Ex�vivo expansion of human peripheral blood natural killer cells and cytotoxic T lymphocytes from lung cancer patients

Abstract

Lung cancer is the most common type of cancer with the highest cancer-associated mortality rates worldwide, as well as in Vietnam. Numerous studies have demonstrated that higher numbers and higher rate of activity of infiltrating natural killer (NK) cells and cytotoxic T lymphocytes (CTLs) in the tumor are closely correlated with positive prognosis, tumor size decrease and longer survival of lung cancer patients. In the present study, the effectiveness of BINKIT® kit in the ex vivo expansion of NK cells and CTLs in the peripheral blood of 7 patients aged between 30 and 84 years with metastatic lung cancer was evaluated. After 21 days of culture, the average number of CTLs (CD3+CD8+) increased by 742.3-fold in the CTL culture, accounting for 72.2% of the cultured cell population, and the mean cell viability was 95.7%. In the NK cell culture, the average number of NK cells (CD3−CD56+) increased by 637.5-fold, accounting for 84.3% of the cultured cell population, with an average viability of 94.7%. The percentage of active NK cells (CD3−CD56+ bright) was 82.1%, which increased by 408.9-fold. Notably, a close correlation was identified between the numbers of cytokine-induced killer (CD3+CD56+) and NK (CD3−CD56+) cells in the NK cell culture (P<0.05). In the two culture conditions (namely NK cell and CTL cultures), no clear correlation was identified between the rate of initial immune cells in the peripheral blood and the corresponding number following ex vivo expansion (P>0.05). These results revealed that the method of expansion and activation of NK cells and CTLs from peripheral blood was successfully applied using BINKIT, and reached the requirements for clinical applications in cancer treatment in Vietnam.

Phase I Clinical Trial Using Autologous Ex Vivo Expanded NK Cells and Cytotoxic T Lymphocytes for Cancer Treatment in Vietnam

“…Based on the method we had already developed [26], the immune cell expansion in this cohort study was carried out for three types of cancer (lung, liver, and colon), with a total of 29 sitting across patients. Compared to the literature, our results demonstrated that we were successful in expanding NK cells and CTLs with high average numbers of 2570.1 × 10 6 and 4105.9 × 10 6 , respectively.…”

“…More importantly, the mean survival time for patients in our study was 18.7 months at 12 months after the transplantation, hence it increased more than twice compared to the estimated survival time at the point of enrollment. From a published report in early 2018 by Hoang et al [26], the mean survival time of seven lung cancer patients were assessed at 17.1 ± 2 months and increased to 19.4 ± 1.7 months after treatment. The 1-year survival rate of our study was 80%, which was higher compared to other reports previously published [36,37].…”

“…The method for immune cell expansion had already been published [26]. In summary, peripheral blood mononuclear cells (PBMNCs) were obtained by density gradient centrifugation.…”

“…The combination of two kinds of immune cells, including NK cells and CTLs, promises to be advantageous in the detection and killing of cancer cells with or without MHC-I expression. Many clinical studies have demonstrated the efficacy of the combination between these two kinds of immune cells in pre-clinical research models and clinical trials [5,23,25,26,27,28,29,30,31,32]. Expansion of immune cells by using BINKIT, developed by Dr. Terunuma Hiroshi (Biotherapy Institute of Japan, Tokyo, Japan), were proven to efficiently induce the number of selective cells and activate them to kill cancer cells [25,26,27,28,29,30,31,32].…”

Immune Cells Expansion From Peripheral Blood of Patients of Three Leading Cancers in Vietnam

“…The method for immune cell expansion we already published [22]. Briefly, peripheral blood mononuclear cells (PBMNCs) were obtained by density gradient centrifugation using Ficoll-Paque (GE Healthcare, Sweden) and were cultured using BINKIT® (Biotherapy Institute of Japan, Japan) at density of 1 × 10 6 cells/ml in cell initial Medium containing 0.01 KE/ml OK432 (Chugai Pharmaceutical, Japan), 700IU/ml recombinant human IL-2 (rhIL-2) (Chiron, Netherlands), supplemented with 5% of heat-inactivated autologous plasma, cultured in an anti-CD16 monoclonal antibody-immobilized culture flask for NK cell expansion.…”