Preliminary Exploration of Simple BH3 Profiling in Screening Drug Susceptibility of Blood Tumors

Objective To establish a simple BH3 profiling for drug-sensitive screening of anti-apoptotic protein inhibitors of the BCL-2 family in blood tumors.

Method To establish the BH3 profiling method based on the microscope and JC-1 staining, describe the dependence of K562 and NB4 cell lines on anti-apoptotic proteins of the BCL-2 family by BH3-only simulated peptide, and verify it by polymerase chain reaction (PCR) method. Clinical drug efficacy was retrospectively reviewed to check the conformity of BH3 drug sensitivity analysis results.

Results BH3 profiling results showed that the anti-apoptotic protein dependence of the NB4 cell line was BCL2 > MC1-1 > BCL-XL; the K562 cell line was BCL-XL > MC1-1 > BCL2. PCR analysis of BCL-2 family expression showed that anti-apoptotic proteins MCL-1, BCL2, and BCL-XL were highly expressed in K562, while BIM, NOXA, and PUMA were highly expressed in BH3-only protein and HRK was low. The anti-apoptotic ability of cells depended more on BCL-XL. In NB4, the anti-apoptotic protein BCL-XL was significantly lower than MCL-1 and BCL-2, while NOXA in BH3-only protein was lower. NB4 cells were more dependent on MCL-1 and BCL-2 than BCL-XL; Which was consistent with BH3 profiling results. Bone marrow BH3 profiling results of 6 patients with hematologic tumors showed that HRK (BCL-XL inhibitor) and VEN (BCL-2 inhibitor) were not sensitive. NOXA (MCL-1 inhibitor) was sensitive to examples 1, 2, 3 and 5, while the sensitivity of examples 4 and 6 was low (< 50%). It was speculated that all 6 cases were resistant to BCL-2 inhibitor; Cases 1, 2, 3 and 5 were sensitive to MCL1 inhibitor, and cases 4 and 6 might be resistant to MCL1 inhibitor. Retrospective clinical efficacy (4~6 sessions): Cases 1~5 were not sensitive to the treatment regimen containing BCL-2 inhibitor, which was considered to be related to insensitivity to BCL-2 inhibitor. Chidamide was effective in case 5, but not effective in cases 4 and 6, which were considered to be related to MCL-1 inhibitor insensitivity. The above clinical outcomes were consistent with the predicted results of BH3 profiling.

Conclusions BH3 profiling based on fluorescence microscopy and JC-1 staining is simple and feasible and can be used to detect the dependence of blood tumor cells on different anti-apoptotic proteins, thus providing evidence for the selection of inhibitors related to the Bcl-2 family.