NATure BIoMedIcAl eNgINeerINgdeveloped an approach for in-cell site-specific protein phosphorylation to synthesize bioactive proteins fused with a phosphorylated alum-binding peptide (ABP) tag. We used this approach to produce a series of ABP-labelled cytokines, which rapidly adsorbed to alum after simple mixing, and upon i.t. injection were retained in tumours for more than a week. Applied to the cytokine IL-12, this approach dramatically increased i.t. retention of the cytokine and eliminated systemic toxicities seen upon i.t. injection of the free drug, while also increasing anti-tumour efficacy. Moreover, a single i.t. dose of alum-anchored IL-12 elicited strong IFN-γ-dependent collaboration between innate and adaptive immune cells, producing robust systemic anti-tumour responses in multiple poorly immunogenic preclinical models when combined with systemic checkpoint blockade therapy. ResultsTargeted phosphorylation via an in-cell approach is robust. A single kinase, Fam20C, is responsible for phosphorylation of

Understanding and enabling the control of the properties of foams is important for a variety of commercial processes and consumer products. In these systems, the role of surface active compounds has been the subject of many investigations using a wide range of techniques. The study of their influence on simplified geometries such as two bubbles in a liquid or a thin film of solution (such as in the well-known Scheludko cell), has yielded important fundamental understanding. Similarly, in this work an interferometric technique is used to study the dynamic evolution of the film formed by a single bubble being pressed against a planar air-liquid interface. Here interferometry is used to dynamically measure the total volume of liquid contained within the thin-film region between the bubble and the planar interface. Three different small-molecule, surfactant solutions were investigated and the data obtained via interferometry were compared to measurements of the density of bulk foams of the same solutions. The density measurements were collected with a simple, but novel technique using a conical-shaped bubbling apparatus. The results reveal a strong correlation between the measurements on single bubbles and complete foams. This suggests that further investigations using interferometric techniques can be instrumental to building a more detailed mechanistic understanding of how different surface-active compounds influence foam properties. The results also reveal that the commonly used assumption that surfactant-laden interfaces may be modeled as immobile, is too simplistic to accurately model interfaces with small-molecule surfactants.

Although co-stimulation of T cells with agonist antibodies targeting 4-1BB (CD137) improves antitumor immune responses in preclinical studies, clinical development has been hampered by on-target, off-tumor toxicity. Here, we report the development of a tumor-anchored ɑ4-1BB agonist (ɑ4-1BB-LAIR), which consists of an ɑ4-1BB antibody fused to the collagen binding protein LAIR. While combination treatment with an antitumor antibody (TA99) displayed only modest efficacy, simultaneous depletion of CD4+ T cells boosted cure rates to over 90% of mice. We elucidated two mechanisms of action for this synergy: ɑCD4 eliminated tumor draining lymph node Tregs, enhancing priming and activation of CD8+ T cells, and TA99 + ɑ4-1BB-LAIR supported the cytotoxic program of these newly primed CD8+ T cells within the tumor microenvironment. Replacement of ɑCD4 with ɑCTLA-4, a clinically approved antibody that enhances T cell priming, produced equivalent cure rates while additionally generating robust immunological memory against secondary tumor rechallenge.

SummaryCytokines have long been considered promising cancer immunotherapy agents due to their endogenous role in activating and proliferating lymphocytes. However, since the initial FDA approvals of Interleukin‐2 (IL‐2) and Interferon‐ɑ (IFNɑ) for oncology over 30 years ago, cytokines have achieved little success in the clinic due to narrow therapeutic windows and dose‐limiting toxicities. This is attributable to the discrepancy between the localized, regulated manner in which cytokines are deployed endogenously versus the systemic, untargeted administration used to date in most exogenous cytokine therapies. Furthermore, cytokines' ability to stimulate multiple cell types, often with paradoxical effects, may present significant challenges for their translation into effective therapies. Recently, protein engineering has emerged as a tool to address the shortcomings of first‐generation cytokine therapies. In this perspective, we contextualize cytokine engineering strategies such as partial agonism, conditional activation and intratumoral retention through the lens of spatiotemporal regulation. By controlling the time, place, specificity, and duration of cytokine signaling, protein engineering can allow exogenous cytokine therapies to more closely approach their endogenous exposure profile, ultimately moving us closer to unlocking their full therapeutic potential.

No abstract