A novel anti-cancer L19-interleukin-12 fusion protein with an optimized peptide linker efficiently localizes in vivo at the site of tumors

Ongaro, Tiziano; Matasci, Mattia; Cazzamalli, Samuele; Gouyou, Baptiste; Luca, Roberto De; Neri, Dario; Villa, Alessandra

doi:10.1016/j.jbiotec.2018.12.004

Cited by 26 publications

(23 citation statements)

References 42 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Indeed, pharmacological approaches similar to the ones described in this article for L19-TNF may be applicable to other TNF-based pharmaceuticals or to other cytokine-based products. In particular, toxicities related to the administration of IL12- or IL2-antibody products [16, 46, 47] might benefit from the transient and selective inhibition of key intracellular mediators of their activity.…”

Section: Discussionmentioning

confidence: 99%

“…L19-murine TNF (L19-mTNF) gene was cloned into the mammalian cell expression vector pcDNA3.1(+) (Invitrogen) by HindIII/NotI restriction sites. The L19-mTNF fusion protein was expressed by transient gene expression in CHO-S cells and purified from the cell culture supernatant to homogeneity by protein A (Sino Biological) chromatography, as described previously [46]. After dialyses into PBS pH 7.4, the quality of the proteins was assessed by SDS-PAGE, by Size-exclusion chromatography on a Superdex 200 Increase 10/300 GL column on an ÄKTA FPLC (GE Healthcare) and by mass spectrometry (Waters Xevo G2XS Q-TOF) (Supplementary Information).…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Targeted enhancement of the therapeutic window of L19-TNF by transient and selective inhibition of RIPK1-signaling cascade

et al. 2019

Self Cite

View full text Add to dashboard Cite

IntroductionCytokine-based products are gaining importance for cancer immunotherapy. L19-TNF is a clinical-stage antibody-cytokine fusion protein that selectively accumulates to tumors and displays potent anticancer activity in preclinical models. Here, we describe an innovative approach to transiently inhibit off-target toxicity of L19-TNF, while maintaining antitumor activity.MethodsGSK’963, a potent small molecule inhibitor of RIPK1, was tested in tumor-bearing mice for its ability to reduce acute toxicity associated with TNF signaling. The biological effects of L19-TNF on tumor cells, lymphocytes and tumor vessels were investigated with the aim to enable the administration of TNF doses, which would otherwise be lethal.ResultsTransient inhibition of RIPK1 allowed to increase the maximal tolerated dose of L19-TNF. The protective effect of GSK’963 did not affect the selective localization of the immunocytokine to tumors as evidenced by quantitative biodistribution analysis and allowed to reach high local TNF concentrations around tumor blood vessels, causing diffused vascular shutdown and hemorrhagic necrosis within the neoplastic mass.ConclusionsThe selective inhibition of RIPK1 with small molecule inhibitors can be used as a pharmaceutical tool to transiently mask TNF activity and improve the therapeutic window of TNF-based biopharmaceuticals. Similar approaches may be applicable to other pro-inflammatory cytokines.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Targeted enhancement of the therapeutic window of L19-TNF by transient and selective inhibition of RIPK1-signaling cascade

et al. 2019

Self Cite

View full text Add to dashboard Cite

show abstract

“…In parallel, the concentration of product in the solid tumor mass increases over time, in a process that slows down with the progressive disappearance of antibody-cytokine fusion from circulation. Products targeting stable antigens (e.g., components of the modified tumor extracellular matrix) exhibit long residence times within the neoplastic mass [59,60]. It is therefore conceivable that by fractionating the administration of antibody-cytokine fusions, we may achieve a progressive build-up of product concentration within the solid tumor mass, while keeping blood levels below a critical threshold.…”

Section: Figure 1e Depicts a Biodistribution Rationale For Using Intrmentioning

confidence: 99%

Comparative evaluation of bolus and fractionated administration modalities for two antibody-cytokine fusions in immunocompetent tumor-bearing mice

Puca

Luca

Seehusen

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

Antibody-cytokine fusion proteins are being considered as biopharmaceuticals for cancer immunotherapy. Tumor-homing cytokine fusions typically display an improved therapeutic activity compared to the corresponding unmodified cytokine products, but toxicity profiles at equivalent doses are similar, since side effects are mainly driven by the cytokine concentration in blood. In order to explore avenues to harness the therapeutic potential of antibody-cytokine fusions while decreasing potential toxicity, we compared bolus and fractionated administration modalities for two tumor-targeting antibody-cytokine fusion proteins based on human interleukin-2 (IL2) and murine tumor necrosis factor (TNF) (i.e., L19-hIL2 and L19-mTNF) in two murine immunocompetent mouse models of cancer (F9 and C51). A comparative quantitative biodistribution analysis with radio-labeled protein preparations revealed that a fractionated administration of L19-hIL2 could deliver comparable product doses to the tumor with decreased product concentration in blood and normal organs, compared to bolus injection. By contrast, L19-mTNF (a product that causes a selective vascular shutdown in the tumor) accumulated most efficiently after bolus injection.Fractionated schedules allowed the safe administration of a cumulative dose of L19-mTNF, which was 2.5-times higher than the lethal dose for bolus injection. Dose fractionation led to a prolonged tumor growth inhibition for F9 teratocarcinomas, but not for C51 colorectal tumors, which responded best to bolus injection. Thus, dose fractionation may have different outcomes for the same antibody-cytokine product in different biological contexts.

