Degradable Hydrogel for Sustained Localized Delivery of Anti-Tumor Drugs.

Disadvantages of systemically administered immunomodulatory anti-tumor therapies include poor efficacy and high toxicity. Direct intratumoral injection of a drug is often associated with rapid efflux from the site of administration, thus reducing local exposure and therapeutic efficacy, while potentially increasing systemic adverse events. To address this, a sustained release prodrug technology was developed using a transient conjugation (TransConTM) technology to provide long-term high local drug exposure after injection in the tumor while minimizing systemic exposure. TransCon technology for systemic delivery is clinically validated, with multiple compounds in late-stage clinical development and approval of a once-weekly growth hormone for pediatric growth hormone deficiency. As a further application of this technology, this report describes the design, preparation, and functional characterization of hydrogel microspheres as insoluble, yet degradable carrier system. Microspheres were obtained after reaction of PEG-based polyamine dendrimers and bifunctional crosslinkers. Resiquimod, a TLR7/8 agonist, and axitinib, a vascular endothelial growth factor tyrosine kinase inhibitor, were chosen as anti-cancer drugs. The drugs were covalently attached to the carrier by linkers, which released the drugs under physiological conditions. Essentially all resiquimod or axitinib was released over weeks before physical degradation of the hydrogel microsphere was observed. In summary, TransCon Hydrogel technology allows localized sustained-release drug delivery for cancer therapy enabling high local drug concentrations while at the same time ensuring low systemic drug exposure over weeks with a single injection, which may improve the therapeutic index and improve efficacy, while minimizing systemic adverse events. A hydrogel prodrug of resiquimod, TransCon TLR7/8 agonist, is currently being investigated in clinical trials of patients with solid tumors (NCT04799054).

Intratumoral delivery of TransCon™ TLR7/8 Agonist promotes sustained anti-tumor activity and local immune cell activation while minimizing systemic cytokine induction.

BACKGROUND: Intratumoral (IT) delivery of toll-like receptor (TLR) agonists has shown encouraging anti-tumor benefit in preclinical and early clinical studies. However, IT delivery of TLR agonists may lead to rapid effusion from the tumor microenvironment (TME), potentially limiting the duration of local inflammation and increasing the risk of systemic adverse events. METHODS: To address these limitations, TransCon™ TLR7/8 Agonist-an investigational sustained-release prodrug of resiquimod that uses a TransCon linker and hydrogel technology to achieve sustained and predictable IT release of resiquimod-was developed. TransCon TLR7/8 Agonist was characterized for resiquimod release in vitro and in vivo, in mice and rats, and was assessed for anti-tumor efficacy and pharmacodynamic activity in mice. RESULTS: Following a single IT dose, TransCon TLR7/8 Agonist mediated potent tumor growth inhibition which was associated with sustained resiquimod release over several weeks with minimal induction of systemic cytokines. TransCon TLR7/8 Agonist monotherapy promoted activation of antigen-presenting cells in the TME and tumor-draining lymph nodes, with evidence of activation and expansion of CD8+ T cells in the tumor-draining lymph node and TME. Combination of TransCon TLR7/8 Agonist with systemic immunotherapy further promoted anti-tumor activity in TransCon TLR7/8 Agonist-treated tumors. In a bilateral tumor setting, combination of TransCon TLR7/8 Agonist with systemic IL-2 potentiated tumor growth inhibition in both injected and non-injected tumors and conferred protection against tumor rechallenge following complete regressions. CONCLUSIONS: Our findings show that a single dose of TransCon TLR7/8 Agonist can mediate sustained local release of resiquimod in the TME and promote potent anti-tumor effects as monotherapy and in combination with systemic immunotherapy, supporting TransCon TLR7/8 Agonist as a novel intratumoral TLR agonist for cancer therapy. A clinical trial to evaluate the safety and efficacy of TransCon TLR7/8 Agonist, as monotherapy and in combination with pembrolizumab, in cancer patients is currently ongoing (transcendIT-101; NCT04799054).

Synthesis and conformational analysis of stevastelin C3 analogues and their activity against the dual-specific vaccina H1-related phosphatase.

The biological activity of macrocyclic natural products depends on their conformational properties. For both the elucidation of enzyme binding affinities as well as the development of selective drugs, rigid macrocyclic scaffolds carry high potential. In this study, 13-membered cyclodepsipeptides based on the structure of naturally occurring stevastelins were studied in detail. Six diastereomeric stevastelin C3 analogues and four phosphorylated derivatives were synthesized. The synthesis of linear precursors was achieved on solid support by starting from stereoisomerically pure 2-methyl-3-hydroxy acids. Subsequent macro-lactamization gave the cyclic depsipeptides in very good yields (36-62%). The conformational space of these stevastelin C3 analogues was computationally investigated. On the basis of NMR spectroscopic data, homogeneous conformations were determined for each benzylated depsipeptide and the influence of phosphorylation on the overall conformation was investigated. Importantly, phosphorylation was found to significantly weaken the conformational preferences of the 13-membered depsipeptides. Finally, the cyclic depsipeptides were tested for activity against phosphatases. Inhibitory activity on vaccina H1-related phosphatase was observed depending on the derivatization of the cycles. The activity profiles are discussed in the light of the structural data.

Understanding protease catalysed solid phase peptide synthesis.

A protease (thermolysin) was used to directly synthesise a number of dipeptides from soluble Fmoc-amino acids onto a solid support (PEGA1900) in bulk aqueous media, often in very good yields. This shift in equilibrium toward synthesis is remarkable because for soluble dipeptides in aqueous solution hydrolysis rather than synthesis is observed. Three possible reasons for the equilibrium shift were considered: (i) using a solid support makes it easy to use an excess of reagents, so mass action contributes towards synthesis; (ii) reduction in the unfavourable hydrophobic hydration of the Fmoc group within the solid support compared with the free amino acid in solution and (iii) suppression of the ionization of amino groups linked to the solid phase due to mutual electrostatic repulsion. It was found that under the conditions studied the second effect was most important.

Enzymatic optical resolution via acylation-hydrolysis on a solid support.

By taking advantage of the reversibility of the thermolysin-catalysed amide synthesis-hydrolysis reaction on a solid support, both L,L and L,D diastereoisomers of dipeptides and L-amino acids are accessible in good yields starting from enantiomeric mixtures of amino acids.