One-Pot Identification of BCR/ABL^p210 Transcript Isoforms Based on Nanocluster Beacon

Area of Science:

• Biochemistry
• Molecular Biology
• Medical Diagnostics

Background:

• The BCR/ABL fusion gene, particularly the p210 subtype, is a hallmark of chronic myeloid leukemia (CML).
• Distinct BCR/ABL^p210 isoforms, e13a2 and e14a2, exhibit varied clinical behaviors, impacting treatment and prognosis.
• Accurate and rapid differentiation of these isoforms is crucial for personalized CML management.

Purpose of the Study:

• To develop a sensitive and simultaneous detection method for e13a2 and e14a2 BCR/ABL^p210 isoforms.
• To utilize nanocluster beacon (NCB) fluorescence for distinguishing between the two major CML-associated fusion gene subtypes.
• To establish a rapid diagnostic tool for CML isoform identification in clinical settings.

Main Methods:

• Development of a DNA-templated silver nanocluster (AgNCs) system as a molecular reporter.
• Design of specific thymine-rich (T-rich) and guanine-rich (G-rich) enhancer sequences for isoform recognition.
• Exploitation of proximity-induced fluorescence of AgNCs upon binding to specific isoform-enhancer complexes, forming three-way junction structures.
• Distinguishing isoforms based on unique fluorescence emission wavelengths generated by AgNCs.

Main Results:

• Successful one-pot identification of e13a2 and e14a2 isoforms using the NCB fluorescence strategy.
• Achieved high sensitivity with detection limits of 16 pM for e13a2 and 9 pM for e14a2.
• Demonstrated effective isoform identification in leukemia cells and within 40 minutes for newly diagnosed CML patients.

Conclusions:

• The developed NCB fluorescence strategy offers a powerful and rapid tool for simultaneous differentiation of BCR/ABL^p210 fusion gene isoforms.
• This method provides a significant advancement for precise diagnosis and subtype classification in chronic myeloid leukemia.
• The approach holds promise for improved clinical decision-making and patient management in CML.