CD38 Protects Tumor Cells from NK Lysis

Expression of the type II glycoprotein CD38 in hematological malignancies is abnormally high compared to that in normal cells of hematopoietic origin. Across all malignancies, the highest CD38 expression levels are detected on the surface of multiple myeloma (MM) cells. The high CD38 level has prompted the development of anti-CD38 antibody-based therapies, like daratumumab and isatuximab (SAR650984), that are promising therapeutics for the treatment of patients with MM. Cellular functions associated with CD38 include co-receptor mediated signaling, cell adhesion and ecto-enzymatic conversion of NAD to modulate calcium signaling and adenosine synthesis; but other than allowing antibody-mediated engagement of innate immunity, the role of CD38 in MM is not fully understood.

In order to investigate a potential immune-protective role for CD38, we evaluated a panel of 15MM and 6 diffuse large B-cell lymphoma (DLBCL) cell lines with varying levels of CD38 for their susceptibility to NK-mediated lysis. Briefly, calcein AM pulsed target cells were incubated with NK-92.CD16 NK cells (E:T=3:1) for 1 hr and cell lysis measured as a function of the fluorescent calcein released into the supernatant. In an initial experiment, we observed that the percentage of cell lysis for all lines with >100,000 CD38s per cell was low, below 10%. The cell lines with <100,000 CD38s per cell were more susceptible to NKs, with a median percentage cell lysis of around 25%. The observed levels of cell lysis were not correlated with the levels of released IFNγ or TNFα.

The differential lytic activity against the target cells led to the hypothesis that CD38 might have a role in protecting the tumor cells from NK-mediated lysis. To investigate this, we first knocked down CD38 in SUDHL-8 cells using shRNA, decreasing the levels of CD38 from 246,000 to 51,000 per cell. This resulted in an increase in the percentage of lysed cells from 25% to more than 50%, likely reflecting a loss of protection against the NK cells. We next overexpressed CD38 in U266 cells, increasing the CD38 levels from 9,000 to 327,000 per cell. As a result, the percentage of lysis decreased from more than 60% to around 30%, indicating that U266 cells are now more resistant to the lysis by NK cells. The protection that CD38 seems to confer to CD38-overexpressing U266 cells is completely lost by addition of isatuximab which triggers CD16-mediated lysis (antibody dependent cellular toxicity, ADCC).

Whether the protective function of CD38 described here is a direct or an indirect protection mechanism is currently under investigation.