Two-Dimensional Liquid Chromatography (2D-LC) for Biopharmaceuticals

Abstract

Biopharmaceuticals, such as monoclonal antibodies (mAbs), interferons/cytokines, vaccines etc contribute progressively more to clinical practice. They are complex macromolecules produced from living cells by means of biotechnology, usually produced to treat lifethreatening diseases, such as tumors, autoimmune diseases, diabetes, etc. Compared to traditional drugs, biopharmaceuticals are much larger in size, which ranges from 2,000 to 2,000,000 Daltons, while traditional pharmaceuticals are typically within the range 100-1500 Daltons. Another significant difference between them is the number of active sites (functional groups) in biopharmaceuticals, which is significantly higher usually 10-2000. All these characteristics make their investigation of great analytical challenge. Due to their complexity, biopharmaceuticals require multiple modes of chromatographic separations. Moreover the fact that exact copies of biopharmaceuticals cannot be produced; this results to the production of biosimilars that refer to drugs with similar physicochemical characteristics, as well as efficacy and safety with the originators. The ongoing development in biosimilar manufacturing has led to the demand for complementary analytical methods in order to achieve the efficient comparison with originators. At present there are more than 200 approved biopharmaceuticals in the market and this is predicted to increase. Although monoclonal antibodies are the prevailing biopharmaceuticals, novel drugs like antibody drug conjugates (ADCs) have become of particular oncological interest. These are capable to deliver the chemotherapeutic cytotoxic agent directly to the tumor site antigen. By this approach the risk of damaging healthy tissues is intensely reduced. Thereby they take advantage of the benefits of large molecule specificity with small molecule toxicity [1-5].