Introduction
SARS-CoV-2 is a positive-sense, single stranded RNA in theCoronaviridae family of viruses1. While most cases of infection present with mild to moderate symptoms consisting of fever, fatigue, dry cough, headache, hypogeusia, anosmia, nausea and diarrhea2; roughly 15% of patients develop a severe disease phenotype requiring hospitalization, most commonly due to dyspnea and hypoxia3,4. Laboratory findings of severe infection include leukopenia, prolonged prothrombin time, and elevated serum concentrations of D-dimer, lactate dehydrogenase (LDH), ferritin, and c-reactive protein (CRP)5. Chest computed tomography classically demonstrates bilateral ground glass opacities1. Severe pathology associated with SARS-CoV-2 infection involves a hyperactive immune response to the virus resulting in a sudden, acute increase in pro-inflammatory cytokines, termed “the cytokine storm”6. Key pro-inflammatory cytokines upregulated in this process include interleukin 6 (IL-6) and tumor necrosis factor-alpha (TNF-α)6. Elevated cytokine levels prompt an influx of various immune cells into the site of infection, leading to tissue destruction, acute respiratory distress syndrome, septic shock and multiorgan failure7. Several mRNA and adenovirus vaccines targeting the spike protein have been approved8, but they appear to have reduced efficacy against the multiple higher infectivity and immune evading variants (e.g., spike protein mutations A222V, N501Y, or E484K/N501Y/K417N) that have arisen9,10. Thus, general SARS-CoV-2 therapies are needed. Early administration of monoclonal antibodies to the spike protein, RNA-dependent RNA polymerase inhibitor remdesivir and dexamethasone and late administration of enoxaparin may modify the natural history of the infection11-15, but additional treatments are needed. We previously reported the activity of a ruxolitinib plus interferon combination16. In this case report, we examine another combination that may show utility for SARS-CoV-2 infections.
Well differentiated small intestinal neuroendocrine tumors arise from serotonin-producing enterochromaffin cells and show epigenetic alterations frequently leading to PI3K/mTOR activation17. Upon metastases to liver and other sites, serotonin escapes liver metabolism, and the carcinoid syndrome occurs with flushing, diarrhea, and abdominal pain18. Treatments include sandostatin analogues, telotristat ethyl,177Lu-DOTATATE, everolimus, sunitinib, interferon-α, and surgical resection or ablation of metastases19. Metastatic small intestinal NET patients have a 70% 5-year survival20.
PI3K/mTOR pathway inhibitors have been proposed as a therapy to target viral protein synthesis and the hyperinflammation associated with SARS-CoV-221. mTOR is the serine/threonine protein kinase catalytic subunit of the mTORC1 complex with LST8, PRA540 and raptor. Targets of mTOR include phosphorylation inactivation of 4E-BP-1 and LARP-122,23. The result is release of eIF-4E translation initiation factor and stimulation of cap-dependent, 5’-terminal oligopyrimidine tract viral RNA translation. SARS-CoV-2 proteins N and nsp1 assist in this seminal virus infectivity step24,25. Inhibitors of mTOR block viral RNA synthesis23. SARS-CoV-2 hyper-inflammation is associated with the NLRP3 inflammasome containing the tripartite NLRP3 sensor protein, ASC adapter protein, and caspase-126. Infection produces a “priming” signal with viral RNA binding and signaling through Toll-like receptors or NOD-like receptors leading to NF-κB nuclear translocation, and NLRP3 and procaspase gene expression27. Then SARS-CoV-2 viroporins E, Orf3a and Orf8a cause cellular potassium efflux, NLRP3 oligomerization, ASC recruitment, and, finally, procaspase-1 recruitment and cleavage27. Orf8b aids the steps by binding the leucin-rich repeat domain of NLRP3 facilitating oligomerization. Gasdermin D is cleaved by caspase 1 and pyroptosis occurs along with inflammatory cytokine release. mTOR stimulates the NLRP3 inflammasome by increased mitochondrial reactive oxygen species formation and inhibiting autophagy28. As predicted, mTOR inhibitors inhibit NLRP3 inflammasome induced hyper-inflammation in vitro andin vivo 29,30.
Interferons signal through cell surface interferon receptors to JAK and STAT proteins and ultimately induce interferon-stimulated genes (ISGs) that halt viral translation via protein kinase receptor (PKR) and block viral NLRP3-induced inflammation via STAT131,32. SARS-CoV-2 proteins block interferon activity via multiple pathways. SARS-CoV-2 induced inflammasome caspases degrade interferon and interferon signaling polypeptides33. Over a dozen SARS CoV-2 proteins block early interferon induction or activity31. Late in the natural history, infection is associated with excessive inflammatory cytokinemia including interferons with dysfunctional T and NK cell responses34.
We present the case of a 61-years-old male found to be SARS-CoV-2 positive who was relatively asymptomatic while taking everolimus for co-existing metastatic neuroendocrine tumor but displayed a prolonged period of nasal swab PCR positivity. Administration of pegylated interferon was followed by prompt clearance of viral RNA by PCR. We hypothesize that the combination of everolimus with interferon may be useful in the acute COVID-19 setting to induce viral clearance with reduced risk of cytokine storm.