Background: Multiple myeloma (MM) is a neoplasm of plasma cells. These cells express high levels of CD38 and are sensitive to daratumumab (DARA), a CD38-targeting monoclonal antibody. DARA kills MM cells via several mechanisms including antibody dependent cellular cytotoxicity (ADCC). Although DARA has improved the outcome of patients, CR is not achieved in all the cases. Besides, most patients that achieve CR will experience relapse. A potential explanation for blunted efficacy of DARA is downregulation of CD38 levels on MM cells and impaired ADCC due to depletion of CD38^high NK cells during therapy. ATRA upregulates CD38 levels on target cells and was proposed to improve the efficacy of DARA. NK cell supplementation was tested in a preclinical model, but that had only a modest effect on DARA efficacy, likely due to their short life via DARA-mediated elimination. Methods: We generated CD38 knockout (CD38^KO) NK cells from healthy donors via Cas9 RNPs and investigate if these cells boost DARA activity against MM. RNA-seq and cellular metabolic analysis were performed to examine the effect of CD38 deletion on NK cell function. Results: Knockout efficiency was 81.9 ± 6.9% (mean ± SD, N = 5). Very low off-target effects were seen by whole genome sequencing. When compared to paired CD38 wild type (CD38^WT) NK cells, CD38^KO NK cells showed lower conjugation (2.57% vs. 11.85%, N = 3, p = 0.04), less fratricide (97.3% vs. 54.2%, mean viability, N = 3, p = 0.01), and superior persistence in mice (18.16% vs. 0.42%, mean frequency in blood, N = 5, p < 0.01) in the presence of DARA. Additionally, CD38^KO NK cells exhibited enhanced ADCC against all tested MM cell lines and primary samples including CD38^low cell lines and primary MM cells from a patient with disease relapse on DARA. Transcriptomic and cellular metabolic analysis revealed that CD38^KO NK cells have favorable metabolic shift with higher mitochondrial respiratory capacity (N = 3, p < 0.01). Although ATRA upregulates CD38 levels on MM cell lines, we observed that ATRA treatment upregulates CD38 levels on circulating NK cells in patients. ATRA upregulated CD38 levels on CD38^WT NK cells, but not on CD38^KO NK cells, and increased their fratricide. Thus, CD38^WT NK cells showed impaired overall DARA-mediated cytotoxicity against MM cells in the presence of ATRA, even though those MM cells were sensitized to DARA. Conclusions: We present proof of concept that adoptive CD38^KO NK cell therapy has the potential to maximize the efficacy of DARA against MM. Further investigation into the role of metabolic reprogramming of CD38^KO NK cells is warranted.