Nanoparticle Encapsulated L-Asparaginase

Amino acid depletion with catabolizing enzymes is a potential therapeutic modality for many cancer types. L-asparaginase has been used to treat Acute Lymphoblastic Leukemia (ALL) for many decades and is part of treatment regimen that has a roughly 80% cure rate. The current frontline formulation is L-asparaginase from E.coli that has been conjugated to polyethylene glycol (PEG) for extended circulation half life and protection from immune responses. However, neutralizing antibodies still occur in a subset of patients, necessitating a switch to a different L-asparaginase, such as the less active enzyme from Erwinia chrysanthemi. This complicates therapy, risks severe allergic reactions, and if antibodies develop to the second enzyme the options become limited. The threat of immune reactions, as well as other side effects, has largely restricted L-asparaginase therapy to ALL, despite in vitro evidence that many other cancers might be sensitive to asparagine depletion.

Our group has developed a novel nanoparticle enzyme encapsulation technology that overcomes the limitation of immune responses to enzymes or PEG. These particles, termed Synthetic Hollow Enzyme Loaded Porous nanoShells (SHELS), are produced in a two-step process where the first creates silica shells with large pores through which the enzyme can be loaded, and a second sealing step that encapsulates the mesoporous shell and the loaded enzyme cargo with a nanoporous silica layer. The work below was done with 200nm particles that have a total shell wall thickness of approximately 10nm.

SHELS were loaded with L-asparaginase (Elspar). No loss of activity was seen when enzyme activity was measured in vitro. Groups of mice were injected IM with 5 IU of either free or SHELS encapsulated L-asparaginase. In each case, half of the mice were passively immunized with rabbit polyclonal anti-asparaginase antibodies to a blood concentration of ∼0.5 mg/ml. Serum asparaginase levels pre and post injection were measured by HPLC at 2, 5 and 8 days. Both free and encapsulated enzyme produced complete depletion of asparagine at day 2, but by day 5 asparagine levels had returned to baseline in the animals given free enzyme, whereas those given encapsulated enzyme maintained depletion at day 5 with restoration to baseline at day 8. There was no asparagine depletion in immunized animals with the free L-asparaginase, but SHELS encapsulated asparaginase produced an identical depletion both in the presence or absence of the neutralizing antibodies, indicating that the SHELS effectively shield the enzyme from the antibodies. Experiments in tumor models are ongoing as well as dose response and repeated administration studies.