As biotherapeutics grow and gain increasing importance, great opportunities arise for pharmaceutical development and human health. These large molecule (LM) drugs are poised to offer some revolutionary treatment options. As with conventional pharmaceutical development, the bioanalytical method continues to play a big role here as well.
And therein lies the challenge of large molecule bioanalysis. It has to contend with small molecule drugs. While that isn’t the problem by itself, the larger development of biotherapeutics is relatively recent. That poses some challenges to consider for the bioanalysis of these drugs.

Segment Maturity

Small molecule bioanalysis forms the method for the most widely used set of therapeutics and drugs. It is a mature segment within the pharmaceutical industry and has excellent methods for development and operation.
Large molecules form a relatively younger segment. There is a surge in the development of various biotherapeutics, partially fueled by many drugs going off-patent. In any case, antibody-drug conjugates (ADC), monoclonal antibodies (mAb), fusion proteins, and PEGylated proteins are seeing a big surge.

While there are experts who can handle large molecule bioanalysis, the lack of an extant framework poses some issues. The evolving set of standards and protocols is challenging as well.
This is not to say that the protocols don’t exist and this is an entirely novel field. It is, however, challenging to keep up with the spike in development.

Species Specificity of Large Molecule Drugs

Large molecule drugs are usually extracted from, or semi synthesized from biological sources. This also means that the intended therapeutic action can be species-specific. This can potentially limit the use, study design, and safety evaluation of the biologic before human trial. One way around this problem is by using relevant animal models and transgenic animals where necessary.
There’s also the fact that the delivery methods for large molecule drugs are limited. They cannot be administered orally and are generally delivered intravenously or subcutaneously.

Method Development and Transfer

Method development and transfer are a challenging part of any bioanalytical method. They get more challenging with large molecule bioanalysis due to the complexity involved. The reproducibility and robustness of assays play a big role. Assays for large molecule drugs can be very complex and thus pose additional difficulties in the transfer.

Immunogenicity

Due to the very nature of their development, large molecule drugs are especially at risk of provoking a response from the immune system. The immune system responds with the creation of antibodies that may neutralize the drug. In some cases, the large molecules may create even larger macromolecules and change the immunogenicity profile of the underlying drug.

Quantification

Quantification for large molecule drugs requires special attention to the complex process. Defining various factors for measurement is important and equally important is deciding what factors to measure. The assays to employ in such methods are another consideration.

Ligand Binding Assays (LBA) are perhaps most popular. Some would even consider these as the gold-standard for LM quantification. ELISA assays have proven themselves to be very useful and very popular. Another method used for quantification is liquid chromatography with tandem mass spectrometry. This method often offers a higher dynamic range, multiplexing, and better management and handling of the generated data.