Response Monitoring of Acute Lymphoblastic Leukemia Patients Undergoing l-Asparaginase Therapy: Successes and Challenges Associated with Clinical Sample Analysis in Plasmonic Sensing

Abstract

Monitoring the response of patients undergoing chemotherapeutic treatments is of great importance to predict remission success, avoid adverse effects and thus, maximize the patients’ quality of life. In the case of leukemia patients treated with E. coli l-asparaginase, monitoring the immune response by the detection of specific antibodies to l-asparaginase in the serum of patients can prevent extended immune response to the drug. Here, we developed a surface plasmon resonance (SPR) biosensor to rapidly detect anti-asparaginase antibodies directly in patients’ sera, without requiring sample pretreatment or dilution. A direct assay with SPR sensing to detect anti-asparaginase antibodies exhibited a limit of detection of 500 pM and a high sensitivity range between 100 nM and 1 μM in pooled and undiluted human serum from a commercial source. While the SPR assay showed excellent reproducibility (12% RSD) in pooled serum, challenges were encountered upon analyzing clinical samples due to high sample-to-sample variability in color and turbidity and, in all likelihood, in composition. As a result, direct detection in clinical samples was unreliable due to factors that may generally affect assays based on plasmonic detection. Addition of a secondary detection step overcame sample variability due to bulk differences in patients’ sera. By those means, the SPR biosensor was successfully applied to the analysis of clinical samples from leukemia patients undergoing asparaginase treatments and the results agreed well with the standard ELISA assay. Monitoring antibodies against drugs is common such that this type of sensing scheme could serve to monitor a plethora of immune responses in sera of patients undergoing treatment.

Development of Escherichia coli Asparaginase II for Immunosensing: A Trade-Off between Receptor Density and Sensing Efficiency

“…Although the His-tagged EcAII displayed lower sensing efficiency than the native EcAII, it provided increased surface coverage and reproducibility of immobilization, ultimately procuring significantly increased immunodetection sensitivity. These results shed light on the challenges encountered in our recently reported detection of serum anti-asparaginase antibodies in the sera of children undergoing chemotherapy 23 and the challenges expected to be encountered by others developing sensors to monitor the immunogenic response of patients undergoing therapy with biologic-type drugs.…”

“…Calibration of the sensor was performed with serial injections of increasing concentrations of anti-asparaginase antibodies on a single sensor chip, as previously described. 23 The SPR shifts were calculated with MATLAB software and served to calculate the surface density of the analyte bound onto the immobilized antigenic receptors, as described above. Taking into account the exact MW of each EcAII variant, we calculated the number of EcAII molecules immobilized on the sensing surface (molecule cm –2 ).…”

The Fundamentals of Real‐Time Surface Plasmon Resonance/Electrogenerated Chemiluminescence

“…Hence, it was revealed that SPR and ECL are complementary and offer similar performances for the detection of antibodies . SPR sensors provide quantitative real‐time information on the adsorption of molecules to a surface, although the analysis of clinical samples in biofluids may be difficult . ECL provides high sensitivity, high selectivity, and low detection limits even in crude matrices as it is relatively insensitive to nonspecific adsorption of biofluids, but lacks information on the adsorption processes occurring on the electrode.…”

The Fundamentals of Real‐Time Surface Plasmon Resonance/Electrogenerated Chemiluminescence

“…[3] SPR sensors provide quantitative real-time information on the adsorption of molecules to as urface,a lthough the analysis of clinical samples in biofluids may be difficult. [4] ECL provides high sensitivity,h igh selectivity,a nd low detection limits even in crude matrices as it is relatively insensitive to nonspecific adsorption of biofluids,b ut lacks information on the adsorption processes occurring on the electrode.H ence,t he combination of SPR and ECL in as ingle instrument would be beneficial to monitor interfacial processes involved in ECL and to design biosensors with improved performances. To fully benefit from the combination of the techniques, at horough understanding of the underlying principles is needed.…”