show abstract

“…Aptamers have several advantages over protein antibodies, including very small molecular weight (5-40 kDa), rapid plasma clearance, non-immunogenicity, easy tissue penetration, high binding affinity and specificity, and the ease of synthesis 28. Aptamers also display remarkable stability under various pH and temperature conditions, as well as in a wide variety of organic solvents 29, 30. Moreover, the mass production of aptamers is simpler and cost-effective, compared to that of proteins.…”

Section: Introductionmentioning

confidence: 99%

Enhanced cytotoxic T lymphocytes recruitment targeting tumor vasculatures by endoglin aptamer and IP-10 plasmid presenting liposome-based nanocarriers

et al. 2019

View full text Add to dashboard Cite

Background : Adequate recruitment of highly active tumor antigen-specific cytotoxic T lymphocytes (CTLs) remains a major challenge in cancer immunotherapy. Objective : To construct liposome (LP)-based nanocapsules with surface endoglin aptamer (ENG-Apt) encapsulating mouse interferon-inducible protein-10 (mIP-10), with the ability to target mouse tumor vascular endothelial cells (mTECs) and enhance CTLs targeting and recruitment to the tumor vasculature. Methods : ENG-Apt/mIP-10-LP nanocapsules were prepared by grafting DSPE-PEG 2000 -ENG-Apt on the surface of liposomes containing mIP-10 plasmids, characterized and assessed for the cell binding specificity in vitro . The tumor-targeting ability of ENG-Apt/mIP-10-LP nanocapsules was evaluated in vivo . The anti-tumor efficacy of ENG-Apt/mIP-10-LP nanocapsules treatment, as well as the combination treatment of ENG-Apt/mIP-10-LP nanocapsules and adoptive TRP2CD8 + T cells, were both tested in melanoma-bearing mice, by evaluation of the tumor volume and the mouse survival time. To discuss the anti-tumoral mechanism of ENG-Apt/mIP-10-LP nanocapsules-based therapies, IFN-γ secretion, proportion of TRP2CD8 + T cells among TILs, MDSCs in the tumor microenvironment and Tregs in the spleen, were determined after the treatments. Proliferation and apoptosis of tumor cells, and tumor angiogenesis were also assessed. Results : The prepared ENG-Apt/mIP-10-LP nanocapsules possess an adequate nanometric size, good stability, high specificity to mTECs and tumor sites, along with the ability to induce mIP-10 expression in vitro and in vivo . Treatment of ENG-Apt/mIP-10-LP nanocapsules demonstrated CTLs enrichment into the tumor site, which inhibited tumor cell proliferation and angiogenesis, as well as promoted tumor-cell apoptosis, leading to a decrease in tumor progression and prolonged survival time in melanoma tumor-bearing mice. In addition, the proportion of MDSCs and Tregs was found to decrease. The combination of ENG-Apt/mIP-10-LP nanocapsules with adoptive TRP2CD8 + T cells, showed stronger abilities in inhibiting tumor growth and increasing animal survival time, thereby displayed an enhanced anti-melanoma tumor efficacy, due to the recruitment of both endogenous CD8 + T cells and exogenous TRP2CD8 + T cells in vivo . Conclusion : ENG-Apt/mIP-10-LP nanocapsules could enhance the recruitment of both endogenous and exogenous CTLs specifically targeting melanoma tumor vasculatures and exert anti-tumoral effect, therefore provides a potentially novel strategy for tumor immunotherapy.

show abstract

A novel anti-cancer L19-interleukin-12 fusion protein with an optimized peptide linker efficiently localizes in vivo at the site of tumors

Cited by 26 publications

References 42 publications

Targeted enhancement of the therapeutic window of L19-TNF by transient and selective inhibition of RIPK1-signaling cascade

Targeted enhancement of the therapeutic window of L19-TNF by transient and selective inhibition of RIPK1-signaling cascade

Comparative evaluation of bolus and fractionated administration modalities for two antibody-cytokine fusions in immunocompetent tumor-bearing mice

Enhanced cytotoxic T lymphocytes recruitment targeting tumor vasculatures by endoglin aptamer and IP-10 plasmid presenting liposome-based nanocarriers

Contact Info

Product

Resources

About