The study, led by a team of scientists from Faculty of Medicine Siriraj Hospital, Mahidol University focused on targeting CD133-expressing CCA cells, which are known to contribute to tumor growth and resistance to current therapies. The researchers engineered T cells to secrete αCD133-αCD3 bispecific T-cell engagers, molecules capable of simultaneously binding CD133 on cancer cells and CD3 on T cells, triggering an immune response against the tumor cells.

To evaluate the effectiveness of this new therapeutic approach, the cDNA encoding the αCD133-αCD3 bispecific T-cell engager was cloned into a lentiviral construct and expressed in T cells obtained from healthy donors. The engineered T cells were then tested for their antitumor activity against CD133-expressing CCA cells.

The results of the study demonstrated promising antitumor effects of the engineered T cells. In co-culture systems, the T cells secreting αCD133-αCD3 engager exhibited significantly higher cytolytic activity against CD133-expressing CCA cells compared to untransduced T cells. At an effector-to-target ratio of 5:1, the engineered T cells showed cytolytic activity of 49.0±9.76% and 64.10±13.18% against KKU-100 and KKU-213A CCA cells, respectively, compared to only 10.97±10.65% and 9.80±11.05% for untransduced T cells.

Importantly, the secreted αCD133-αCD3 engager not only enhanced the killing activity of the engineered T cells but also redirected bystander T cells to attack the target CCA cells. The combination of transduced and bystander T cells resulted in a remarkable 73.20±1.68% cytolytic activity against CCA cells, demonstrating the potential for a potent immune response.

Furthermore, the engineered T cells and bystander T cells exhibited enhanced killing activity against CCA spheroids, three-dimensional structures that mimic tumor behavior, with a rate approximately five times higher than the control condition without treatment.

These findings provide compelling proof-of-principle that T cells secreting αCD133-αCD3 engager hold promise as an alternative therapeutic approach for CD133-positive CCA. The successful preclinical results pave the way for further investigation in animal models and future clinical trials, bringing hope for a much-needed effective treatment option for CCA patients.