Case report: Understanding the impact of persistent tissue-localization of SARS-CoV-2 on immune response activity via spatial transcriptomic analysis of two cancer patients with COVID-19 co-morbidity.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected half a billion people, including vulnerable populations such as cancer patients. While increasing evidence supports the persistence of SARS-CoV-2 months after a negative nasopharyngeal swab test, the effects on long-term immune memory and cancer treatment are unclear. In this report, we examined post-COVID-19 tissue-localized immune responses in a hepatocellular carcinoma (HCC) patient and a colorectal cancer (CRC) patient. Using spatial whole-transcriptomic analysis, we demonstrated spatial profiles consistent with a lymphocyte-associated SARS-CoV-2 response (based on two public COVID-19 gene sets) in the tumors and adjacent normal tissues, despite intra-tumor heterogeneity. The use of RNAscope and multiplex immunohistochemistry revealed that the spatial localization of B cells was significantly associated with lymphocyte-associated SARS-CoV-2 responses within the spatial transcriptomic (ST) niches showing the highest levels of virus. Furthermore, single-cell RNA sequencing data obtained from previous (CRC) or new (HCC) ex vivo stimulation experiments showed that patient-specific SARS-CoV-2 memory B cells were the main contributors to this positive association. Finally, we evaluated the spatial associations between SARS-CoV-2-induced immunological effects and immunotherapy-related anti-tumor immune responses. Immuno-predictive scores (IMPRES) revealed consistent positive spatial correlations between T cells/cytotoxic lymphocytes and the predicted immune checkpoint blockade (ICB) response, particularly in the HCC tissues. However, the positive spatial correlation between B cells and IMPRES score was restricted to the high-virus ST niche. In addition, tumor immune dysfunction and exclusion (TIDE) analysis revealed marked T cell dysfunction and inflammation, alongside low T cell exclusion and M2 tumor-associated macrophage infiltration. Our results provide in situ evidence of SARS-CoV-2-generated persistent immunological memory, which could not only provide tissue protection against reinfection but may also modulate the tumor microenvironment, favoring ICB responsiveness. As the number of cancer patients with COVID-19 comorbidity continues to rise, improved understanding of the long-term immune response induced by SARS-CoV-2 and its impact on cancer treatment is much needed.

Reversible time-dependent inhibition of cytochrome P450 enzymes by duloxetine and inertness of its thiophene ring towards bioactivation.

Duloxetine is a selective serotonin-norepinephrine reuptake inhibitor (SNRI) approved to treat major depressive disorder and diabetic peripheral neuropathic pain. It is known to cause hepatotoxicity, in some cases leading to death. It has been reported that duloxetine causes time-dependent inhibition (TDI) of CYP1A2, CYP2B6, CYP2C19 and CYP3A4/5; but the nature of these TDI (whether reversible or irreversible) is not known. Irreversible TDI can cause clinically significant drug-drug interactions and also immune-mediated hepatotoxicity. Structurally, duloxetine possesses several toxicophores, i.e. the naphthyl and thiophene rings. It has been reported that the naphthyl ring undergoes epoxidation and was subsequently adducted to glutathione, but bioactivation related to the thiophene ring has not been completely elucidated. In this paper, the potential of duloxetine in causing irreversible TDI and generating reactive metabolites was investigated. Human liver microsomal assays demonstrated that duloxetine did not cause irreversible TDI of CYP1A2, CYP2B6, CYP2D6, CYP2C19 and CYP3A4/5. Subsequently, reactive metabolite trapping assays using soft nucleophiles (glutathione and glutathione ethyl ester) revealed a previously reported adduct at the naphthyl ring of duloxetine but not at the thiophene ring. Trapping assays utilizing a hard nucleophile (semicarbazide) did not demonstrate adducts with the thiophene ring, indicating an absence of thiophene ring opening. The hepatotoxicity of duloxetine is possibly not related to the irreversible TDI of CYP450 or the bioactivation of its thiophene moiety, but might be due to the epoxidation of its naphthyl ring.