Current cancer immunotherapy predominately focuses on eliciting type 1 immune responses fighting cancer; however, long-term complete remission remains uncommon1,2. A pivotal question arises as to whether type 2 immunity can be orchestrated alongside type 1-centric immunotherapy to achieve enduring response against cancer3,4. Here we show that an interleukin-4 fusion protein (Fc-IL-4), a typical type 2 cytokine, directly acts on CD8+ T cells and enriches functional terminally exhausted CD8+ T (CD8+ TTE) cells in the tumour. Consequently, Fc-IL-4 enhances antitumour efficacy of type 1 immunity-centric adoptive T cell transfer or immune checkpoint blockade therapies and induces durable remission across several syngeneic and xenograft tumour models. Mechanistically, we discovered that Fc-IL-4 signals through both signal transducer and activator of transcription 6 (STAT6) and mammalian target of rapamycin (mTOR) pathways, augmenting the glycolytic metabolism and the nicotinamide adenine dinucleotide (NAD) concentration of CD8+ TTE cells in a lactate dehydrogenase A-dependent manner. The metabolic modulation mediated by Fc-IL-4 is indispensable for reinvigorating intratumoural CD8+ TTE cells. These findings underscore Fc-IL-4 as a potent type 2 cytokine-based immunotherapy that synergizes effectively with type 1 immunity to elicit long-lasting responses against cancer. Our study not only sheds light on the synergy between these two types of immune responses, but also unveils an innovative strategy for advancing next-generation cancer immunotherapy by integrating type 2 immune factors.
© 2024. The Author(s).

To determine the role of the AKT pathway in the regulating of natural Killer-induced apoptosis of acute myeloid leukemia cells and to characterize the associated molecular mechanisms.
BALB/c nude mice were injected with HL60 cells to induce a xenogenic model of subcutaneous leukemic tumors. Mice were treated with perifosine, and their spleens were analyzed using biometry, histopathology, and immunohistochemistry. Gene expression analysis in leukemia cells was performed by real-time PCR. Protein analysis of leukemia and natural Killer cells was performed by flow cytometry. AKT inhibition in HL60 cells, followed by co-culture with natural Killer cells was performed to assess cytotoxicity. Apoptosis rate was quantified using flow cytometry.
Perifosine treatment caused a reduction in leukemic infiltration in the spleens of BALB/c nude mice. In vitro , AKT inhibition reduced HL60 resistance to natural Killer-induced apoptosis. AKT inhibition suppressed the immune checkpoint proteins PD-L1, galectin-9, and CD122 in HL60 cells, but did not change the expression of their co-receptors PD1, Tim3, and CD96 on the natural Killer cell surface. In addition, the death receptors DR4, TNFR1, and FAS were overexpressed by AKT inhibition, thus increasing the susceptibility of HL60 cells to the extrinsic pathway of apoptosis.
The AKT pathway is involved in resistance to natural Killer-induced apoptosis in HL60 cells by regulating the expression of immune suppressor receptors. These findings highlight the importance of AKT in contributing to immune evasion mechanisms in acute myeloid leukemia and suggests the potential of AKT inhibition as an adjunct to immunotherapy.

