The NLP protein is a novel regulator of G2-m Phase of the cell cycle critical for proliferation of human peripheral T-cell lymphomas

Abstract

Abstract Peripheral T-cell lymphomas (PTCL) are a diverse group of aggressive T- and NK-cell malignancies distinguished by differences in cellular origin, genetic alterations, molecular signatures, morphology, and antigen expression. Representing roughly 15% of non-Hodgkin lymphomas, PTCLs carry a poor prognosis. Standard regimens such as CHOP or R-CHOP, along with approved targeted agents (belinostat, romidepsin, pralatrexate), have yielded limited long-term benefit, with 5-year survival rates of only 20–30%. Thus, meaningful advances in PTCL therapy will require the discovery of new molecular vulnerabilities important for lymphoma cell growth and survival. Using functional screening, we identified an uncharacterized human gene C17ORF58 (termed NLP - Netrin-Like Protein) to be critical for the survival of PTCL cells in vitro. NLP encodes a 37-kDa protein featuring an N-terminal signal peptide, nuclear localization signals, an intrinsically disordered region, and a C-terminal Netrin-like domain. Consistent with its predicted features, the protein localizes to the nucleus and is also secreted. Furthermore, structural modeling analysis predicts NLP possesses a druggable architecture. The gene is overexpressed in >95% of PTCL cases, including Anaplastic Large Cell Lymphoma, Angioimmunoblastic T-cell Lymphoma, and PTCL-Not Otherwise Specified (PTCL-NOS), as well as in multiple solid tumors such as breast, renal, and liver cancers. Importantly, loss of NLP in cancer cell lines induces G2–M arrest and mitotic catastrophe, culminating in apoptosis, suggesting a critical role in regulating malignant T-cell proliferation. The growth inhibition was associated with deregulation of genes involved in stress responses (PDCD5, NDUFAB1, RPL10), G2–M phase progression (TOM1, RPLP2, HMGA1, MYL12B), and cell death (PUMA, EPAS1, ATF5). Furthermore, NLP loss was accompanied by an increase in γH2A.X, Phospho-Chk2 (Thr68) and Phospho-BRCA1 (Ser1524), and the decrease in Phospho-Chk1 (Ser345) indicating the induction of double-strand DNA breaks and the activation of DNA repair mechanisms. Interestingly, NLP depletion did not suppress proliferation of normal human T-cells, mouse embryonic fibroblasts or human kidney embryonic cells. Homozygous NLP knockout mice are viable but show ~10% reduction in the body size suggesting that mouse NLP is important but not essential for normal development. Altogether, our findings identify NLP as a novel regulator of the G2–M phase of the cell cycle with a critical role in the proliferation of malignant T-cells. Given its predicted druggable structure, NLP represents a promising candidate for the development of novel targeted PTCL therapies.