Role of RPL39 in Metaplastic Breast Cancer

Abstract

Background: Metaplastic breast cancer is one of the most therapeutically challenging forms of breast cancer because of its highly heterogeneous and chemoresistant nature. We have previously demonstrated that ribosomal protein L39 (RPL39) and its gain-of-function mutation A14V have oncogenic activity in triple-negative breast cancer and this activity may be mediated through inducible nitric oxide synthase (iNOS). The function of RPL39 and A14V in other breast cancer subtypes is currently unknown. The objective of this study was to determine the role and mechanism of action of RPL39 in metaplastic breast cancer. Methods: Both competitive allele-specific and droplet digital polymerase chain reaction were used to determine the RPL39 A14V mutation rate in metaplastic breast cancer patient samples. The impact of RPL39 and iNOS expression on patient overall survival was estimated using the Kaplan-Meier method. Co-immunoprecipitation and immunoblot analyses were used for mechanistic evaluation of RPL39. Results: The RPL39 A14V mutation rate was 97.5% (39/40 tumor samples). High RPL39 (hazard ratio = 0.71, 95% confidence interval = 0.55 to 0.91, P = .006) and iNOS expression (P = .003) were associated with reduced patient overall survival. iNOS inhibition with the pan-NOS inhibitor NG-methyl-L-arginine acetate decreased in vitro proliferation and migration, in vivo tumor growth in both BCM-4664 and BCM-3807 patient-derived xenograft models (P = .04 and P = .02, respectively), and in vitro and in vivo chemoresistance. Mechanistically, RPL39 mediated its cancer-promoting actions through iNOS signaling, which was driven by the RNA editing enzyme adenosine deaminase acting on RNA 1. Conclusion: NOS inhibitors and RNA editing modulators may offer novel treatment options for metaplastic breast cancer.

Pharmacological Inhibition of NOS Activates ASK1/JNK Pathway Augmenting Docetaxel-Mediated Apoptosis in Triple-Negative Breast Cancer

“…Mechanistically relevant to these effects were the RPL39 A14V and MLF2 R158W mutations (8). Furthermore, in metaplastic breast cancer, a highly chemotherapy-resistant form of breast cancer, we determined that the RPL39 A14V mutation and iNOS expression are both associated with reduced patient overall survival (18). To test the efficacy of combining docetaxel and L-NMMA, we used the HM-3818 PDX, which was derived from an ascites sample from a patient with metastatic TNBC (Table 1).…”

“…Droplet digital PCR (ddPCR) was performed using a standard protocol with custom RPL39 (A14V) and MLF2 (R158W) ddPCR probes and primers (Bio-Rad Laboratories) as previously described (18).…”

“…These results suggest that iNOS may be the main NOS upregulated in TNBC, in response to docetaxel. L-NMMA has been previously used in TNBC cell lines to decrease iNOS production of total nitrites (16,31). We evaluated the effect of L-NMMA on docetaxel-induced iNOS.…”

“…S3A). We previously showed decrease in tumor volume growth by inhibition of iNOS with L-NMMA in PDX model BCM-4664 (18). We then evaluated the effect of these drugs by the addition of chemotherapy on PDX BCM-2147.…”

“…Breast cancer patients harboring RPL39 (A14V) and MLF2 (R158W) mutations demonstrate a shorter median time to relapse with lung metastases, compared with those without these mutations (17). In metaplastic breast cancer, a subtype mutation in RPL39 (A14V) has been correlated with higher levels of iNOS, increased metastatic relapse in the lung, and worse overall survival (18).…”