Aquaporins are a family of ubiquitously expressed transmembrane water channels implicated in a broad range of physiological functions. We have previously reported that aquaporin 4 (AQP4) is expressed on T cells and that treatment with a small molecule AQP4 inhibitor significantly delays T cell mediated heart allograft rejection. Using either genetic deletion or small molecule inhibitor, we show that AQP4 supports T cell receptor mediated activation of both mouse and human T cells. Intact AQP4 is required for optimal T cell receptor (TCR)-related signaling events, including nuclear translocation of transcription factors and phosphorylation of proximal TCR signaling molecules. AQP4 deficiency or inhibition impairs actin cytoskeleton rearrangements following TCR crosslinking, causing inferior TCR polarization and a loss of TCR signaling. Our findings reveal a novel function of AQP4 in T lymphocytes and identify AQP4 as a potential therapeutic target for preventing TCR-mediated T cell activation.
© The Author(s) 2023. Published by Oxford University Press on behalf of Society for Leukocyte Biology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Graft-versus-host disease (GVHD) significantly contributes to patient morbidity and mortality after allogeneic hematopoietic cell transplantation (allo-HSCT). Sphingosine-1-phosphate (S1P) signaling is involved in the biogenetic processes of different immune cells. In the current study, we demonstrated that recipient sphingosine kinase 1 (Sphk1), but not Sphk2, was required for optimal S1PR1-dependent donor T-cell allogeneic responses by secreting S1P. Using genetic and pharmacologic approaches, we demonstrated that inhibition of Sphk1 or S1PR1 substantially attenuated acute GVHD (aGVHD) while retaining the graft-versus-leukemia (GVL) effect. At the cellular level, the Sphk1/S1P/S1PR1 pathway differentially modulated the alloreactivity of CD4+ and CD8+ T cells; it facilitated T-cell differentiation into Th1/Th17 cells but not Tregs and promoted CD4+ T-cell infiltration into GVHD target organs but was dispensable for the CTL activity of allogeneic CD8+ T cells. At the molecular level, the Sphk1/S1P/S1PR1 pathway augmented mitochondrial fission and increased mitochondrial mass in allogeneic CD4+ but not CD8+ T cells by activating the AMPK/AKT/mTOR/Drp1 pathway, providing a mechanistic basis for GVL maintenance when S1P signaling was inhibited. For translational purposes, we detected the regulatory efficacy of pharmacologic inhibitors of Sphk1 and S1PR1 in GVHD induced by human T cells in a xenograft model. Our study provides novel mechanistic insight into how the Sphk1/S1P/S1PR1 pathway modulates T-cell alloreactivity and validates Sphk1 or S1PR1 as a therapeutic target for the prevention of GVHD and leukemia relapse. This novel strategy may be readily translated into the clinic to benefit patients with hematologic malignancies and disorders.
© 2022. The Author(s), under exclusive licence to CSI and USTC.

Despite intensive risk-based treatment protocols, 15% of paediatric patients with B-Cell Precursor Acute Lymphoblastic Leukaemia (BCP-ALL) experience relapse. There is urgent need of novel strategies to target poor prognosis subgroups, like PAX5 translocated.
We considered 289 childhood BCP-ALL cases consecutively enrolled in Italy in the AIEOP-BFM ALL2000/R2006 protocols and we performed extensive molecular profiling, integrating gene expression, copy number analyses and fusion genes discovery by target-capture NGS. We developed preclinical strategies to target PAX5 fusion genes.
We identified 135 cases without recurrent genetic rearrangements. Among them, 59 patients (43·7%) had a Ph-like signature; the remaining cases were identified as ERG-related (26%), High-Hyperdiploid-like (17%), ETV6::RUNX1-like (8·9%), MEF2D-rearranged (2·2%) or KMT2A-like (1·5%). A poor prognosis was associated with the Ph-like signature, independently from other high-risk features. Interestingly, PAX5 was altered in 54·4% of Ph-like compared to 16·2% of non-Ph-like cases, with 7 patients carrying PAX5 fusions (PAX5t), involving either novel (ALDH18A1, IKZF1, CDH13) or known (FBRSL1, AUTS2, DACH2) partner genes. PAX5t cases have a specific driver activity signature, extending to multiple pathways including LCK hyperactivation. Among FDA-approved drugs and inhibitors, we selected Dasatinib, Bosutinib and Foretinib, in addition to Nintedanib, known to be LCK ligands. We demonstrated the efficacy of the LCK-inhibitor BIBF1120/Nintedanib, as single agent or in combination with conventional chemotherapy, both ex vivo and in patient-derived xenograft model, showing a synergistic effect with dexamethasone.
This study provides new insights in high-risk Ph-like leukaemia and identifies a potential therapy for targeting PAX5-fusion poor risk group.
Ricerca Finalizzata-Giovani Ricercatori (Italian Ministry of Health), AIRC, Transcall, Fondazione Cariparo.
Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.