Single chain variable fragment as probe

Single chain variable fragment (scFv) Abs are one of the recombinant antibody(rAb)fragments, which are popular therapeutic alternatives to full length of monoclonal Abs. Compared to generating whole Abs from animal cell culture, scFv are smaller and can be expressed rapidly, economically and in large quantities in a bacterial host, such as E. coli. A scFv possesses the complete antigen binding site, which contains the variable heavy (VH) and variable light domain of an antibody. The VH domain is linked to a VL domain by an introduced flexible polypeptide linker. A scFv is capable of binding its target antigens with an affinity similar to that of the parent mAb. Due to containing the specific antigen binding unit, scFv fragments show tremendous versatility and importance in human therapeutics and diagnostics. ^[1] In addition, scFv fragments can be envisaged to be applied in the non-pharmaceutical sector, such as in food, cosmetic or environmental industries. The unique and highly specific antigen-binding ability might, for example, be exploited to block specific enzymes (e.g. enzymes that cause food spoilage), bacteria (e.g. in toothpaste or mouthwashes) or to detect environmental factors present in very low concentrations (as biosensors).^[2]

Figure 1.

Properties and development of targeted drugs

This year, we decided to utilize the scFv as probes to detect cancer markers and aid in the prescription of targeted drugs in cancer treatments. Targeted drugs therapy utilize compounds that are capable of inhibiting target molecules, the cancer markers which send messages along signaling pathways in cell growth, cell division or cell death.^[3] Via specific binding to target molecules, targeted drugs show more accurate attack to cancer cells and less harmful damage to normal tissues. ^[4] The precision of targeting the cancer cells has enhanced the efficiency of treatment by a large margin. The targeted therapy is a major step forward for many cancers, especially advanced cancers, and physicians and researchers are now focusing on the development of targeted drugs, creating a new era of personalized cancer treatment.^[5]Targeted therapy are so-called "personalized medicine" because health care professionals can use clinical test results from a patient to select a specific drug that has a higher likelihood of being effective for that particular person.

Figure 2. The usage of targeted drugs.

According to the statistics, the usage rate of targeted drug therapy has increased within ten years. In Figure 2, in 2003, targeted drug therapy is not commonly used compared with other therapies, accounting for only 11% usage. Over one decade, it is estimated that the usage of targeted drug therapy dramatically increases to 46%. It indicates targeted drugs therapy is a potential growing field and will become the commonly used therapy in cancer treatments in the near future.

Pre-diagnosis of targeted drugs treatment

To create the new era of tailored targeted drugs, doctors must aim at appropriate target molecules for patients with particular diseases. In 2014, the U.S. Food and Drug Administration (FDA) issued a guidance to facilitate the development and review of diagnostics tests. The diagnostics tests are the steps to identify the abnormal cancer biomarkers. Moreover, the purpose of diagnostics tests are to help medical practitioners determine which patients could benefit from the certain drugs, conversely, those who should not receive the medication. If the treatment decisions is not optimal, it would not only cause the fatal body damage, but also lead to the waste of time, money and medical resources. FDA encourages the joint of targeted drugs therapies and precise diagnostics tests which are essential for the safe and effective use of targeted drugs.^[6]

The concept of combination therapy

Although targeted drugs treatments can lead to the dramatic regressions of solid tumors, the responses are often short-lived because resistant cancer cells arise after a period of treatment. The major strategy proposed for overcoming the resistance is combination therapy.^[7] The clinical and preclinical researches further indicated that targeted drug therapy combined with another targeted drug therapy or other types of therapies to treat cancers simultaneously may attain greater effects than using only one therapy. With the concept of combination therapy, we can not only improve the treating effect but also reduce the occurrence of cancer cells resistance toward the targeted drugs as there are less probability that a single mutation will cause cross-resistance to both drugs.^[8]

APPOllO E.Cotector

To enhance the efficiency of diagnosis and provide reference for proper usage of targeted drugs and combination therapy, we come up with the idea of detecting multimarker at the same time and this was how our marvelous E.Cotector is borned. This year, NCTU_Formosa commits to creating a multimarker diagnosis platform via scFv as probes for helping physicians to determine and prescribe the usage of targeted drugs in cancer patients, especially the monoclonal-antibody-targeted drugs.

Customized Detection Platform

This year, APOllO, introduce a revolutionary product, a customized detecting platform. We call it the E.Cotector.

In our product, we built three individual biobrick libraries of scFv probes and color signal and Gold Binding Polypeptide (GBP). Probes detect the target we want. Fluorescence protein or chromoprotein expresses signal for observation. GBP acts as a bridge to connect our product to gold, which enables our product to be applied to biosensors. Our customers can spontaneously choose from our libraries, pair up any biobricks. Next, comes the most crucial part of our project. Our team will then co-transform the plasmids into E. coli. Our customized detecting platform, The APOllO E.Cotector, is therefore born. By co-transforming plasmids, it not only helps us customize our product every which way for our customers but also brings us a major advantage in our manufacturing procedure. We no longer have to build lengthy biobricks, which tremendously increases our manufacturing efficiency.

Health and Medical Detection Platform

With our hit product, E.Cotector, we can focus on health and medical dimension. We want to provide a convenient and efficient diagnosis for prescribing personalized targeted drug therapy via tissue biopsies. Therefore, we chose single chain variable fragments (scFv), which are directly from monoclonal antibody, targeted drugs. ScFv acts as our probe to detect specific biomarkers. Our customer can pair up any scFv with any color signal. By changing various kinds of scFv on the surfaces of E.Cotector, it can identify multimarkers and suggest appropriate different kinds of targeted drugs.

Our E.Cotector can consequently bind to various targeted antigen also simultaneously express color as signal for instant observation. We provide another plasmid of Gold Binding Polypeptide for customers to pair up. Pairing up scFv and GBP can achieve quantitative detection as a biosensor. In conclusion, the customized APOllO E.Cotector will bring a brand new era for pre-diagnosis target drugs treatment.