# Heterogeneous Activation of Signaling Pathways and Therapeutic Vulnerabilities in KSHV‐Associated Primary Effusion Lymphoma Cell Lines

## TL;DR

This study explores how different signaling pathways are activated in KSHV-related lymphoma and finds that combining inhibitors may be more effective for treatment.

## Contribution

The study reveals heterogeneous pathway activation and identifies dual PI3K/mTOR inhibitors as a promising therapeutic strategy for PEL.

## Key findings

- KSHV-infected PEL cell lines show constitutive AKT and mTORC1 activation and varied sensitivity to inhibitors.
- Dual PI3K/mTOR inhibitors like BGT226 and Dactolisib effectively inhibit proliferation and induce apoptosis across PEL cell lines.
- Responses to inhibitors do not always correlate with baseline pathway activation, indicating complex signaling networks in PEL.

## Abstract

Primary effusion lymphoma (PEL) is a rare and aggressive B‐cell malignancy caused by Kaposi's sarcoma‐associated herpesvirus (KSHV), with limited treatment options and poor prognosis. KSHV‐encoded proteins and miRNAs activate multiple signaling pathways that promote cell proliferation and survival. However, the heterogeneity in pathway activation and therapeutic responses among PEL cases remains poorly characterized. In this study, we investigated the activation status of key oncogenic and survival signaling pathways, including PI3K/AKT/mTOR, FOXOs and NF‐κB, and assessed the efficacy of targeted inhibitors in three KSHV‐positive EBV‐negative PEL cell lines BC3, BCP1 and BCBL1, KSHV‐negative BJAB cells, and KSHV‐infected BJAB‐KSHV cells. We observed heterogeneous activation of these pathways among PEL cell lines and differential sensitivity to pathway‐specific inhibitors. All KSHV‐infected cell lines exhibited constitutive AKT and mTORC1 activation and were sensitive to their respective inhibitors, though with varying efficacy. FOXO1 and FOXO3a, downstream targets of AKT, were frequently downregulated or inactivated by phosphorylation, consistent with AKT hyperactivation. Inhibition of FOXO1 suppressed proliferation and induced apoptosis in a cell line‐specific manner. Canonical and noncanonical NF‐κB pathways were differentially activated, and contributed to cell survival, as pathway‐specific inhibition suppressed proliferation. Interestingly, responses to inhibitors did not always correlate with basal pathway activation levels, highlighting the complexity of PEL signaling networks. Importantly, dual PI3K/mTOR inhibitors BGT226 and Dactolisib demonstrated superior efficacy by potently inhibiting proliferation and inducing apoptosis and cell cycle arrest in all PEL cell lines, suggesting an advantage in overcoming compensatory feedback mechanisms. These findings underscore the heterogeneity of PEL and the need for personalized therapeutic strategies. Our results support the potential of combinatorial or multi‐targeted approaches to improve treatment outcomes for PEL patients and warrant further preclinical and clinical investigations.