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Application of conventional chemotherapy regardless of its unique effectiveness have been gradually being edged aside due to limited targeting capability, lack of selectivity and chemotherapy-associated side effects. To this end, colon-targeted nanoparticles  via  combination therapy have shown great therapeutic potential against cancer. Herein, pH/enzyme-responsive biocompatible polymeric nanohydrogels based on poly(methacrylic acid) (PMAA) containing methotrexate (MTX) and chloroquine (CQ) were fabricated. PMAA-MTX-CQ exhibited high drug loading capacity of which MTX was 4.99% and was CQ 25.01% and displayed pH/enzyme-triggered drug release behavior. Higher CQ release rate (76%) under simulated acidic microenvironment of tumor tissue whereas 39% of CQ was released under normal physiological conditions. Intestinally, MTX release was facilitated in the presence of proteinase K enzyme. TEM image demonstrated spherical morphology with particle size of less than 50 nm.  In vitro  and  in vivo  toxicity assessments indicated that developed nanoplatforms possessed great biocompatibility. These nanohydrogels did not cause any adverse effects against  Artemia Salina  and HFF2 cells (around 100% cell viability) which highlight the safety of prepared nanohydrogels. There was no death in mice received different concentrations of nanohydrogel through oral administration and less than 5% hemolysis was found in red blood cells incubated with PMAA nanohydrogels.  In vitro  anti-cancer results showed that combination therapy based on PMAA-MTX-CQ can effectively suppress the growth of SW480 colon cancer cells (29% cell viability) compared to monotherapy. Altogether, these findings suggest that pH/enzyme-responsive PMAA-MTX-CQ could effectively inhibit cancer cell growth and progression  via  site-specific delivery of its cargo in a safe and controlled manner.

Targeted co-delivery of methotrexate and chloroquine via a pH/enzyme-responsive biocompatible polymeric nanohydrogel for colorectal cancer treatment