amount of blood plasma filtered into the Bowman's space, we tested the hypothesis that nephrons of genital kidneys have reduced urine formation function by comparing glomerular size between nephrons of pelvic and genital kidney regions in Eurycea longicauda with general histological techniques. Light microscopy analysis revealed that glomeruli of pelvic kidneys were significantly larger than those measured from genital kidneys. Transmission electron microscopy analysis also revealed modifications in genital kidney nephrons when compared to pelvic kidney nephrons that suggested a decrease in urine formation function in genital kidneys. Such modifications included a decrease in basal and lateral plasma membrane folding in genital kidney proximal and distal tubules compared to that of pelvic kidney proximal and distal tubules. Genital kidney proximal tubules were also ciliated, which was not observed in pelvic kidney proximal tubules. In conclusion, although structurally similar at the histological level, it appears that nephrons of genital kidneys have decreased urine formation function based on glomerular size comparison and nephron ultrastructure.

Chimeric antigen receptors designed to overcome transforming growth factor-ß-mediated repression in the adoptive T-cell therapy of solid tumors.

Adoptive cell therapy with chimeric antigen receptor (CAR)-engineered T cells produced lasting remissions in the treatment of advanced, so far refractory B-cell malignancies; however, the elimination of solid tumors remains so far elusive. The low efficacy of CAR T cells is thought to be due to the immune-repressive milieu within the tumor lesion, predominantly mediated by transforming growth factor-ß (TGF-ß) that represses effector T-cell activities and drives differentiation towards regulatory T cells (Tregs). Seeking to boost antitumor immunity, TGF-ß is currently targeted by different means in pre-clinical studies. While a recent clinical trial showed the utility of shielding CAR T cells from TGF-ß repression, further strategies in counteracting TGF-ß in the adoptive cell therapy warrant exploration. We here discuss the most recent advances in the field and draw future developments to make CAR T-cell therapy more potent in the treatment of solid cancer.

Acceptance of televideo technology by adults aging with a mobility impairment for health and wellness interventions.

Televideo technology (e.g., Skype) has potential to support adults as they age by facilitating their interactions with people remotely; this potential may be even greater for adults aging with pre-existing mobility impairments, who face challenges interacting with others in person due to lack of transportation or accessibility. Our research employed questionnaire and interview methodologies to investigate this understudied population about their attitudes toward televideo technology for supporting social engagement, healthcare provider access, and physical activity. Participants were 14 adults aging with self-reported mobility impairments (50-70 years of age). Overall, participants were open to accepting televideo technology for social engagement, healthcare provider access, and physical activity. Participants perceived these technologies to be useful and perceived additional benefits, including the feeling of "being there" by enabling the viewing of facial expressions and the environment of the other person. Concerns included perceptions that televideo technologies lack security and privacy. Participants also expressed concerns that these technologies are difficult to use and are difficult to learn to use. The findings have implications for education, training, and deployment of televideo technology for home-based interventions for adults aging with pre-existing mobility impairments.

Understanding older adults' perceptions of and attitudes towards exergames.

PURPOSE: Maintaining physical activity is a key component of successful aging and has benefits for both physical and cognitive functioning in the older adult population. One promising method for engaging in physical activity is through exergames, which are video games designed to promote exercise. Exergames have the potential to be used by a wide range of people, including older adults, in a variety of settings, such as at home, in community living environments, or senior centers. However, exergames have not been designed for older adults (e.g., with respect to their attitudes, needs). Thus, older adults may not adopt these systems if they perceive them as not useful or relevant to them. METHOD: Twenty older adults (aged 60-79) interacted with two exergames, and were then interviewed about their perceptions of the system's ease of use and usefulness, as well as their general attitudes towards the system. RESULTS: Participants identified the potential for exergames' usefulness for various goals, such as to increase their physical activity. However, they also reported negative attitudes concerning the system, including perceiving barriers to system use. Overall, participants said they would use the system in the future and recommend it to other people at their age for improving health, despite these use challenges. CONCLUSION: The older adults were open to adopting exergames, which could provide opportunities to increase physical activity. Given the participants' overall positive perceptions of the usefulness of exergames, designers must address the perceived challenges of using these systems. Understanding barriers and facilitators for older adults' use of exergames can guide design, training, and adoption of these systems.

Assessing Older Adults' Usability Challenges Using Kinect-Based Exergames.

Exergames have been growing in popularity as a means to get physical exercise. Although these systems have many potential benefits both physically and cognitively, there may be barriers to their use by older adults due to a lack of design consideration for age-related changes in motor and perceptual capabilities. In this paper we evaluate the usability challenges of Kinect-based exergames for older adults. Older adults rated their interaction with the exergames system based on their perceived usefulness and ease-of-use of these systems. Although many of the participants felt that these systems could be potentially beneficial, particularly for exercise, there were several challenges experienced. We discuss the implications for design guidelines based on the usability challenges assessed.