Inhibition of transcription by dactinomycin reveals a new characteristic of immunogenic cell stress

Abstract

Chemotherapy still constitutes the standard of care for the treatment of most neoplastic diseases. Certain chemotherapeutics from the oncological armamentarium are able to trigger pre-mortem stress signals that lead to immunogenic cell death (ICD), thus inducing an antitumor immune response and mediating long-term tumor growth reduction. Here, we used an established model, built on artificial intelligence to identify, among a library of 50,000 compounds, anticancer agents that, based on their molecular descriptors, were predicted to induce ICD. This algorithm led us to the identification of dactinomycin (DACT, best known as actinomycin D), a highly potent cytotoxicant and ICD inducer that mediates immune-dependent anticancer effects in vivo. Since DACT is commonly used as an inhibitor of DNA to RNA transcription, we investigated whether other experimentally established or algorithm-selected, clinically employed ICD inducers would share this characteristic. As a common leitmotif, a panel of pharmacological ICD stimulators inhibited transcription and secondarily translation. These results establish the inhibition of RNA synthesis as an initial event for ICD induction.

An unexpected link between immunogenic cell death and inhibition of gene transcription

“…Intrigued by this observation, we used multiple tests to measure RNA synthesis (and downstream protein synthesis, downstream of RNA transcription) to conclude that most ICD inducers (including anthracyclines, oxaliplatin, lurbinectedin, crizotinib and thiostrepton) actually cause an inhibition of DNA-to-RNA transcription (and hence a subsequent inhibition of RNA-to-protein translation) and that this effect may cause a peculiar ER stress response consisting in the phosphorylation of eIF2α by eIF2α kinase 3 (EIF2AK3, better known as PERK) without any other signs of the unfolded stress response such as activation of the ATF6 and the IRE1-XBP1 axes. [8][9][10] The aforementioned results suggest that a vast class of ICD inducers (with the exception of microtubular poisons such as vinca alkaloids and taxanes) is able to reduce RNA synthesis, which resembles a response to viral infection. This has two major implications.…”

“…On one hand, it is possible to consider anticancer agents with known transcription-inhibitory properties as candidates for ICD induction. 9 On the other hand, it is relatively easy to measure inhibition of RNA synthesis in vitro, on cultured cells, for example using a chemically derivatized uridine analogue whose incorporation into nascent RNA, can be visualized by click chemistry to yield a fluorescent signal (Figure 1a). An alternative method to diagnose stalled transcription consists in detecting the separation of two proteins involved in RNA synthesis, nucleolin and fibrillarin, that normally (when RNA synthesis is active) colocalize in the nucleus, yielding an overlapping immunofluorescence staining (Figure 1b).…”

“…An alternative method to diagnose stalled transcription consists in detecting the separation of two proteins involved in RNA synthesis, nucleolin and fibrillarin, that normally (when RNA synthesis is active) colocalize in the nucleus, yielding an overlapping immunofluorescence staining (Figure 1b). 9 Hence, these types of tests might be easily added to the current compendium of assays to measure immunogenic stress events induced by anticancer agents (Figure 1c), adding yet another criterion to discriminate agents with a high potential of ICD induction from agents with a lower probability to kill cancer cells in an immunogenic fashion. Thus, this information could be fed into existing and yet-to-be-developed databanks to improve the algorithm calculating the 'ICD score', further refining the approach leading to the identification of ICD inducers.…”

Repurposing anticancer drugs for the management of COVID-19

“…This intravenously administered cytotoxic antibiotic acts by intercalation to DNA and interruption of RNA transcription ( Hollstein, 1974 ; LiverTox, 2012). Recently, dactinomycin was shown to induce immunogenic cell death, suggesting a direct antineoplastic activity enhanced by immunomodulatory properties ( Humeau et al 2020 ). In fact, this drug is used in combination with other cytotoxic drugs in the management of a broad array of solid malignancies, such as gestational trophoblastic disease, Wilms tumor and Ewing sarcoma ( Li et al 2018 ; Alazzam et al 2016 ; van den Heuvel-Eibrink et al 2017 ; Gaspar et al 2015 ).…”

Novel galenic formulations for delivering immunochemotherapies: kill intelligently, remove the checkpoint and let the T cells in!

“…2 Recently, the inhibition of DNA-to-RNA transcription by cytotoxic drugs was described as a cardinal mechanism governing ICD. 3 Several preclinical studies have shown that ICD induced by chemotherapy or radiotherapy sensitizes to subsequent immunotherapy. 4 Thus, ICD induction with oxaliplatin and cyclophosphamide sensitized non-small cell lung cancers (NSCLC) expressing oncogenic Kras and lacking Trp53 to anti-CTLA-4 and anti-PD-1-mediated double ICB.…”